Cheok 2014

Extraction and quantification of saponins: A Review
Choon Yoong Cheok, Hanaa Abdel Karim Salman, Rabiha Sulaiman
PII:
DOI:
Reference:
S0963-9969(14)00074-X
doi: 10.1016/j.foodres.2014.01.057
FRIN 5055
To appear in:
Food Research International
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Accepted date:
1 October 2013
19 January 2014
Please cite this article as: Cheok, C.Y., Salman, H.A.K. & Sulaiman, R., Extraction
and quantication of saponins: A Review, Food Research International (2014), doi:
10.1016/j.foodres.2014.01.057
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Extraction and quantification of saponins: A Review
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Choon Yoong Cheok, Hanaa Abdel Karim Salman, Rabiha Sulaiman*

Department of Food Technology
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Faculty of Food Science and Technology
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Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia
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* Corresponding author: Tel.: +60-3-89468520; Fax: +60-3-89423552;
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Email address: rabiha@upm.edu.my
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Abstract
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Saponins, a second metabolites mainly derived from plant materials, have been used
extensively in drug-related industry due to the pharmaceutical properties. These have driven the
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emergence of various new extraction technologies with the main purpose to optimize the yield in
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order to accommodate the recent need. The plants contain saponins is discussed, and its
pharmaceutical properties and applications in food are highlighted. This review focuses on the
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saponins extraction with emphasis on conventional and green technologies techniques employed
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in previous works by relating to their specific objective in each study. The quantification
methods of saponins yield, ie., spectrophotometric and chromatographic, are summarized and
discussed. In addition, this review aims to provide a point of reference to researchers who wish
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to design experiment to suit their particular objective in swift.
Keywords: Saponins; conventional extraction; green extraction technologies; quantification;

spectrophotometric; chromatographic.
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Contents
Introduction ........................................................................................................................................... 4
2.
Plant materials contain saponins ........................................................................................................... 6
3.
Pharmaceutical properties of saponins .................................................................................................. 7
4.
Applications of saponins in foods ....................................................................................................... 10
5.
Extraction techniques .......................................................................................................................... 12
7.
Conventional extraction techniques ............................................................................................ 14
5.2.
Green extraction technologies .................................................................................................... 16
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5.1.
Quantification of Saponins.................................................................................................................. 20
6.1.
Spectrophotometric method ........................................................................................................ 20
6.2.
Chromatograhic method ............................................................................................................. 23
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6.
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1.
Conclusions ......................................................................................................................................... 24
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References ................................................................................................................................................... 25
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1. Introduction
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Saponins are second metabolites which widely distributed in plant kingdom. It acts as a
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chemical barrier or shield in the plant defense system to counter pathogens and herbivores
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(Augustin, Kuzina, Anderson, & Bak, 2011). Therefore, it is found in plant tissues that are most
vulnerable to fungal or bacterial attack or insect predation (Wina, Muetzel, & Becker, 2005).
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Saponins divided into two major classes which are triterpenoid and steroid glycosides which
their structure characterization are varied by the numbers of sugar units attached at dierent
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positions (Hostettmann & Marston, 1995). The classification and occurrence of saponins in the
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plant kingdom is reviewed in detail by Vincken et al. (2007).
Saponins, which derived from soapwort (Saponaria officinalis L.), has been widely used for
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centuries as household detergent (Sparg, Light, & van Staden, 2004) due to its amphiphilic
nature with presence of a lipid-soluble aglycone and water-soluble chain(s) in their structure
(Gl-ntnda & Mazza, 2007). The seeds of Barringtonia asiatica Kurz (Lecythidaceae)
which have known to contain saponins, have been used traditionally by native Asian and Pacic
sherman as fish poison to enhance their catches (Sparg et al., 2004). Saponin-containing plant
materials, i.e., Yucca schidigera, alfafa, were used as feed additives to increase growth, milk or
wool in ruminant production (Wina et al., 2005). The molluscicidal saponins derived from
soapnut (Sapindus mukorossi Gaerth) has been found having inhibitory effects against golden
apple snail, which is the major pests of rice and other aquatic crops in Asian countries (Huang,
Liao, Kuo, Chang, & Wu, 2003).
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The discovery of biological activities of saponins is not only limited to the traditional uses,
but more recently, in pharmaceutical applications (Gl-ntnda & Mazza, 2007; Sparg et al.,
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2004). Saponins have been found having pharmaceutical properties of haemolytic, molluscicidal,
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anti-inflammatory, antifungal or antiyeast, antibacterial or antimicrobial, antiparasitic, antitumor,

and antiviral (Sparg et al., 2004). It employs as a starting point for the semi-synthesis of steroidal
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drugs in pharmaceutical industry. Sheng and Sun (2011) reviewed the clinical significance of
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triterpene saponins in prevention and treatment of metabolic and vascular disease.
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The pharmaceutical properties discoveries, especially anticancer, have intensified the seeking
of saponins from plant materials. These have driven the emergence of various new extraction
technologies with the main purpose of maximizing the yield in order to accommodate the recent
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need. Saponins are also known possessing mineral complexes of iron, zinc, and calcium (Milgate
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& Roberts, 1995). The beneficial effect of saponins intake in plasma cholesterol for human is
another important factor contributes to the continuous sorting of saponins (Milgate & Roberts,
1995). Besides anticancer (Cheng et al., 2011; Man, Gao, Zhang, Huang, & Liu, 2010; Waheed
et al., 2012), saponins have been discovered scientifically having pharmaceutical properties of
antioxidant (Chan, Khong, Iqbal, & Ismail, 2013; Dini, Tenore, & Dini, 2009; Li et al., 2010b),
immunological adjuvant activities (Estrada, Katselis, Laarveld, & Barl, 2000; Sun, Chen, Wang,
& Zhou, 2011; Verza et al., 2012), and haemolytic activities (Hassan et al., 2010; Sun et al.,
2011).
Since saponins are currently the most interested subject of their potential for industrial
processes and pharmacology, a correct selection of extraction technique through a review of
appropriate literature is essential. Researchers from a variety of scientific backgrounds are often
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challenged by the initial extraction process prior to isolation and identification of specific
saponins responsible for biological activities. The aim of this review is to summarize the
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selection of extraction methods from previous literature in respect to research focus in order to
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provide a quick reference in future experimental design.
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2. Plant materials contain saponins
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Saponins are mainly derived from various plant materials (Sparg et al., 2004; Vincken et al.,
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2007), but several of them are found in sea cucumber and starfish (Augustin et al., 2011;
Demeyer et al., 2014). The most widely studied plant material that found having saponins is
ginseng (Kwon, Blangar, Pare, & Yaylayan, 2003; Qian, Lu, Gao, & Li, 2009; Vongsangnak,
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Gua, Chauvatcharin, & Zhong, 2004; Wu, Lin, & Chau, 2001; Zhang & Cheng, 2006; Zhang,
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Liu, Qi, Li, & Wang, 2013b), even though saponins derived from alfafa has been carried out as
early by Van Atta et al. (1961). Other plant materials which have been discovered containing
saponins were soymilk (Lai, Hsieh, Huang, & Chou, 2013), sugar beet (Ridout, Price, Parkin,
Dijoux, & Lavaud, 1994), soy and chickpea (Serventi et al., 2013), asparagus (Vzquez-Castillo
et al., 2013), marion blackberry, strawberry, and plum fruit (Yoon & Wrolstad, 1984).
Saponins distribution has been found to vary in individual plant parts. For example, the roots
of Medicago truncatula (Huhman, Berhow, & Sumner, 2005) and Allium nigrum L. (Mostafa et
al., 2013) have been revealed containing the greatest total amount of saponins accumulation. The
yam tuber cortex has been discovered possess the highest amount of saponins of 582.53 g/g dw
which was about 2.55 times higher than tuber flesh of 227.86 g/g dw (Lin & Yang, 2008).
However, the total saponins concentration has been reported contain highest level in leaves from
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four varieties of Swithchgrass (Lee et al., 2009) and greenhouse grown Maesa lanceolata
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(Theunis et al., 2007). Table 1 tabulates the saponins derived from different plant parts.
Since saponins fall in two categories which on water-soluble sugar units attached to a lipophilic
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steroid (C27) or triterpenoid (C30) moiety (Challinor & De Voss, 2013; Gl-ntnda &
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Mazza, 2007; Harborne & Baxter, 1999), therefore, the isolation and structure elucidation of
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triterpenoid (Connolly & Hill, 2010) and steroidal (Challinor & De Voss, 2013) saponins have
been reviewed. Sparg et al. (2004) reviewed a list of plant species from which saponins have
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been isolated by categorizing them into triterpenoid and steroidal in period from 1998 to 2003.
However, recent review on the triterpenoid and steroidal saponins derived from various plants is
shown in Table 2. The elucidation and characterization of saponins structure are conducted on
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the basis of EI-MS (electrospray ionization-mass spectra), 1H and
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C NMR (nuclear magnetic
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resonance) data, such as in Ipomoea batatas (Dini et al., 2009), Aralia taibaiensis (Bi et al.,
2012), and Allium ampeloprasum var. porrum L. (Ado et al., 2011).
3. Pharmaceutical properties of saponins
Saponins rich in pharmaceutical properties and recently many studies focus on saponins
ability to increase immune responses (Estrada et al., 2000; Sun, 2006; Sun et al., 2011; Verza et
al., 2012), possess of antibacterial (Hassan et al., 2010; Mostafa et al., 2013; Iorrizzi, Lanzotti,
De Marino, Ranalli, & Zollo, 2002; Teshima et al., 2013), antioxidant (Bi et al., 2012; Chan et
al., 2013; Dini et al., 2009; Li et al., 2010b; Lin, Yang, & Lin, 2011), anticancer (Cheng et al.,
2011; Man et al., 2010), antidiabetic and anti-obesity (Joseph & Jini, 2013; Kimura, Ogawa,
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Katsube, Yokota, & Jisaka, 2008; Yun, 2010). Thus ginseng, which contains saponins, is
included in most of the Chinese Medicinal Prescriptions, for example, Bianxia Xiexin decoction
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in treating gastroenteritis diseases (Wang et al., 2014). Seven structurally consecutive saponins
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derived from Platycodon grandiflorum have been discovered having haemolytic activities and
adjuvant potentials on the immune responses to Newcastle disease virus-based combinant avian
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inuenza vaccine in mice (Sun et al., 2011). Both Verza et al. (2012) and Sun (2006) revealed
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that saponin fractions derived from Chenopodium quinoa seeds and Bupleurum chinense
enhanced haemolytic activities and adjuvant potentials on immune responses of mice against
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ovalbumin. Saponins obtained from Polygala senega L. were also suggested as potential vaccine
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adjuvants to increase specific immune responses (Estrada et al., 2000).
Hassan et al. (2010) reported that 100% methanol fraction of saponin-rich extracts from guar
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meal exhibited antibacterial activities against Staphylococcus aureus, Salmonella Typhimurium

and Escherichia coli, however the results showed 20% and 60% methanol fractions stimulated
Lactobacillus spp. growth. Aginoside saponins extracted from Allium nigrum L. roots had
significant antifungal activity (Mostafa et al., 2013). Saponins isolated from seeds of Capsicum
annum L. showed higher antimicrobial activity against yeasts compared to common fungi
(Iorrizzi et al., 2002). The n-butanol extract of shallot basal plates and roots exhibited antifungal
activity against plant pathogenic fungi (Teshima et al., 2013). Fruticoside I, a new steroidal
saponins derived from Cordyline fruticosa leaves, has been found showing moderate
antibacterial activity against the Gram-positive Enterococcus faecalis (Fouedjou et al., 2014).
A number of previous literature reported that saponins rich fraction have antioxidant
properties. There were derived from root bark of Aralia taibaiensis (Bi et al., 2012), deffated rice
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bran (Chan et al., 2013), Ipomoea batatas tubers (Dini et al., 2009), yellow horn (Li et al.,
2010b), stems and fruits of Momordica charantia (Lin et al., 2011). The sprouts of soybean,
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mung bean (Vigna radiata L.), and alfafa (Medicago sativa L.) which are rich in saponins, have
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been suggested as a good supplement of bioactive compound in daily diet with health-promoting
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antioxidant (Silva et al., 2013).
Man et al. (2010a) highlighted saponins possess significant anticancer properties and the
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structure-function of saponins influenced the antitumor mechanism. Saponins derived from
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Gynostemma pentaphyllum leaves (Cheng et al., 2011) and a novel steroidal saponin glycoside
derived from Fagonia indica (Waheed et al., 2012) have been discovered having
antiproliferation and apoptosis against prostate, breast and colon cancer cells. In a recent study,
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two new steroidal saponins, fruticoside H and fruticoside I, derived from Cordyline fruticosa
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leaves have been found having moderate cytotoxic activity against human breast, colon, and
melanoma cell lines (Fouedjou et al. 2014).
The lack of physical activity in daily routines and increase in high-calorie fast food intake
have led to a number of health-related consequences such as obesity and diabetes, where 80% of
type 2 diabetes patients are linked to obesity (Yang et al., 2010). Hence, studies on identification
of anti-obesity property from plant materials have become a popular trend. In a recent review,
saponins separated from plant materials of Panax japonicas, Platycodi radix, Kochia scoparia
fruits, Thea sinensis leaf, Scabiosa tschiliensis Grun., and Acanthopanax sessiliflorous were
suggested having anti-obesity property in inhibited pancreatic lipase (Yun, 2010). The possibility
of obesity treatment with saponins derived from leaves of Acanthopanax sessiliflorus
(Yoshizumi et al., 2006) and saponins from Japanese horse chestnut (Aesculus turbinate
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BLUME) after the treatment with wood ashes (Kimura et al., 2008) have been discussed. Panax
notoginseng saponins have both anti-hyperglycemic and anti-obese effects which may beneficial
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to type 2 diabetic patients by improving insulin sensitivity and decreasing leptin resistance (Yang
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et al., 2010). A typical triterpenoid saponin of charantin derived from Momordica charantia is a
well-known anti-diabetic bioactive compound (Joseph & Jini, 2013; Raman & Lau, 1996). Four
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new triterpenoid saponins isolated from the root bark of Aralia taibaiensis exhibited moderate
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effects on antioxidant and antiglycation activities which could be correlated with treatment of
diabetes mellitus (Bi et al., 2012). Both Liu et al. (2012b) and Zheng et al. (2012) demonstrated
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that total saponins from Rhizoma Anemarrhenae and Entada phaseoloides L. were able to
ameliorate diabetes-associated cognitive decline in rats. Saponin rich fractions from Bryonia
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Laciniosa (Patel, Patel, Vyas, Singh, Shah, & Gandhi, 2012a), Momordica charantia (Keller et
al., 2011) and Polygonatum odoratum (Deng et al., 2012) have been proven having anti-diabetic
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property and suggested to use in the treatment of diabetes.
Other therapeutic properties of saponins were reported in previous literature. There were
cardioprotective effects of saponins from Panax japonicas (He et al., 2012), anti-thrombotic
activity from Dioscorea zingiberensis (Li et al., 2010a), anti-inflammatory and antiulcerogenic properties from the bulbs of Allium ampeloprasum (Ado, da Silva, & Parente,
2011), anti-HIV activity from Momordica charantia (Chen et al., 2008; Chen et al., 2009), and
antiurolithiatic activity from fruit of Solanum xanthocarpum (Patel et al., 2012a).
4. Applications of saponins in foods

Apart from pharmaceutical applications, saponins have been used in foods as natural surfactant
and serve as preservative in controlling microbial spoilage of food. More recently, due to
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consumer preference for natural substance, Quillaja saponin has been used as a natural small
molecule surfactant in beverage emulsions in replacing synthetic surfactant of Tweens
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(Piorkowski & McClements, 2013). The effectiveness of the natural surfactant isolated from the
bark of the Quillaja saponaria Molina tree for forming and stabilizing emulsions with a synthetic
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surfactant (Tween 80) has been compared by Yang et al. (2013). After comparing the inuence
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of homogenization pressure, number of passes, and emulsier concentration on the particle size
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produced from these two surfactants, they suggested that the natural surfactant is an effective
surfactant that may be able to replace synthetic surfactants in food and beverage products. This
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natural surfactant has been further proven its stability and effectiveness at forming edible
Vitamin E delivery systems, thus it is recommended for functional food encapsulation and
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beverage applications (Yang & McClements, 2013).
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Due to its natural foam-like characteristic, the application of saponins as a natural bio-surfactant
to improve the surface properties of food is intensively studied recently. Wojciechowski et al
(2014) have conducted a study to evaluate the surface activity between Quillaja bark saponin
with -casein of bovine milk protein. From their results obtained, they suggested that the
Quillaja bark saponin can be used as a natural low molecular weight bio-surfactant. A recent
study indicated that banana cellulose micro and nano bres obtained by steam explosion process
which soaked with saponin, a surfactant extracted from soapnut fruit, showed differences in the
degree of modication and morphology of the cellulose bres (Cordeiro et al., 2013). The results
clearly show that the saponin can provide a continuous path of hydrogen bonds between the bre
surfaces which will thus enhance the hydrophobic and the acidbase nature of the bre surface.
This behaviour will lead to better polymer/bre interaction during the composite preparation.
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Andreuccetti et al. (2010) evaluated the incorporation of hydrophobic plasticizers in a matrix of
gelatin, using the saponin extracted from Yucca schidigera (yucca) as emulsifier, in the
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production of biodegradable emulsified films using the casting technique. Their results showed
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that the gelatin-based films produced have good mechanical resistance, low values of water
vapor permeability and reduced drying times, even though the films presented limited
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elongation, considerable solubility and opacity. Therefore, they suggested that the possibility of
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using this natural surfactant may allow for new applications of biodegradable emulsified films.
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The use of saponins as a natural biochemical substance in inactivation of food-borne viruses has
been reviewed by Li et al. (2013). The saponins-extract from Sapindus saponaria combined with
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heat-treatment was recommended to inactivate Alicyclobacillus acidoterrestris, a spoilagecausing bacterium, in orange juice (Alberice et al., 2013). Tea saponin, a tea seed-derived natural
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surfactant, combining with Bacillus amyloliquefaciens (Hao et al., 2011) and imazalil and
prochloraz (Hao et al., 2010), have been used for postharvest treatment of Mandarin fruit and
results showed that the incidence of green and blue mold and sour rot were reduced.
5. Extraction techniques
The recent advances in extraction of bioactive compound from plant material have been
intensively reviewed (Azmir et al., 2013; Wang & Weller, 2006) and this might due to the
increase in public awareness of preventative health care which could be promoted through the
consumption of plant material extract. In general, the extraction techniques employed in saponins
extraction can be classified into two categories, the conventional and the green technologies. The
conventional extraction techniques are maceration, Soxhlet, and reflux extraction, where the
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green technologies are ultrasound-assisted, microwave-assisted, and accelerated solvent
extraction (Heng, Tan, Yong, & Ong, 2013). The conventional extraction is relied on the
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solubility of solute from plant materials into solvent. Therefore, it often utililizes a large quantity
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of solvent to extract the desired solute, even though sometimes is aided with elevated
temperature by heating, and mechanical stirring or shaking. On the other hand, the green
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extraction techniques involved less hazardous chemical synthesis, safer chemicals used, energy
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efciency, use of renewable feedstock, and pollution prevention (Azmir et al., 2013). The design
of green extraction technologies are governed under these measurements. Consequently, water is
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used as extraction solvent by manipulating the extraction system pressure and temperature, as in
pressurized liquid extraction.
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Based on the importance of saponins as pharmaceutical properties especially in countering

cancer has provoked the invention of new extraction methods in order to obtain the maximum
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output to cope with the increasing demand. Therefore, a synthesis of previous literature in
extraction technique selection may provide useful information in related processing industry.
Fig. 1 clearly demonstrates that researchers are more inclined to selection of the conventional
extraction techniques (70%) which include the subsequent methods, compared to the green
technologies (30%), even though the green techniques use minimal solvent. The selection of
these methods usually was governed by the research focus of the studies being conducted. To
further analyze the selection of extraction technique made by researchers, Table 3 presents an
overview of extraction techniques in accordance to their research objectives. For isolation of new
saponins and pharmaceutical properties studies, 78% and 91% of the previous works were using
the conventional extraction methods. However, in works focused on quantification and
optimization studies, 58% and 67% of the previous works have selected green extraction
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technologies. It is also noteworthy that the ultrasound-assisted extraction is the most selected
green extraction technologies in quantification studies which gave an implication of its capability
Conventional extraction techniques
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5.1.
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and efficacy in obtaining significant saponins yields.
5.1.1. Maceration extraction
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The maceration extraction is a solid-liquid extraction where the bioactive compound
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(solute) inside the plant material is extracted by soaking the plant material in a specific solvent
for a period of time (Takeuchi et al., 2009). The efficacy of maceration process is determined by
two main factors, solubility and effective diffusion. The solubility is governed by basic rule of
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like dissolves like where indicated that a polar compounds dissolve in polar solvents, and
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nonpolar compounds dissolve in nonpolar solvents (Reichardt & Welton, 2011). The rate of
dissolution of a solute in the extraction solvent is determined by the rate of mass transfer of a
solute from the plant material to the solvent (Takeuchi et al., 2009). Due to the concentration
gradient in the solid-liquid interface, the transfer of the solute inside the plant material occurs
showing an effective diffusion takes place (Takeuchi et al., 2009).
No complicated utensil and equipment are needed for the set-up of a maceration extraction
system has made it a popular choice for researchers. The only paramount factor to be paid
attention in enhancing extractability is the knowledge of similarity of bioactive compound
interest and solvent polarity. Table 4 summarizes the maceration extraction procedure carried out
in previous literature in respect to their objective(s).
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Ethanol and methanol were the extraction solvents used to extract saponins from plant
material, and ethanol preferred better probably due to environment friendly concern. The
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duration of extraction time is long and sometimes takes up to weeks using this method, therefore,
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maceration extraction often aided with mechanical shaker (Cheng et al., 2011; Huhman et al.,
2005; Lee et al., 2009; Sylwia, Bogumil, & Wieslaw, 2006) or magnetic stirring (Verza et al.,
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2012) to shorten the extraction time.
5.1.2. Reflux and Soxhlet extractions
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Due to the similar working principle of Soxhlet and reflux extractions, the discussion is
carried out under the same sub-title. The only different between reflux and Soxhlet is that
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Soxhlet apparatus consists of a thimble to house the plant material. Reflux and Soxhlet extraction
involved distillation process which widely used in food and non-food industrial and laboratories.
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The process involves heating a solution to boiling and then returning the condensed vapours to
the original flask (Bart, 2011). The disadvantage of reflux and Soxhlet extractions is time
consuming where it required at least one hour for an extraction. Table 5 presents a summary of
saponins extraction from plant materials using reflux and Soxhlet extraction method. Ethanol is
still the most used solvent in reflux extraction, although there are few used methanol as
extraction solvent. The extraction duration of reflux extraction was varied from 1 to 4 hours,
while for Soxhlet was 24 to 72 hours.
5.1.3. Subsequent extraction

There were numerous studies carried out on extraction of saponins using two extraction
methods subsequently. The purpose of using two extraction methods subsequently might due
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highly purify the extract before subjected to HPLC analysis for isolation and identification
saponins compound from plant material. Table 6 presents the application of two subsequent
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extraction methods in obtaining the specific saponins from various plant materials. For Soxhlet
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and reflux subsequent method, the Soxhlet extraction is carried out first to remove the lipid of
the plant material using solvent such as chloroform (Bialy, Jurzysta, Mella, & Tava, 2004; Bialy
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Jurzysta, Mella, & Tava, 2006; Oleszek et al., 2001; Tava et al., 2009) and hexane (Ncube,
Green extraction technologies
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5.2.
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Ngunge, Finnie, & Staden, 2011).
5.2.1. Ultrasound-assisted extraction (UAE)
The phenomenon of ultrasound in creating cavitation bubbles in the solvent by acting as a
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microjet to denature the plant cell wall when the bubbles collapse at rarefraction resulted in a
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greater extraction yield of biaoactive compounds. Few researchers have reviewed the ultrasound
effect on the technological properties and bioactivity of food (Soria & Villamiel, 2010), the
applications of ultrasound-assisted extraction on bioactive principles from herbs (Vinatoru,
2001), food industry and processing (Mason, 1998; Vilkhu, Mawson, Simons, & Bates, 2008).
Although ultrasound-assisted extraction is commonly employed in many bioactive compounds
extraction (Cheok, Chin, Yusof, Taib, & Law, 2013; de Koning, Janssen, & Brinkman, 2009;
Jadhav, Rekha, Gogate, & Rathod, 2009; Zhang et al., 2008), only few has been found in
saponins extraction (Table 7).
5.2.2. Microwave-assisted extraction (MAE)
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Microwaves are non-ionizing electromagnetic waves with a frequency range from 0.3 to
300 GHz (Heng et al., 2013; Takeuchi et al., 2009). Recently, MAE has drawn attention in
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bioactive compound extraction from plant material due to short extraction time, minimal solvent
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usage, and its special heating mechanism (Heng et al., 2013). The recent applications of MAE of
plant secondary metabolites such as flavonoids, quinones, phenylpropanoids, terpenoids,
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alkaloids and saponins have been reviewed (Zhang, Yang, & Wang, 2011). Microwaves are able
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to penetrate into biomaterials and generate heat by interacting with polar molecules such as water
inside the materials. The penetration depth of microwaves into plant matrix depends on dielectric
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constant, moisture content, temperature, and the frequency of the electrical field (Takeuchi et al.,
2009). The water contained in a plant material is responsible for the absorption of microwave
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energy which led to internal superheating and cell structure disruption, and consequently,
facilitates the diffusion of bioactive compound from the plant matrix (Takeuchi et al., 2009). The
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efficacy of MAE is relied on the effect of microwave on extraction solvent and plant matrix cell
structure (Takeuchi et al., 2009).
Although the potential application of microwave extraction for flavonoids has been reviewed
thoroughly (Routray & Orsat, 2011), only few works of saponins extraction using MAE has been
mentioned in previous reviews (Zhang et al., 2011; Gl-ntnda & Mazza, 2007). Table 8
synthesizes an up-to-date literature of using MAE in saponins extraction in which the content
was not covered in those two reviews. The superiority of MAE in saponins extraction in
comparison with other extraction methods, in terms of higher yield (Chen, Xia, & Gong, 2007b;
Li et al., 2010b; Mandal & Mandal, 2010; Xu et al., 2012) and shorter extraction time (Chen et
al., 2007b; Kwon et al., 2003; Mandal & Mandal, 2010; Xu et al., 2012), has been found in
previous literature.
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5.2.3. Accelerated solvent extraction (ASE)
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Accelerated solvent extraction has been regarded as a green technique in plant material
sample preparation prior to chromatographic analysis (Heng et al., 2013; Azmir et al., 2013).
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This technique was introduced by Dionex Corporation in 1995. It is also known as pressurized
liquid extraction, pressurized solvent extraction, and enhanced solvent extraction. Sometime it is
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referred as pressurized hot water extraction, sub-critical water extraction or superheated water
extraction, when water is used as solvent (Mustafa & Turner, 2011). It is an automated rapid
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extraction technique that uses minimal solvent at elevated temperature and pressure. The merit of
using increased temperature is to enhance the solubility and mass transfer of solute to solvent,
and elevated pressure keeps the solvent below its boiling point, enabling fast, safe, and efcient
extraction of target analytes from plant materials into the extraction solvent (Mottaleb & Sarker,
2012). An extraction process is usually completed in 15-25 minutes using only 15-45 ml
consumption of solvent. Therefore, it has been widely applied in the fields of environmental,
food, polymer, and pharmaceutical researches.
ASE comprises of two main set-ups, there are the static and dynamic instruments
(Mustafa & Turner, 2011). Static setup is replacement of the solvent between cycles if the
extraction process consists of one or several extraction cycles. A high pressure pump is required
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to pump the extraction solvent through the sample vessel continuously in the dynamic setup. The
parameters affecting ASE efficiency are temperature, pressure, type and composition of solvents,
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modifiers and additives, matrix composition, and extraction mode (Sun, Ge, Lv, & Wang, 2012).
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The most commonly applied operating temperature and pressure for ASE are 100 C at 1500 psi
(Mottaleb & Sarker, 2012). Although ASE is a green technology in extracting bioactive
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compound from plant material, the application in saponins extraction is still scarce. Table 9
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summarizes extraction of saponins using ASE from plant materials. Worth noted that this method
is used mostly to extract ginsenoside from ginseng (Qian, Lu, Gao, & Li, 2009; Wan, Zhang, Ye,
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& Wang, 2008; Wan et al., 2006; Zhang, Liu, Qi, Li, & Wang, 2013) could be due to the
precious value of the product.
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The efficacy of ASE in saponins extraction has been studied and compared with other
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extraction methods. A higher saponin yield has been obtained from cow cockle seeds using
accelerated solvent extraction compared to ultrasonic-assisted extraction in pure and aqueous
solvents of ethanol and methanol (Gl-ntnda, Balsevich, & Mazza, 2007). Similarly, the
pressurized hot water system extracted a greater yield of ginsenosides (11.2 mg/g) compared to
ultrasound-assisted method (7.2 mg/g) from Panax quinquefolium (Engelberth et al., 2010).
Although slight increase in saponins yields of escin Ia, escin Ib, isoescin Ia and isoescin Ib were
obtained in extraction from Aesculus chinensis Bunge using ASE, it required shorter extraction
time of 7 minutes compared to reflux and sonication extraction of 1 hour and 30 minutes,
respectively (Chen et al., 2007a). Pressurised liquid extraction showed distinctive advantages of
yielding total amount of saponins of 7.36% over other green extraction methods of ultrasound of
5.77%, and conventional extractions of Soxhlet of 6.99% and maceration of 6.00%, in the
extraction of saponins from Panax notoginseng (Wan et al., 2006).
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6.
Quantification of Saponins
Prior to the quantification of total saponins of a plant source, it is appropriate to carry out a
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simple procedure to test the presence of saponins. This can be done by putting the plant material
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into a test tube filled with distilled water and vigorously shaken for 2 minutes (Ncube et al.,
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2011). The appearance of stable and persistent foam on the liquid surface for 15 minutes
indicated the presence of saponins. The quantification of plant saponins is usually carried out by
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spectrophotometric and chromatographic methods. The difference quantitative expression

between the two methods is the spectrophotometrics gives a total saponins value while the
Spectrophotometric method
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6.1.
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chromatographics quantifies specific saponins compound.
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The spectrophotometric technique has become a popular method in the quantification of

saponins from plant materials could be because it is simple, fast and inexpensive to operate.
Total saponins also known as vanillin-sulphuric acid assay, is the most commonly selected
spectrophotometric method in plant saponins quantification (Table 10). However few factors,
such as selection of standards, wavelength, and others should be considered before using this
method. The basic principle of this method is the reaction of oxidized triterpene saponins with
vanillin (Li et al., 2010b). Sulphuric acid is used as oxidant and the distinctive colour of this
reaction is purple (Hiai, Oura, & Hakajima, 1976) and sometimes perchloric acid is used (Chen
et al., 2007b; Li et al., 2010b; Wu et al., 2001). Due to differences in selection of reagent,
condition to allow full colour development, standard, and wavelength from previous researches
as presented in Table 10, it is hard to compare the results in terms of yields. However, it provides
a good reference for future experimental design.
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Two works (Patel et al., 2012a and Ncube et al., 2011) were found using the conditions of
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60 C and 10 minutes to allow the mixture to have full colour development following procedure
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in Hiai et al. (1976). However, other researchers used 70 C for 15 minutes (Chen et al., 2007b)
and 20 minutes (Li et al., 2010b) to allow full colour development. This inconsistency should be
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standardized because the reaction time may directly attribute to the final absorbance value which
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later translates into quantity. As presented in Table 10, the standards of oleanolic acid,
soyasaponin, Quillaja saponin, and ginsenoside are grouped in triterpenoid saponins, where
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diosgenin and sarsasapogenin are categorized in steroid saponins (Harborne & Baxter, 1999).
Since total saponins method is to quantify total saponins from the reaction of oxidized triterpene
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saponins with vanillin (Li et al., 2010b), these inevitably raise a question whether the selection of
standard to be used in spectophotometeter is essential to express the correct saponins group from
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the plant source. Unfortunately, the related information on selection of the standards is rarely
found. No explanation is stated in previous works on the selection of wavelength, but most
researchers selected wavelength of 544 nm is observed (Table 10). However, the selected
wavelengths are falled within the range of 480-610 nm, except 473 nm (Mostafa et al., 2013) and
283 nm (Liu et al., 2012b), most probably due to the maximum absorption of purple colour falls
within this range (Bruice, 2007).
Despite total saponins, total steroidal sapogenin is employed to quantify specifically the
steroidal saponins content of plant material (Baccou, Lambert, & Sauvaire, 1977). This method
is also based on colour reactions with anisaldehyde (or vanillin), sulphuric acid and ethyl acetate
measured at maximum wavelength (max) of 430 nm (Baccou et al., 1977). The difference
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between total saponin and total steroidal sapogenins is the solvent used to prepare the reagents.
For total saponins, vanillin reagent is prepared by diluting with ethanol and sulphuric acid with
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water, whereas for total steroidal sapogenins, both the anasaldehyde and sulphuric acid are
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diluted with ethyl acetate. Total steroidal sapogenins has been proven stable and reproducible
with a number of standards, ie., diosgenin, tigogenin, hecogenin, smilagenin, yonagenin,
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tokorogenin, etc. without interference from sugars, sterols, fatty acid and vegetable oil (Baccou
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et al., 1977).
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The procedure to quantify total steroidal sapogenin is by weighing 0-40 g of crude

extract and dissolved in 2 ml of ethyl acetate in the test tube. Then mixed with 1 ml of reagent A
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(consists of 0.5 ml anisaldehyde and 99.5 ml ethyl acetate) and 1 ml of reagent C (consists of 50
ml concentrated sulphuric acid and 50 ml ethyl acetate). The test tube with mixtures was placed
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in a water-bath at 60 C for 10 minutes to allow full colour development. Then, it was cooled for
10 minutes at room temperature before measuring at 430 nm using a spectrophotometer. It has
been employed in recent researches to quantify total steroidal saponins from micropropagated
Tulbaghia violacea which obtained 10.03 mg DE/ml (Ncube et al., 2011), and the extract from
defatted rice bran in n-butanol fraction which yielded 5.70 mg DE/g (Chan et al., 2013). Qin et
al. (2009) carried out the total steroidal sapogenin determination of Dioscorea zingiberensis with
some modifications. They used perchloric acid instead of sulphuric acid. The test tube was
placed in a water bath maintained at 70 C for 15 min to develop colour fully, then allowed to
cool for 2 min in 0 C water bath and metered volume to 25 ml with glacial acetic acid. After 30
minutes of stabilization, only then the test tube was subjected to measure its absorbance at 454
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nm using a spectrophotometer and yielded total steroidal saponins of 28.34% (w/w) of
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lyophilized powder.
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Haemolytic method is another spectrophotometric method uses to quantify saponins

content of a plant material (Barve, Laddha, & Jayakumar, 2010). The principle of this method is
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the reaction of saponins with blood reagent to release oxy-hemoglobin which results a
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measurable colour for spectrophotometer. The saponins concentrations in bittergourd varieties

were quantified using haemolytic method (Habicht et al., 2011). The saponin extract was
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dissolved in distilled water and 100 l of this solution were incubated with 1 ml fresh EDTAblood at 30 C for 30 min. After centrifugation for 10 min, haemoglobin was quantied in the
supernatant photometrically at 545 nm and the result was expressed in haemolytic saponins.
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They revealed that white bitter gourd varieties were found to contain signicantly lower saponin
6.2.
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concentrations (0.25%) compared to green varieties (0.67%).
Chromatograhic method
Saponins are separated and purified from plant materials using chromatographic methods in
many studies to identify a specific saponins compound (Ado et al., 2011; Liu et al., 2012a) and
investigate its pharmaceutical property (Gupta et al., 2010; He et al., 2012; Zheng et al., 2012).
The most common chromatographic methods employed are high performance liquid
chromatography (HPLC) (Bi et al., 2012; He et al., 2012; Liu et al., 2012b; Mostafa et al., 2013)
and thin layer chromatography (TLC) (Ado et al., 2011; Liu et al., 2012a; Patel et al., 2012a).
The chromatographic determination of plant saponins for period from 2002 to 2005 has been
reviewed by Oleszek and Bialy (2006). Therefore, the present review looks into the most recent
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works of chromatographic method specifically in quantitation study of plant saponins where
HPLC was found to be the most commonly used method. A summary of quantification of plant
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saponins using HPLC is presented in Table 11. As noted the quantification of specific saponin
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compound is the primary objective of all the studies which apply HPLC method. The specific
saponins content detected not only serve as a good data reference source to future researchers,
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but as a strong scientific reference to drug-related manufacturer who is interested to process the
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particular plant source further. Besides HPLC, ultra pressure liquid chromatography (UPLC)
(Foubert et al., 2010; Ha et al., 2014; Serventi et al., 2013; Verza et al., 2012) was also employed
Conclusions
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7.
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to quantify saponins.
Saponins are important secondary metabolites derived from various plant sources because
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of its invaluable pharmaceutical properties. This review focuses on two different extraction
techniques (conventional and green technology) employed in previous works to obtain crude
saponins extract prior to further analysis. Moreover, spectrophotometric and chromatographic
methods in saponins quantification are described. This synthesis of the range and diversity of
previous studies already active in the field serves as important information to researchers who
wish to embark a new project. This may provide an overview and quick reference for future labscale experimental design. The knowledge of extraction technique employed in respect to
objective is vital and can be extended to address the rising food processing challenges over time.
After highlighting the lack of green extraction technologies utilization in lab-scale saponins
extraction, more attention should be paid by researchers for these technologies exploration in
future.
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Xu, H., Ji, X., Shi, X., Du, Y., Zhu, H., & Zhang, L. (2011). Development of a novel method for
TE
triterpenoidal saponins in rat plasma by solid-phase extraction and high-performance liquid

chromatography tandem mass spectrometry. Analytical Biochemistry, 419, 323-332.Xu, H-
AC
CE
P
j., Shi, X-w., Ji, X., Du, Y-f., Zhu, H., & Zhang, L-t. (2012). A rapid method for
simultaneous determination of triterpenoid saponins in Pulsatilla turczaninovii using
microwave-assisted extraction and high performance liquid chromatographytandem mass
spectrometry. Food Chemistry, 135, 251-258.
Yang, C-y., Wang, J., Zhao, Y., Shen, L., Jiang, X., Liang, N., Zhang, L., Chen, Z-h., & Xie, Zg. (2010). Anti-diabetic effects of Panax notoginseng saponins and its major antihyperglycemic components. Journal of Ethnopharmacology, 130, 231236.
Yang, D-J., Lu, T-J., & Hwang, LS. (2003). Isolation and identification of steroidal saponins in
Taiwanese yam cultivar (Dioscorea pseudojaponica Yamamoto). Journal of Agricultural
and Food Chemistry, 51, 6438-6444.
49
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Yang, Y., & McClements, D.J. (2013). Encapsulation of vitamin E in edible emulsions fabricated
PT
using a natural surfactant. Food Hydrocolloids, 30, 712-720.
Yang, Y., Leser, M.E., Sher, A.A., & McClements, D.J. (2013). Formation and stability of
RI
emulsions using a natural small molecule surfactant: Quillaja saponin (Q-Naturale). Food
SC
Hydrocolloids, 30, 589-596.
NU
Yoon, K.R., & Wrolstad, R.E. (1984). Investigation of marion blackberry, strawberry, and plum
fruit for the presence of saponins. Journal of Agricultural and Food Chemistry, 32, 691-
MA
693.
Yoshizumi, K., Hirano, K., Ando, H., Hirai, Y., Ida, Y., Tsuji, T., Tanaka, T., Satouchi, K., &
TE
Terao, J. (2006). Lupane-type saponins from leaves of acanthopanax sessiliflorus and their
335-341.
AC
CE
P
inhibitory activity on pancreatic lipase. Journal of Agricultural and Food Chemistry, 54,
Yun, J.W. (2010). Possible anti-obesity therapeutics from nature A review. Phytochemistry, 71,
16251641.
Zhang, H., & Cheng, Y. (2006). Solid-phase extraction and liquid chromatographyelectrospray
mass spectrometric analysis of saponins in a Chinese patent medicine of formulated Salvia
miltiorrhizae and Panax notoginseng. Journal of Pharmaceutical and Biomedical Analysis,
40, 429432.
Zhang, H-F., Yang, X-H., & Wang, Y. (2011). Microwave assisted extraction of secondary
metabolites from plants: Current status and future directions. Trends in Food Science &
Technology, 22, 672-688.
50
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Zhang, T., Liu, H., Liu, X-T., Xu, De-r., Chen, X-q., & Wang, Q. (2010). Qualitative and
PT
quantitative analysis of steroidal saponins in crude extracts from Paris polyphylla var.
RI
yunnanensis and P. polyphylla var. chinensis by high performance liquid chromatography

coupled with mass spectrometry. Journal of Pharmaceutical and Biomedical Analysis, 51,
NU
SC
114-124.
Zhang, X., Ito, Y., Liang, J., Su, Q., Zhang, Y., Liu, J., & Sun, W. (2013a). Preparative isolation
MA
and purication of ve steroid saponins from Dioscorea zingiberensis C.H.Wright by

counter-current chromatography coupled with evaporative light scattering detector.
TE
Journal of Pharmaceutical and Biomedical Analysis, 84, 117-123.
AC
CE
P
Zhang, Y., Liu, C., Qi, Y., Li, S., & Wang, J. (2013b). Application of accelerated solvent
extraction coupled with counter-current chromatography to extraction and online isolation
of saponins with a broad range of polarity from Panax notoginseng. Separation and
Purification Technology, 106, 82-89.
Zhang, Z-S., Wang, L-J., Li, D., Jiao, S-S., Chen, X.D., & Mao, Z-H. (2008). Ultrasoundassisted extraction of oil from flaxseed. Separation and Purification Technology, 62, 192198.
Zhao, Y., Wang, X., Wang, H., Liu, T., & Xin, Z. (2014). Two new noroleanane-type triterpene
saponins from the methanol extract of Salicornia herbacea. Food Chemistry, 151, 101-109.
51
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Zheng, T., Shu, G., Yang, Z., Mo, S., Zhao, Y., & Mei, Z. (2012). Antidiabetic effect of total
saponins from Entada phaseoloides (L.) Merr. in type 2 diabetic rats. Journal of
PT
Ethnopharmacology, 139, 814821.
RI
Zhu, N., Sheng, S., Sang, S., Jhoo, J-W., Bai, N., Karwe, M.V., Rosen, R.T., & Ho, C-T. (2002).
List of Figure
TE
MA
NU
SC
Triterpene saponins from debittered quinoa (Chenopodium quinoa) seeds. Journal of
AC
CE
P
Fig. 1. Current extraction techniques employed in extraction of saponins from plant materials
52
PT
ACCEPTED MANUSCRIPT
RI
Maceration (36%)
NU
SC
Ultrasound-assisted (14%)
TE
MA
Reflux (22%)
Green extraction technologies
AC
CE
P
Conventional extractions
Fig. 1. Current extraction techniques employed in extraction of saponins from plant materials
53
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Table 1. Saponins derived from different parts of plant materials
PT
List of Tables
RI
Table 2. Triterpenoid and steroidal saponins derived from various plant materials
SC
Table 3. Selection of saponins extraction technique based on research focus

Table 4. Summary of studies focusing on saponins extraction
NU
Table 5. Summary of studies focusing on saponins extraction using reflux and Soxhlet extraction
MA
Table 6. Summary of studies focusing on saponins extraction using two subsequent extractions
Table 7. Summary of studies focusing on saponins extraction using ultrasound-assisted
extraction
TE
Table 8. Summary of studies focusing on saponins extraction using microwave-assisted

extraction
AC
CE
P
Table 9. Summary of studies focusing on saponins extraction using accelerated solvent

extraction
Table 10. Spectrophotometric method in quantifying total saponins from various plant materials
Table 11. Quantification of plant saponins using HPLC
54
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Table 1. Saponins derived from different parts of plant materials

Plant material
Reference(s)
Seed
Aesculus chinensis Bunge

Aesculus turbinate BLUME
Allium tuberosum
Argania spinosa
Bryonia Laciniosa
Capsicum annum L.
Chenopodium pallidicaule
Chenopodium quinoa
Chen et al. (2007a)

Kimura et al. (2008)
Sang et al. (2001a); Sang et al. (2001b)
Alaoui et al. (2002)
Patel et al. (2012b)
Iorizzi et al. 2002
Rastrelli et al. (1996)
Dini et al. (2001); Verza et al. (2012); Zhu et al. (2002); Woldemichael
& Wink (2001)
Zheng et al. (2012)
Ha et al. (2014)
RI
SC
NU
MA
Petit et al. (1995)
Gl-ntnda et al. (2007)
Mostafa et al. (2013)

Bi et al. (2012)
Hu et al. (2008)
Avula et al. (2011)
Borges et al. (2013)
Wang et al. (2012)
AC
CE
P
TE
Root
Trigonella foenum graecum

L.
Vaccaria segetalis Garcke,
Saponaria Vaccaria L.,
Vaccaria pyramidate
Allium nigrum L.
Aralia taibaiensis
Bupleurum chinense
Caulophyllum thalictroides
Chiococca alba
Dioscorea panthaica
Entada phaseoloides
Leguminous species
PT
Plant part
Leaf
Glycyrrhiza inflate,
Glycyrhiza glabra,
Glycyrrhiza uralensis
Glycyrrhiza yunnanensis
Gypsophila trichotoma
Maesa lanceolata
Medicago hybrid
Momordica charantia
Paris polyphylla var.
chinensis
Paris polyphylla var.
yunnanensis
Pulsatilla chinensis
Tao et al. (2013)
Panax notoginseng
Panax quinquefolius
Platycodon grandiorum
Polygonatum odoratum
Radix Astragali
Vigna radiata L.
Yucca gloriosa L.
Lau et al. (2003); Li et al. (2005); Wu et al. (2001)

Engelberth et al. (2010); Gafner et al. (2004); Wu et al. (2001)
Sun et al. (2011)
Bai et al. (2014); Deng et al. (2012)
Qi et al. (2006)
Waller et al. (1999)
Skhirtlardze et al. (2011)
Acanthopanax sessiliflorus
Allium nigrum L.
Yoshizumi et al. (2006)

Ji et al. (2014)
Voutquenne-Nazabadioko et al. (2013)
Theunis et al. (2007)
Bialy et al. (2006)
Chen et al. (2008)
Liu et al. (2013); Zhang et al. (2010)
Zhang et al. (2010)
Xu et al. (2011)
55
ACCEPTED MANUSCRIPT
Tuber
Flower
NU
PT
RI
SC
Akhov et al. (1999)

Rubio-Moraga et al. (2011)
Lian & Zhang (2013)
Li et al. (2007); Lin et al. (2011); Liu et al. (2012a)
Patel et al. (2012a)
Dinchev et al. (2008)
Alabdul Magid et al. (2006)
Lin et al. (2011)
Masullo et al. (2014)
Huang et al. (2003)
Kowalczyk et al. (2011)
Voutquenne et al. (2005)
Dini et al. (2009)
Prez et al. (2013)
MA
Pericarp
Bark

Guo et al. (2011)
Stem
Tribulus terrestris L.
Vigna radiata L.
Ziziphus jujube, Z. jujuba
var. spinosa
Allium nutans L.
Allium nigrum L.
Crocus sativus
Gleditsia sinensis Lam.
Momordica charantia
Solanumxanthocarpum
Caryocar villosum
Momordica charantia
Silphium asteriscus L.
Vigna radiata L.
Sapindus mukorossi
Yucca schidigera Roezl
Harpullia austro-caledonica
Ipomoea batatas
Agave offoyana
TE
Fruit

Eskander et al. (2011)
Fouedjou et al. (2014)
Mandal & Mandal (2010)
Foubert et al (2010); Theunis et al. (2007)
Masullo et al. (2014)
AC
CE
P
Bulb
Antonia ovate
Beaucarnea recurvata
Cordyline fruticosa
Gymnema sylvestre
Maesa lanceolata
Silphium asteriscus L.
56
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Table 2. Triterpenoid and steroidal saponins derived from various plant materials
Reference(s)
PT
Plant material
Triterpenoid saponins
Antonia ovata
Aralia taibaiensis
Bacopa monnieri
Caryocar villosum
Momordica charantia
medicago truncatula
medicago arabica L.
medicago hydriba
Caulophyllum thalictroides
Chenopodium quinoa
Chiococca alba
Flos Lonicerae
Genista ulicina Spach
MA
NU
SC
RI

Bi et al. (2012)
Ganzera et al. (2004)
Chen et al. (2008); Li et al. (2007)
Kapusta et al. (2005); Huhman et al. (2005)
Bialy et al. (2004); Tava et al. (2009)
Bialy et al. (2006)
Avula et al. (2011)
Verza et al. (2012)
Borges et al. (2013)
Chai et al. (2005)
Boutaghane et al. (2013)
AC
CE
P
TE
Glycyrrhiza inflate, Glycyrhiza glabra,

Glycyrrhiza uralensis
Glycyrrhiza yunnanensis
Gymnema sylvestre
Gypsophila trichotoma
Harpullia austro-caledonica
Lamii albi flos
Pulsatilla chinensis
Salicornia herbacea
Panax quinquefolius
Ganoderma atrum
Ipomoea batatas
Xanthoceras sorbifolia Bunge
Crocus sativus
Maesa lanceolata
Polygala japonica
Pulsatilla turczaninovii
Acanthopanax sessiliflorus
Steroidal saponins
Agave offoyana
Allium ampeloprasum
Beaucarnea recurvata
Cordyline fruticosa
Dioscorea zingiberensis
Yucca schidigera Roezl
Dioscorea panthaica
Dioscorea pseudojaponica Yamamoto
Asparagus officinalis L.
Panicum virgatum L.
Paris polyphylla var. chinensis
Paris polyphylla var. yunnanensis
Polygonatum odoratum
Fagonia indica
Tao et al. (2013)

Ji et al. (2014)
Mandal & Mandal (2010)
Voutquenne-Nazabadioko et al. (2013)
Voutquenne et al. (2005)
Wjciak-Kosior et al. (2013)
Xu et al. (2011)
Zhao et al. (2014)
Gafner et al. (2004)
Chen et al. (2007b)
Dini et al. (2009)
Li et al. (2010b); Ling et al. (2011)
Rubio-Moraga et al. (2011)
Foubert et al. (2010); Theunis et al. (2007)
Wang et al. (2007)
Xu et al. (2012)
Yoshizumi et al. (2006)
Prez et al. (2013)
Ado et al. (2011)
Eskander et al. (2011)
Fouedjou et al. (2014)
Li et al. (2010a); Qin et al. (2009); Zhang et al.
(2013a)
Kowalczyk et al. (2011)
Wang et al. (2012)
Lin et al. (2006); Lin & Yang (2008)
Dawid & Hofmann (2012)
Lee et al. (2009)
Liu et al. (2013); Zhang et al. (2010)
Zhang et al. (2010)
Bai et al. (2014)
Waheed et al. (2012)
57
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Skhirtladze et al. (2011)
AC
CE
P
TE
MA
NU
SC
RI
PT
Yucca gloriosa L.
58
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Table 3. Selection of saponins extraction technique based on research focus

Extraction
method
Pharmaceutical properties investigation
Quantification studies
Plant source (reference)
Maceration
Allium ampeloprasum (Ado et al. 2011)
Allium ampeloprasum (Ado et al. 2011)
Momordica charantia (Chen et al. 2008)

Polygonatum odoratum (Bai et al. 2014; Deng
et al. 2012)
Momordica charantia (Habicht et al. 2011)
Gynostemma pentaphyllum (Cheng et al. 2011)
Medicago truncatula (Huhman et al. 2005)
Ipomoea batatas (Dini et al. 2009)
Polygonatum odoratum (Deng et al. 2012)
Panicum virgatum L. (Lee et al. 2009)
Panicum virgatum L. (Lee et al. 2009)

Dioscorea pseudojaponica Yamamoto (Yang
et al. 2003)
Chenopodium quinoa (Verza et al. 2012)
RhizomaAnemarrhenae (Liu et al. 2012b)
Chenopodium quinoa (Zhu et al. 2002)
Solanum xanthocarpum (Patel et al. 2012)
Fagonia indica (Waheed et al. 2012)
Dioscorea zingiberensis (Qin et al. 2009)
Yucca gloriosa (Skhirtladze et al. 2011)
Chenopodium quinoa (Verza et al. 2012)
Dioscorea zingiberensis (Zhang et al. 2013a)
Fagonia indica (Waheed et al. 2012)
Leguminous species (Ha et al. 2014)

Genista ulicina Spach (Boutaghane et al.
2013)
Panax notoginseng (Yang et al. 2010)

Entada phaseoloides (Zheng et al. 2012)
SC
MA
Processing
parameters/optimization
Panax quinquefolius (Gafner

et al. 2004)
Dioscorea pseudojaponica
Yamamoto (Lin et al. 2006)
Yucca gloriosa (Skhirtladze et al. 2011)
CE
PT
ED
Momordica charantia (Gupta et al. 2010)
Cordyline fruticosa (Fouedjou et al. 2014)
AC
Glycyrrhiza yunnanensis (Ji et al. 2014)
RI
Conventional
extractions:
NU
Saponins isolation
PT
Field of research focus
Silphium asteriscus L. (Masullo et al. 2014)

Harpullia austro-caledonica (Voutquenne et
al. 2005)
Silphium asteriscus L. (Masullo et al. 2014)

Salicornia herbacea (Zhao et al. 2014)
Agave offoyana (Prez et al. 2013)

Salicornia herbacea (Zhao et al. 2014)
Cordyline fruticosa (Fouedjou et al. 2014)
Reflux
Allium nutans L. (Akhov et al. 1999)
Aralia taibaiensis (Bi et al. 2012)
Flos Lonicerae (Chai et al. 2005)
Aralia taibaiensis (Bi et al. 2012)
Xanthoceras sorbifolia (Ling et al. 2011)
Flos Lonicerae (Chai et al. 2005)
Defatted rice bran (Chan et al. 2013)

Paris polyphylla var. chinensis (Liu et al.
2013)
Paris and Trillium plants (Man et al. 2010b)

Glycyrrhiza uralensis (Tao et al. 2013)
Guar meal (Hassan et al. 2010)
Guar meal (Hassan et al. 2010)
Polygala japonica (Wang et al. 2007)
59
Defatted rice bran (Chan et

al. 2013)
Medicago sativa L. (Oleszek et al. 1998)
Panax japonicus (He et al. 2012)
Momordica charantia (Liu et al. 2012a)
Medicago sativa L. (Oleszek et al. 1998)
Maesa lanceolata (Theunis et al. 2007)

Acanthopanax sessiliflorus (Yoshizumi et al.
2007)
Momordica charantia (Popovich et al. 2010)
RI
Bupleurum chinense (Sun 2006)

Acanthopanax sessiliflorus (Yoshizumi et al.
2007)
SC
Radix Astragali (Qi et al. 2006)

Glycyrrhiza uralensis (Tao et al. 2013)
Polygala japonica (Wang et al. 2007)
Subsequent
Medicago arabica L. (Bialy et al. 2004)
Antonia ovate (Alabdul Magid et al. 2012)
methods
Medicago hybrid (Bialy et al. 2006)
Dioscorea zingiberensis (Li et al. 2010a)
Yucca schidigera Roezl.(Oleszek et al. 2001)
Tulbaghia violacea (Ncube et al. 2011)
NU
Soxhlet
Xanthoceras sorbifolia (Ling et al. 2011)

Caryocar villosum (Alabdul Magid et al.
2006)
Chenopodium quinoa (Woldemichael & Wink
2001)
AC
Ultrasoundassisted
CE
Antonia ovate (Alabdul Magid et al. 2012)

Medicago truncatula (Kapusta et al. 2005)
Dioscorea zingiberensis (Li et al. 2010a)
Tribulus terrestris L. (Dinckev et al. 2008)
PT
ED
MA
Chenopodium quinoa (Woldemichael & Wink

2001)
Vigna radiata L. (Waller et al. 1999)
Medicago Arabica (Tava et al. 2009)
Green
extraction
techniques:
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ACCEPTED MANUSCRIPT
Caulophyllum thalictroides (Avula et al.

2011)
Allium nigrum L.(Mostafa et al. 2013)

Panax quinquefolium, Panax ginseng (Wu et
al. 2001)
Bacopa monnieri (Ganzera et al. 2004)
Allium nigrum L.(Mostafa et al. 2013)

Soy and chickpea (Serventi et al. 2013)
Gleditsia sinensis Lam. (Lian & Zhang

2013)
Chiococca alba (Borges et al. 2013)
Chiococca alba (Borges et al. 2013)
Maesa lanceolata (Foubert et al. 2010)

Panax notoginseng (Lau et al. 2003; Li et al.
2005)
Caulophyllum thalictroides (Avula et al. 2011)

Dioscorea panthaica (Wang et al. 2012)
Paris polyphylla var. yunnanensis, Paris
polyphylla var. chinensis (Zhang et al. 2010)
Dioscorea panthaica (Wang et al. 2012)

Paris polyphylla var. yunnanensis, Paris
polyphylla var. chinensis (Zhang et al.
2010)
Platycodi Radix (Ha et al. 2006)

Ziziphus jujube, Z.jujuba var. spinosa (Guo et
al. 2011)
Platycodi Radix (Ha et al. 2006)

Ziziphus jujube, Z.jujuba var. spinosa (Guo
et al. 2011)
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Micowaveassisted
PT
Pulsatilla turczaninovii (Xu et al. 2012)
NU
CE
PT
ED
MA
Aesculus chinensis Bunge (Chen et al. 2007a)

Folium ginseng, Radix ginseng (Qian et al.
2009)
Panax notoginseng (Zhang et al. 2013b)
AC
Accelerated
solvent
SC
RI
Panax ginseng (Kwon et al. 2003;

Vongsangnak et al. 2004; Wang et al. 2008)
61
Aesculus chinensis Bunge (Chen et al.

2007a)
Panax quinquefolium (Engelberth et al
2010)
Xanthoceras sorbifolia
Bunge (Li et al. 2010b)
Dioscorea zingibernsis (Li et
al. 2010a)
Panax ginseng
(Vongsangnak et al. 2004)
Bupleurum chinense (Hu et
al. 2008)
Gymnema sylvestre (Mandal
& Mandal 2010)
Aesculus chinensis Bunge
(Chen et al. 2007a)
ACCEPTED MANUSCRIPT
Table 4. Summary of studies focusing on saponins extraction using maceration extraction
Pretreatment and extraction condition
Genista ulicina To isolate

Spach
triterpene
saponins.
1.16
kg/5 L
methanol
72 hours
Room
temperature
Air-dried
roots
10 kg/ -
70%
ethanol
Air-dried
powder
1 kg/10
L
80%
methanol
30 g/150
ml
-.
Ado et al.
(2011)
Three novel
cholestane-type
steroidal glycosides
and three known
steroidal glycosides
saponins have been
isolated and
identified.
Bai et
(2014)
Six triterpene
saponins have been
isolated.
Boutaghane
et al. (2013)
3 hours
60 C
Both saponin and

flavonoid fractions
showed
antiproliferation of
prostate cancer
cells effectively.
Cheng et al.
(2011)
Ethanol
extract
partitioned with nbuthanol showed
the highest antidiabetic potential.
Two new saponins
have been isolated
and identified.
Spectrophotometric
method detected
yield: 200.01
mg/100 g. HPLCDAD: saponin1:
161.20 mg/100 g;
saponin 2: 14.67
mg/100 g.
The antioxidant
activities tested of
total phytochemical
fraction and of
single saponins
were moderate in
relation to
commercial
standards.
Deng et al.
(2012)
To investigate
the
antiproliferation
and apoptosis
mechanism of
saponin and
flavonoid
fractions of
Gynostemma
pentaphyllum.
Powder
Polygonatum
odoratum
(Mill.) Druce
To characterize
the anti-diabetic
active fractions
in this herb.
Dried
5 kg/40
L
80%
ethanol
2 hours
80 C
Ipomoea
batatas tubers
To isolate,
identify and
quantify
triterpenes
glycosides.
To evaluate
their antioxidant
properties.
Flour
6 g/ -
80%
methanol
AC
CE
P
Identified a new
saponins.
Showed haemolytic
effects in vitro and
demonstrated antiinammatory
activity and
gastroprotective
property in vivo.
Boiling point
2 hours
Gynostemma
pentaphyllum
ethanol
Reference
Mechanical
aid
RI
Extraction
Temperature
SC
To isolate
steroidal
saponins.
Extraction
duration
NU
Polygonatum
odoratum
Solvent
used
MA
To isolate a
new steroidal
saponin.
To investigate
its antiinflammatory
and
antiulcerogenic
properties in
vitro and in
vivo.
Solid/
solvent
Allium
ampeloprasum
var. porrum
State of
input
material
Fresh
bulbs
Outcome(s)
PT
Objective(s)
TE
Plant
material(s)
Shaking
62
al.
Dini et al.
(2009)
ACCEPTED MANUSCRIPT
To isolate new
steroidal
saponins and
investigate their
cytotoxic and
antimicrobial
activity.
Dried
pulverize
d leaves
3 kg/ -
methanol
24 hours
Panax
quinquefolius
To investigate
the extraction
efficiency of
three solvent
systems on
triterpene
saponins from
root of Panax
quinquefolius.
Dried
roots
1:5
50%
ethanol; a
mixture
of 20%
ethanol,
40%
glycerin,
and 40%
water; or
65%
glycerin
6 weeks
Momordica
charantia
To investigate
antileishamanial
activity of the
aqueous extract
in vitro and in
vivo.
Air dried
5 kg/ -
To isolate and
identify
trinocucurbitane
and cucurbitane
triterpenoids
from roots.
To investigate
the anti-HIV
activity.
Air-dried
To quantify the
saponin extract,
the lipid extract,
and the
hydrophilic
extract in
bittergourd
varieties.
Powder
NU
SC
RI
PT
Cordyline
fruticosa
TE
MA
48 hours
Room
temperature
1.9 kg/5
L
methanol
6 hours
60 C
10 g/
150 ml
Ethyl
acetate
2 hours
AC
CE
P
water
63
Three new steroidal

saponins,
fruticoside H,
fruticoside I, and
fruticoside J, have
been isolated.
Fruticoside H and
fruticoside I
showed moderate
cytotoxic activity
against human
breast, colon, and
melanoma cell line.
Fruticoside I
showed moderate
antibacterial
activity against the
Gram-positive
Enterococcus
faecalis.
Fouedjou et
al. (2014)
The amount of total

saponins was
highest in the 50%
ethanol extract
(61.7 0.1 mg/g
dry root), although
similar to the
ethanol-glycerinwater extract (59.4
0.5 mg/g dry
root).
Gafner et al.
(2004)
Crude extract and

momordicatin
concentrations of
16mg/L and 0.4
mg/L were found
inhibiting 100%
parasites growth in
vitro. In vivo, no
parasites were
detected in hamster
at doses of
300mg/kg and
10mg/kg of crude
extract and
momordicatin.
Gupta et al.
(2010)
Five new saponins

have been isolated
and elucidated.
Possessed anti-HIV
property.
Chen et al.
(2008)
White bitter gourd

varieties were
found to contain
signicantly lower
saponin
concentrations
(0.25%) compared
to green varieties
(0.67%). The lipid
Habicht et
al. (2011)
ACCEPTED MANUSCRIPT
extract contained
high amounts of
conjugated linoleic
and linolenic acids
(up to 65.89%).
To characterize
saponins of nine
different species
of Leguminous
using UPLC.
Dried
pulverise
d seeds
-/10 ml
80%
methanol
24 hours
25 C
A total of twenty
saponins
were
characterized.
Ha et
(2014)
Sapindus
mukorossi
To investigate
the
molluscicidal
effects of
saponins from
Sapindus
mukorossi
against the
golden apple
snail.
Powder
35.5 g/ -
methanol
72 hours
Bioassay
data
revealed that the
seven
isolated
saponins
from
soapnut
were
molluscicidal,
causing 70-100%
mortality at 10 ppm
against the golden
apple snail.
Huang et al.
(2003)
Medicago
truncatula
To quantify
saponins in M.
truncatula root,
leaves, stem,
seedpod and
seeds.
Lyophili
zed
powder
10
mg/0.5
ml
80%
methanol
2 hours
Room
temperature
Shaking
Roots (5924
ng/mg/dw)
contained the
greatest total
amount of saponins
followed by leaf
(1064 ng/mg/dw)
and seed (991
ng/mg/dw),
respectively.
Huhman et
al. (2005)
Glycyrrhiza
yunnanensis
To isolate
triterpene
saponins.
Powder
18 kg/30
L
70% and
95%
ethanol
2 hours
One new oleananetype triterpenoid,

seven new
triterpene saponins
and four known
saponins have been
isolated.
Ji et
(2014)
al.
Panicum
virgatum L.
To isolate,
characterize,
and quantify of
steroidal
saponins in
Switchgrass
(Panicum
virgatum L.)
Dried
100
mg/5 ml
methanol
30 minutes
Shaking
Three types of
steroidal saponins
have been isolated
from 4 types of
Switchgrass
variety. Differences
in the relative
concentrations of
different saponins
were observed
between
switchgrass
cultivars and plant
parts.
Lee et
(2009)
al.
Freezedried
40 g/1 L
methanol
24 hours
25 C
Results showed
that the contents of
saponins were
decreased along
with increasing
cooking
temperature and
time except for the
steaming treatment.
None of the
steamed yam slices
Lin et
(2006)
al.
AC
CE
P
TE
MA
NU
SC
RI
PT
Leguminous
species
Dioscorea
To investigate
pseudojaponica
the effects of
Yamamoto
domestic
processing on
steroidal
saponins and
furostanol and
spirostanol
glycosides.
64
al.
ACCEPTED MANUSCRIPT
Freezedried
600 g/6
L
methanol
24 hours
Room
temperature
Commin
uted
1 kg/8 L
75%
ethanol
1 hour
80 C
Silphium
asteriscus L.
To isolate
saponins and
investigate their
antiproliferative
activity.
Dried
leaves
and
stems
500 g/ -
CH2Cl2methanol,
methanol,
50%
methanol
48 hours
Solanum
xanthocarpum
To establish the
scientic
rationality of
Solanum
xanthocarpum
fruit saponin
rich fraction on
antiurolithiatic
activity using in
vitro calcium
oxalate (CaOx)
crystallization
and in vivo
ethylene glycol
induced
urolithiasis in
the male albino
Wistar rats.
Fruit
powder
100 g/
500 ml
SC
NU
TE
MA
Rhizoma
Anemarrhenae
RI
To isolate and
identify
steroidal
saponins.
To investigate
the cognitionenhancing
effects of total
saponins.
PT
significantly
change their initial
compositions or
quantities of
furostanol and
spirostanol
glycosides.
50%
ethanol
24 hours
AC
CE
P
Agave offoyana To isolate

steroidal
saponins.
Dioscorea
zingiberensis
To determine
the biochemical,
hematological
and
histopathologica
l toxicity of
steroidal
saponins from
Dioscorea
zingiberensis
rhizome after
Six types of new

steroidal saponins
have been isolated
and identified.
Fndings
demonstrated that
diabetes-associated
cognitive declined.
Yang et al.
(2003)
Eighteen saponins
with highly
hydroxylated
oleanane-type
aglycones were
isolated. The
isolated
compounds showed
antiproliferative
activity against
human epitheloid
cervix carcinoma,
leukaemic T-cell
line, and colorectal
adenocarcinoma.
Masullo et
al. (2014)
The antiurolithiatic
activity in Solanum
xanthocarpum is
mediated possibly
through the
inhibition of CaOx
crystal formation
and its effect on the
urinary
concentration of
stone-forming
constituents and
nephrolithiasis
inducing factors.
Patel et al.
(2012a)
Liu et
(2012b)
al.
Dried
powder
0.5 kg/ -
70%
ethanol
48 hours
Room
temperature
Five unknown and

six known steroidal
saponins have been
isolated.
Prez et al.
(2013)
Air-dried
600 g/2
L
ethanol
12 hours
Room
temperature
The steroidal
saponins did not
show any sign of
toxicity up to oral
dose of 562.5
mg/kg in mice. No
signicant changes
of biochemical and
hematological
parameters in rats
(except at 510
Qin et
(2009)
65
al.
ACCEPTED MANUSCRIPT
acute oral
dosing in mice
and sub-chronic
oral
administration
in rats.
70%
ethanol
Powder
200 g/ -
80%
methanol
Room
temperature
1 g/100
ml
70%
methanol
PT
3.5 kg/ -
To investigate
the effect of
high and low
saponin lines of
alfalfa on pea
aphid
performance.
To evaluate the
differences in
saponins within
the studied lines
and estimate an
inuence of
some isolated
compounds on
the pea aphid
probing
behavior.
Freezedried
Chenopodium
quinoa
Seeds
To investigate
the
immunoadjuvan
t activity,
toxicity assays.
To determine
triterpenic
saponins using
UPLC/Q-TOFMS.
Dried
To isolate and
characterize of
twelve
Dried
1 hour
60 C
Shaking
AC
CE
P
TE
Medicago
sativa L.
MA
NU
SC
Yucca gloriosa To isolate and

L.
quantify
steroidal
saponins.
Dried
RI
To isolate five
steroidal
saponins
mg/kg/day).
100 g/ 1
L
40%
hydroetha
nol
solution
1 hour
4 kg/ -
90%
ethanol
3 days
50 C
Stirring
66
Room
temperature
Five steroidal
saponins have been
isolated and
identified.
Five steroidal
glycosides
including three
spirostane, one
furostane and one
cholestane
glycosides, along
with seven known
compounds have
been isolated and
characterized, The
dried extract
obtained more than
25% w/w of
glycosides.
Zhang et al.
(2013a)
The high-saponin
line of alfalfa
differed from the
low-saponin one by
the presence of
zanhic acid
tridesmoside and a
higher level of 3GlcA,28AraRhaXyl
medicagenic acid
glycoside.
The saponins
incorporated into
sucroseagarose
gels signicantly
reduced number of
the aphid probes
into the gels and
extended their
duration in
comparison to the
control gels
(without tested
compounds).
Sylwia et al.
(2006)
The two quinoa

saponin fractions
enhanced
significantly the
production of
humoral and
cellular immune
responses to
ovalbumin in mice.
Two quinoa
saponin fractions
were obtained.
Verza et al.
(2012)
Twelve triterpene
saponins have been
isolated, and their
Zhu et
(2002)
Skhirtlardze
et al. (2011)
al.
ACCEPTED MANUSCRIPT
structures were
characterized on
the basis of
hydrolysis and
spectral data,
especially NMR
evidence.
PT
triterpene
saponins.
Harpullia
austrocaledonica
To isolte
triterpenoid
saponins.
Dried
stem
bark
powder
1.110
kg/10 L
20%
methanol
17 hours
and boiled
3 hours
Fagonia indica
To isolate a
novel saponin
glycoside.
To perform the
apoptosis and
necrosis test on
three cancer cell
lines.
Dried
powder
200 g/1
L
ethanol
14 days
Room
temperature
Panax
notoginseng
To investigate
the mechanisms
of antihyperglycemic
and anti-obese
effects of Panax
notoginseng
saponins (PNS)
in KK-Ay mice.
Powder
75 g/1 L
75%
ethanol
4 hours
Salicornia
herbacea
To isolate
triterpene
saponins and
investigate their
antiproliferative
activity.
Fresh
parts
22.6 kg/
-
80%
acetone
24 hours
Ambient
temperature
Entada
phaseoloides
L.
To evaluate the
potential
therapeutic
effects of total
saponins from
Entada
phaseoloides
(TSEP) in
experimental
type2 Diabetes
mellitus
(T2DM) rats.
Dried
powder
70%
ethanol
2 hours
RI
NU
SC
AC
CE
P
TE
MA
67
Eight new and one

known acylated
triterpenoid
saponins were
isolated.
Voutquenne
et al. (2005)
A novel steroidal
saponin has been
isolated.
It was able to
induce apoptosis or
necrosis in cancer
cells depending on
the cell type.
Waheed et
al. (2012)
PNS possess antihyperglycemic and

anti-obese
activities by
improving insulinand leptin
sensitivity, and
ginsenoside Rb1 is
responsible for the
anti-hyperglycemic
effect among the
ve saponins in
KK-Ay mice.
Yang et al.
(2010)
Two new
noroleanane-type
triterpene saponins,
Salbige A and
Salbige B, have
been isolated. Both
compounds
exhibited potent
antiproliferative
activities against
cancer cells.
Total saponins
demonstrated both
hypoglycemic and
hypolipidemic in
type 2 diabetic rats
which indicating
possess antidiabetic property.
Zhao et al.
(2014)
Zheng et al.
(2012)
ACCEPTED MANUSCRIPT
Table 5. Summary of studies focusing on saponins extraction using reflux and Soxhlet extraction
Objective(s)
State of
input
material
Solid/
solvent
solvent used
duration (hr)
Reflux
extraction:
To isolate
triterpenoid
saponins
Dried
powder
400
g/3.5 L
methanol
Allium nutan L.
Isolation of four
steroidal
saponins from
underground
parts of A.
nutans.
Oven-dried
powder
300
g/500
ml
80%
methanol
Aralia
taibaiensis
To isolate four
new triterpenoid
saponins.
To investigate
the four newly
derived saponins
on antioxidant
and antiglycation
activities.
Air-dried
powder
1.2 kg/1
L
Defatted rice
bran
To fractionate
with different
solvent systems.
To investigate
the antioxidant
activity.
Oven-dried
Flos Lonicerae
To determine
triterpenoid
saponins in five
specicies of Flos
Lonicerae both
qualitatively and
quantitatively.
Xanthoceras
sorbifolia
To isolate and
quantify saponins
from parts of
husks, twig bark
and xylem, seed
coats and
kernels, flowers,
and leaves.
Alabdul
Magid et al.
(2006)
Four steroidal
glycosides including
deltoside and
nolinofuroside D and
two novel saponins
were isolated.
Akhov et al.
(1999)
70% ethanol
Four new oleanane type

triterpenoid
saponins
have been isolated.
Compounds exhibited
moderate effects on
antioxidant
and
antiglycation activities
which correlated with
treatment of diabetes
mellitus.
Bi
et
(2012)
al.
10
g/150
ml
50% ethanol
The highest yield

(236.14 17.72 mg
DE/g extract of
fraction) was obtained
in n-buthanol fraction.
Phenolic-saponins rich
fraction showed higher
antioxidant activity.
Chan et
(2013)
al.
Oven-dried
pulverized
2.0 g/
25 ml
60% ethanol
Seven major saponins

were found.
53.7 mg/g of
dipsacoside B was
found in L.hypoglauca,
more than 41.0 mg/g of
macranthoidin B was
found in
L.macranthoides, L.
confuse, and L. similis.
Chai et
(2005)
al.
Dried
1 kg/1
L
70% ethanol
Two new triterpenoid

saponins, sorbifoside A
and B, were isolated.
Among the parts, husk
was found having the
highest sorbifoside A
(157.0 g/g) and B
(58.33 g/g).
Ling et
(2011)
al.
Oven-dried
powder
-/250
ml
70% ethanol
The number of sugar

chains of Paris steroidal
saponins played a
crucial role in
hemolysis, while the
Liu et
(2013)
al.
NU
MA
D
TE
AC
CE
P
Paris polyphylla To evaluate

var. chinensis
cytotoxic and
hemolytic
activities of
steroidal
Reference
Seven triterpenoid
saponins have been
isolated and identified.
SC
Caryocar
villosum
Finding(s)
PT
Plant material(s)
Conditions
RI
Extraction
68
ACCEPTED MANUSCRIPT
Panax japonicus To study the

cardioprotective
effects of
saponins.
Momordica
charantia
50% ethanol
Saponins of Paris VI
was found majorly in
Paris fargesii (19.520
mg/g), and Trillium
tschonoskii (11.770
mg/g).
Crude saponin-rich guar
meal extract was
puried by RP-HPLC
eluting 20%, 60% and
100% methanol
fractions with 2.04
0.32%, 0.91 0.16%
and 1.55 0.15% DM
of crude saponin-rich
GM extract,
respectively.
Results indicated that
only 100% methanol
fraction and its 16 min
peak sub-fraction
exhibited both
haemolytic and
antibacterial activities
against Staphylococcus
aureus, Salmonella
Typhimurium and
Escherichia coli, but
20% and 60% MeOH
fractions stimulated
Lactobacillus spp.
growth.
Man et
(2010b)
RI
25
g/250
ml
1.5
SC
80% ethanol
MA
Ground
0.5 g/50
ml
NU
Oven-dried
powder
AC
CE
P
Guar meal
To determine
major saponins in
Paris and
Trillium plant
qualitatively and
quantitatively.
To isolate
saponin rich
extract.
To evaluate its
fractions for
haemolytic and
antibacterial
activities against
two gram
positive bacteria
(Lactobacillus
spp. and
Staphylococcus
aureus) and two
gram negative
bacteria
(Escherichia coli
and Salmonella
Typhimurium) in
a dose dependent
manner.
TE
Paris and
Trillium plants
kinds of saponins
inuenced the
mechanism and
cytotoxicity. Also the
results of the cytotoxic
and hemolytic activities
of plants suggested that
the existed correlation
of the steroidal saponins
affected the activities.
PT
saponins.
al.
Hassan et al.
(2010)
Dried
pieces
1 kg/4
L
60% ethanol
Saponins from Panax

japonicus exerted
benecially
cardioprotective effects
on myocardial ischemia
injury rats, mainly
scavenging oxidative
stress-triggered
overgeneration and
accumulation of
reactive oxygen species,
alleviating myocardial
ischemia injury and
cardiac cell death.
He et
(2012)
al.
To investigate
the chemical
constituents from
fresh fruit.
Fresh fruits
3 kg/24
L
90% ethanol
1.5
A new C30 sterol

glycoside was isolated.
Liu et
(2012a)
al.
To isolate and
identify new
triterpenoid
saponins.
To investigate
Air-dried
powder
30
kg/100
L
ethanol
Identified
fourteen
cucurbitane
triterpenoids.
Exhibited weak antiHIV activity.
Chen et
(2009)
al.
69
ACCEPTED MANUSCRIPT
their anti-HIV
activity in vitro.
methanol
Saponins containing
extract reduced
preadipocyte
proliferation and lipid
accumulation of the
adipocyte.
Popovich
al. (2010)
et
200
mg/10
ml
1.5
It was shown that

monodesmosidic
medicagenic acid
glycoside was
synthesized after 4 days
of germination and
subsequently followed
by bidesmosidic
saponin production.
The total saponin
concentration increased
from 2.12 mol/g of dry
matter at the beginning
of germination toaround
6mol/g after 8-16 days
of seedling growth.
Oleszek
(1998).
methanol
Four main saponins

have been isolated
successfully.
Bupleurum chinense
saponins showed a
slight haemolytic effect
and enhanced
signicantly a specic
antibody and cellular
response against
ovalbumin in mice.
Qi et al.
(2006)
30%
methanol
NU
Freezedried
powder
SC
RI
PT
TE
Medicago sativa To determine the

var. Boja
occurrence of
individual
saponins.
To verify early
biological data
on their
quantification
with new
analytical
techniques.
Lyophilized
pieces
MA
To investigate
two classes of
saponins, ie.,
cucurbitane and
oleanane type
triterpenoids, for
the potential to
reduce
preadipocyte
viability, lipid
accumulation and
adiponectin
expression in
3T3-L1 cells.
Radix Astragali
To isolate four
main saponins.
Bupleurum
chinense
To evaluate the
haemolytic
activities of
Bupleurum
chinense
saponins and its
adjuvant
potentials on the
immune
responses of ICR
mice against
ovalbumin.
Powder
1 kg/ -
70% ethanol
Glycyrrhiza
inflate,
Glycyrhiza
glabra,
Glycyrrhiza
uralensis
To isolate and
quantify
triterpenoid
saponins from
species of
Glycyrrhiza.
Dried
powder
0.5 g/30
ml
50%
methanol
Ten saponins have been

isolated and the total
saponins yields were
found in the range of
28.958-119.750 mg/g.
Tao et
(2013)
Maesa
lanceolata
To determine
saponins using
LC-UV method.
Dried
powder
1.5 g/70
ml
50%
methanol
Compounds of oleanane
type triterpenes were
identified.
Theunis et al.
(2007)
Polygala
japonica
To isolate and
quantify
triterpenoid
saponins.
Dried
powder
1.0 g/50
ml
methanol
Six triterpenoid
saponins have been
isolated. The total
saponins found were
4.45-27.32 mg/g.
Wang et al.
(2007)
AC
CE
P
Oven-dried
powder
1.5 g/60
ml
70
Sun (2006)
al.
ACCEPTED MANUSCRIPT
Air-dried
powder
10
kg/80 L
Hot water
To identify and
determine
biological
activities of
triterpenoid
saponins.
Dried
powder
900 g/ -
petroleum
ether,
methanol
Yoshizumi et
al. (2006)
72
Sixteen saponins were

detected and isolated.
Both bidesmosides and
derived
monodesmosides
showed little or no
antifungal
activity,
whereas
a
comparatively
higher
degree of hemolytic
activity
could
be
determined
for
monodesmosides.
Woldemichael
&
Wink
(2001)
24
Saponins produced by
mungbean plants added
to the soil enhanced the
growth
of
new
mungbean plants as an
allelochemical
plant
growth regulator.
Waller et al.
(1999)
MA
NU
SC
Soxhlet
extraction:
Chenopodium
quinoa seed
Three known and one

novel saponins have
been isolated.
Sessiloside and
chiisanoside inhibited
pancreatic lipase
activity in vitro, and
addition of the saponinrich fraction to a highfat diet suppressed the
body weight gain of
mice.
PT
To isolate four
saponins.
To determine
their inhibitory
activities on
pancreatic lipase
in vitro and in
vivo.
RI
Acanthopanax
sessiliflorus
leaves
Chloroform,
80% ethanol
TE
Fresh plant
AC
CE
P
Vigna radiata L. To focus on plant

adaptability,
stages of growth,
concentration
and type of
allelochemical
measurement of
biological
activity, etc. to
fully explore the
allelochemical
response.
71
ACCEPTED MANUSCRIPT
Table 6. Summary of studies focusing on saponins extraction using two subsequent extractions
Objective(s)
Pretreatment/Procedures
Finding(s)
References
Subsequent methods employed:
Soxhlet reflux
Eight major triterpene

saponins have been
isolated.
al.
al.
To isolate eight major triterpene

saponins.
The powdered plant material was

defatted with chloroform in a
Soxhlet apparatus and then
extracted with 80% methanol
under reflux.
Bialy et
(2004)
Medicago
hybrid roots
To isolate fourteen triterpene

saponins.
The ground roots were defatted with

chloroform in a Soxhlet apparatus
for 48 hours and then extracted
with 95% methanol under reflux
for 24 hours..
Fourteen triterpene
saponins have been
isolated.
Bialy et
(2006)
Yucca
schidigera
Roezl
To isolate and characterize the

dominant saponins of yucca trunk.
Yucca powder (300 g) was defatted

with chloroform in a Soxhlet. After
drying, it was extracted three times
by boiling with water for half an
hour.
Eight steroidal saponins

have been isolated.
Oleszek et al.
(2001)
Powdered leaves (500 g) were

defatted with chloroform in a
Soxhlet apparatus (fats 4.3% DM).
Defatted material (200 g) was then
extracted with 80% methanol
under reux for 24 hours.
Fourteen known saponins

have been isolated, and
five new saponins with a
hydroxy group at the 30methyl position of the
triterpenic skeleton were
identified.
Tava et
(2009)
SC
NU
MA
TE
Medicago
To acquire further evidence of the
arabica (L.)
previously reported compounds in
leaves
M. arabica, and elucidate the
chemical structure of these newly
identied saponins.
RI
Medicago
Arabia L.
PT
Plant
materials
al.
Maceration reflux
To isolate six new

polyhydroxyoleanene saponins.
To test the effects on the growth of
human oral epithelium carcinoma
(KB) cell line.
Dried powdered leaves were

macerated in 5 litres of petroleum
ether for 6 hours and percolated
during 12 hours.
The solvent was evaporated and the
dried extract was then macerated for
another 2 hours with 5 litres of
90% aqueous methanol and
further reuxed for 3 hours.
Six pentacyclic
triterpenoid saponins,
named antoniosides EJ
along with two known
alkaloids, were isolated.
Inhibited the growth of
oral epithelium carcinoma
cell.
Alabdul Magid
et al. (2012)
Tribulus
terrestris L.
To quantify the distribution of

steroidal saponins of the plant from
different geographical regions.
One gram of the plant material

(aerial parts, fruits, leaves and
stems) was extracted with 50 ml
chloroform for 1 hour, by shaking
at room temperature.
The extract was ltered and the
plant material was extracted again,
three times by reuxing with 50 ml
of 70% ethanol for 1 hour.
Samples from Bulgaria

were found to contain a
rather high percentage
(0.245-1.337%) of
protodioscin, whereas
samples from China
contained only small
amounts (0.063% and
0.089%) of this
compound.
Dinchev et al.
(2008)
Medicago
truncatula
To identify and determine saponins

in aerial parts of barrel medic using
LC-ESI/MS/MS.
One gram of dried and finely

powdered barrel medic tops (leaves
and stems) was extracted overnight
with 50 mL of 80% methanol at
room temperature.
The extract was filtered, and the
residues were additionally extracted
twice by refluxing with 50 mL of
80% methanol for 1 hour.
Aerial parts of all three

subspecies contained 17
saponins previously
identified and also a
substantial amount of
astragaloside VIII (3GlcA-Xyl-Rha
soyasapogenol B), not
previously reported in M.
truncatula.
Kapusta et al.
(2005)
AC
CE
P
Antonia
ovata leaves
72
ACCEPTED MANUSCRIPT
A chemical ngerprint
method was rstly
established and validated
to quantify and
standardize rhizomes
including parvioside,
protodeltonin,
protodioscin,
protogracillin,
zingiberensis saponin,
deltonin, dioscin and
trillin.
The results indicate that
total steroidal saponins
could inhibit thrombosis
by both improving the
anticoagulation activity
and inhibiting platelet
aggregation action,
suggesting that total
steroidal saponins from
Dioscorea zingiberensis
rhizomes have the
potential to reduce the risk
of cardiovascular diseases
by antithrombotic action.
Li et al. (2010a)
Extracts of
micropropagated plants
and outdoor grown plants
showed good antibacterial
activity in vitro.
Ncube
(2011)
PT
An accurately weighed sample of

600 g powder was put into a round
bottom ask, soaked with 2000 ml
ethanol at room temperature for 24
hours, and then reuxed with a
mechanical stirrer at 78 C for 2
hours. The heat-reux extraction
process was repeated twice.
The dried and ground plant samples

were defatted with hexane in a
Soxhlet apparatus for 3 hours.
After air drying, saponins were
extracted twice from the defatted
samples (10 g) in 100 ml of 50%
aqueous methanol by incubating at
room temperature overnight with
continuous stirring.
TE
To compare the antimicrobial

and phytochemical properties of
in vitro cultured and outdoor
grown Tulbaghia violacea plants
in the quest to validate the use of
micropropagated plants as
alternatives to outdoor grown
plants in traditional medicine.
AC
CE
P
Tulbaghia
violacea
Soxhlet maceration
MA
NU
SC
RI
Dioscorea
To characterize the major active
zingiberensis
constituents.
To evaluate the anti-thrombotic
activity.
73
et
al.
ACCEPTED MANUSCRIPT
Findings
Reference
Eight triterpene saponins

(cauloside H, leonticin D,
cauloside G, cauloside D,
cauloside B, cauloside C,
cauloside A and saponin PE)
were separated within 35 min
using HPLC method and within
8.0 min using UPLC method
with detection limits of 10
g/mL for saponins.
Avula
(2011)
et
al.
RI
Pretreatment/Extraction
procedure
Dry plant samples were weighed
and sonicated in 2 ml of methanol
for 30 minutes.
NU
SC
Plant
Objectives
material(s)
Caulophyllum To compare
thalictroides
chromatographic
performance of HPLC and
UPLC in determining
saponins from the roots.
PT
Table 7. Summary of studies focusing on saponins extraction using ultrasound-assisted

extraction (UAE)
To isolate five triterpenoid

saponins from the roots.
To investigate their
inflammatory activity in
vitro.
The ethanolic extract of the

pulverized dried roots of C. alba
(1 kg)was obtained with the
assistance of an ultrasonic bath,
after extraction with hexane and
methylene chloride.
Five types of triterpenoid

saponins have been isolated and
identified.
The saponin fractions have been
observed against in vitro
lipopolysaccharide-induced
inflammation.
Borges
(2013)
et
al.
Maesa
lanceolata
To evaluate the efficiency

of UPLC in quantifying 14
saponins from the leaves.
Hundred grams of dried plant

material was sonicated in 5 ml of
50% methanol (v/v) for 1 hour.
Foubert
(2010)
et
al.
Bacopa
monnieri
To separate and isolate

triterpenoid saponins.
To determine and quantify

ten major saponins.
TE
A rapid and sensitive UPLC

MS/MS method was developed
to relatively quantify the
amount of 14 individual
maesasaponins present in the
plant material of M. lanceolata.
One gram of finely powdered plant

material was extracted three times
with 3 ml methanol by sonication
for 10 minutes.
Several B. monnieri samples

(extract, plant material,
commercial products) were
successfully analyzed, each of
them containing at least four of
the seven reference
compounds.
Main components were either
bacoside A3 or bacopaside II,
least dominant showed to be
bacopasides IV and V.
The total saponin content in the
samples varied from 1.1 to
13.0%.
Ganzera et al.
(2004)
The dried powders of Z. jujube and

Z. jujube var. spinosa leaves (1.0 g,
40 mesh) were extracted with 20
ml of 80% methanol by
ultrasonication (40kHz) at room
temperature for30 minutes.
,
Approximately 1 g of the nely
powdered Platycodi Radix was
extracted three times with 10 ml of
each solvent system (water, 30%
MeOH, 50% MeOH, 70% MeOH
and MeOH) by sonication for 10
Two saponins were separated

and quantified having 1.41
mg/g of zizyphus saponins I
and 4.52 mg/g of zizyphus
saponins II, respectively.
Guo et al. (2011)
Ten major saponins have been

identified with the most
majorly discovered were
platycodin D of 2431 g/g,
polygalacin D of 2116 g/g,
and platycoside E of 1088
g/g.
Ha et al. (2006)
AC
CE
P
Ziziphus
To perform simultaneous
jujuba,
qualitative and quantitative
Z.jujuba var.
analysis in the leaves.
spinosa
Platycodi
Radix
MA
Chiococca
alba
74
ACCEPTED MANUSCRIPT
minutes.
The contents of ginsenosides

Rg1, Re, Rb1, Rd, and
notoginsenoside R1 in most of
the raw samples were found to
be higher than those in the
corresponding steamed
samples.
Lau et al. (2003)
To quantify six major active

saponins using HPLC.
Li et al. (2005)
RI
PT
To analyze saponins in raw

and steamed P.notoginseng.
MA
NU
SC
Panax
notoginseng
Gleditsia
To determine the contents
sinensis Lam.
of saponins.
Each of the ground gleditsia

materials (1.501.60 g) was placed
in a 250 mL ask with 50 ml of
AC
CE
P
TE
methanol and then by ultrasonic

treatment for three times (each 5
min).
Soy
and To investigate the stability
chickpea
during breadmaking and in
vitro bioaccessibility of
saponins from soy and
chickpea.
The established HPLC analytic

method was successfully used
to determine the concentrations
of 29 compounds including 19
gleditsia saponins and ten
unidentied eight commercial
gleditsia fruits from different
sources. The results from this
study suggested that this newly
developed HPLC method could
be used for qualitative and
quantitative analysis of the
saponins in the ingredients.
Lian & Zhang

(2013)
Entire loaves of breads were

processed to a ne paste with a
grinder and aliquots (150 mg) were
mixed with 3 mL of 70% ethanol
in 4 mL vials.
Mixtures were sonicated for 2 min,
passed through a 0.2 m nylon
lter.
Saponin structure and food

matrix aect the stability of
saponins during processing and
digestion and that uptake of
saponins by enterocyte-like cells
is poor despite moderate
apparent bioaccessibility.
Serventi
(2013)
et
al.
et
al.
Allium
nigrum L.
To determine the content of

cysteine sulfoxides, total
polyphenols, and total
saponins in different organs
of A. nigrum by using highperformance liquid
chromatograph (HPLC) and
spectral techniques.
To isolate, quantify, and
assess the antifungal
activity of the aginoside
compound in a wide range
of phytopathogens.
Fresh rootbulb basal stem of A.

nigrum (80 g) was hand-cut and
air-dried at room temperature, and
the nal dry weight (30 g) was
obtained and used for this study.
The dry weight was exhaustively
extracted at room temperature with
the following solvents: n-hexane
and 70% methanol.
Each solvent extraction step was
conducted for 1 day and repeated
three times with 30 min of
sonication.
The HPLC and spectral analyses

of cysteine sulfoxides (CSOs),
total polyphenols (TP), and total
saponins revealed quantitative
variations within the different
organs of Allium nigrum L. A
large accumulation of CSOs was
detected in the bulb (0.367 mg/g
fw), of TP in the leaf (116.05
mg CE/100 g fw), and of
saponins in the root
(19.38mg/gdw).
Aginoside saponins showed
significant antifungal activity
depending on the concentration.
Mostafa
(2013)
Dioscorea
panthaica
To isolate and quantify

steroidal saponins.
The ne powder of plant (0.4 g)

was accurately weighed and placed
in a conical ask. After adding 20
Six steroid saponins were

determined. The saponins found
were in the range of 2571.3-
Wang
(2012)
75
et
al.
ACCEPTED MANUSCRIPT
Powder of plant material (0.5g)

was extracted in 50 ml of
methanol with sonicated for 30
minutes.
Ten and seven saponins

were determined in P. polyphylla
var. yunnanensis and P. polyphylla
var. chinensis, respectively,
including four unknown
compounds. Total saponins for P.
polyphylla var. yunnanensis was
19.62 mg/g and P. polyphylla var.
chinensis was 1.63 mg/g.
TE
MA
NU
SC
RI
To isolate and quantify

steroidal saponins.
AC
CE
P
Paris
polyphylla
var.
yunnanensis,
Paris
polyphylla
var.
chinensis
37435.1 g/g.
PT
mL of 75% ethanol to theask,

the mixture was sonicated for 60
min.
76
Zhang
(2010)
et
al.
ACCEPTED MANUSCRIPT
Table 8. Summary of studies focusing on saponins extraction using microwave-assisted

extraction (MAE)
Extraction procedure
To develop a novel
MAE method, and
to evaluate MAE
and conventional
extraction
techniques for the
extraction of
triterpenoid
saponins from
Ganoderma atrum.
MAE was performed in the closed vessel unit

MDS2002 (Xinyi Microwave Extraction
Instrument Company, Shanghai) equipped with
temperature sensor. Maximum oven power for
this system is 800 W. For the rst series of
experiments 3 g of the whole dried and ground
material were placed in 100 ml PFTE (CEM)
extraction vessels and appropriate amount of
solvent was added (30 or 75 ml). The extraction
temperature was set at dierent degrees due to
dierent solvents and programmed as follows:
ramp to the temperature for 5 min step by step,
hold at temperature for a few minutes.
Microwave power was 800 W (100%). After
extraction, the vessels were left for 30 min to
cool down below 35 C.
Panax
notoginseng
To compare the
efficiency of MAE
with the
conventional
extraction methods
The extractions were performed using MAP

extractor (Prolabo, France) at full power (300
W) and an emission frequency of 2450 MHz for
30 s on a mixture consisting of different amounts
of ginseng powders (2.5, 5.0, 10.0 g) and 50 ml
of 80% methanol.
Finding
Reference
Compared with shaking

extraction method, heat
reux extraction,
supercritical uid carbon
dioxide extraction (SFE) and
normal ultrasound-assisted
extraction (UAE), MAE only
need 5 min to give the
highest yield of triterpenoid
saponins at 0.968%, while
the other extraction methods
need several hours or even
more than 10 h and give
lower yield.
Chen et al.
(2007b)
Results indicated that the

MAP was more superior
than the conventional
method in its capability to
extract target components
without causing any
degradation. Additionally, it
dramatically reduced the
extraction time from 12 h to
a few seconds, suggesting
that it can be an alternative
technique to the timeconsuming conventional
reux method.
Kwon et al.
(2003)
Vongsangnak
et al. (2004)
PT
Objectives
AC
CE
P
TE
MA
NU
SC
RI
Plant
material(s)
Ganoderma
atrum
To find an efficient
extraction method
and optimize the
MAE for saponin
from cultured cells
of Panax
notoginseng.
Microwave-assisted extraction was performed

using a household microwave oven of 2450 MHz
(P182, Whirlpool, USA). Fasks with 100 mg of
dried cells and 15 ml of water-saturated nbutanol were exposed to the microwave (at a
power of 90 or 125 W). A temperature of 20, 50
or 80 C was controlled by interrupting
the irradiation every 15 or 30 s and cooling the
irradiated samples in an ice-bath.
To evaluate the
efficiency of high
pressure MAE as a
fast extraction of
ginsenosides from
Panax notoginseng.
The high pressure MAE was performed in a

WRT-C microwave preparation system with a
pressure, temperature and time control system
(Meicheng Technology Co., Ltd., Beijing,
China). A sample powder of 1.00 g was
accurately weighed, transferred into the liner
vessel and immersed by 40 mL extraction
solvent. After the closed control and extraction
vessels were put into the microwave preparation
system, the pressure was turned on and began to
gradually increase. When the pressure reached
preset pressure, irradiation time was counted and
77
The optimal extraction

method was microwaveassisted extraction for 4 min
of extraction time at 50 C
of extraction temperature (at
a radiation power of 125 W)
and 1:150 of the solid/liquid
(w/v) ratio.
Wang et al.
(2008)
ACCEPTED MANUSCRIPT
the extraction was carried out continuously at the
preset pressure.
Xanthoceras
sorbifolia
Bunge.
kernel
To optimize the
microwave-assisted
extraction
procedure for
triterpene saponins
from the defatted
residue of yellow
horn kernel.
MAE is carried out using MASII Microwave

Extraction Testing Equipment. One gram
defatted residue was placed into the extraction
vessel.
The initial conditions were as follows:
temperature 30 C, duration 3 min, irradiation
power 300 W, ratio of solvent to material 20
ml/g, 10% ethanol and 1 extraction cycle.
For the subsequent condition optimization of
MAE, the above parameters were chosen
differently according to the experimental design.
Dioscorea
zingibernsis
To extract the total

steroids in the
culture broth from
the fermentation of
Dioscorea
zingibernsis using
microwave-assisted
aqueous-two-phase
extraction
(MAATPE) and a
novel three-liquidphase system was
developed to
separate diosgenin
from untransformed
steroidal saponins.
Extraction of total steroid was performed using

MAATPE.
Thirty milliliters of culture broth obtained from
the fermentation of 1 g DZW powder was mixed
with water, (NH4)2SO and absolute ethanol in a
ask and placed into a household microwave
oven retted with a condensator outside, and
irradiated at medium level power for 2 min.
The mixture was cooled to room temperature and
the top phase was taken for analysis.
Gymnema
sylvestre
To evaluate the
applicability of
MAE on oleanolic
acid extraction from
Gymnema sylvestre
leaves.
One gram of homogenous leaf powder was

mixed with 20 ml of ethanol. After allowing a
preleaching time of 5 min, the suspension was
irradiated with a microwave extractor (CATAR,
Catalyst Systems, Pune, India) at different
experimental conditions.
Pulsatilla
turczaninovii
To develop a
method for
quantifying and
MAE was carried out in a QW-6HME (6000 W,

2450 MHz) microwave accelerated reaction
system (KeWei Microwave energy Co., Ltd.,
Hu et
(2008)
al.
The optimum extraction

parameters were 51 C, 7
min, 900 W, 32 ml/g, 42%
ethanol and 3 cycles,
obtaining the highest
extraction yield of triterpene
saponins of 11.62 0.37 %
of defatted kernel.
Li
et
(2010b)
al.
An improved MAATPE
Li
et
(2010a)
al.
Under optimum conditions,

8 min of MAEproduced a
maximum yield of 7.6%
(w/w) of oleanolic acid
which was found to be 4
times, 3 times and 1.2 times
more efcient than
maceration, stirring
extraction and heat reux
extraction, respectively.
Extracts obtained from 8 min
of MAE showed better
antioxidant activity when
compared to other
conventional methods. No
degradation of the target
analyte was observed at the
optimum conditions as
evidenced from the stability
studies performed with
standard oleanolic acid.
Mandal
Mandal
(2010)
&
The total triterpenoid

saponins content was the
highest in April (18328.6
Xu et
(2012)
al.
PT
Experiments were carried out using a custom

made microwave reactor with cylindrical cavity
(model 961, Microwave Power Consultants, VIC,
Australia). This microwave reactor is a
continuously variable microwave power system
with power output up to 1000 W and a bre
optical temperature controller.
AC
CE
P
TE
MA
NU
SC
RI
To study the
relationships
between the
extraction yields of
saikosaponins and
the operating
parameters of
MAE.
Bupleurum
chinense
78
could thoroughly extract the

steroids from the
fermentation and subsequent
application of three-liquidphase system with optimum
conditions of 30% ethanol,
17% (NH4)2SO (w/w) and 40%
petroleum ether yielded top
phase diosgenin recovery of
97.24%.
ACCEPTED MANUSCRIPT
PT
g/g), the seedling stage, and

decreased from May
(18295.9 g/g) to August
(17157.6 g/g). The contents
of total triterpenoid saponins
decreased signicantly after
the blooming period.
MAE in closed system was
the most effective technique.
The best results for ursolic
obtained (111.2 g/g DM)
with use of MAE in
closed system for 10 min
and 100% of generator
power. Oleanolic acid (22.2
g/g DM) was better
extracted with use of milder
conditions (30% generator
power and 30 min).
SC
RI
MAE was carried out using Plazmotronika

UniClever (350W) BMZ I(Wroclaw, Poland).
Plant material was placed into the extraction
vessel and extracted with 40 mL of acetone
using various generator powers (30%, 65%,
100%) during 10, 20 and 30 min. The obtained
extracts were ltered, evaporated, transferred
into 5 mL asks and made up to the mark with
acetone.
TE
MA
To compare six
different extraction
methods, i.e.,
maceration,
Soxhlet, heat reflux
extraction,
ultrasonic
extraction,
microwave
extraction, and
accelerated solvent
extraction, on
oleanolic acid and
ursolic acid from
Lamii albi flos.
AC
CE
P
Lamii albi
flos
Guangzhou, China). One gram of powder was

weighed accurately into a round bottom ask
(100 ml) and extracted under optimized
condition, including a solid/liquid ratio of 1:20
(g/ml), a microwave power of 500 W, an
extraction temperature of 80 C and an
irradiation time of 3 min with 70% ethanol for
one cycle.
NU
comparing the
triterpenoid
saponins in nine
different seasonal
batches of P.
turczaninovii from
the Liaoning
province.
79
WjciakKosior et al.
(2013)
ACCEPTED MANUSCRIPT
Plant
material(s)
Aesculus
chinensis
Bunge
Objectives
Pretreatment/Extraction conditions
To identify and
quantify four major
saponins using HPLC
method.
To introduce a new
extraction process,
accelerated solvent
extraction, to optimize
the extraction yield.
An ASE 100 System (Dionex,

Sunnyvale, CA, USA) with 34-mL
stainless steel ASE vessels was used
for the pressurized liquid extraction.
.
Folium
Ginseng and
Radix Ginseng
To develop a rapid
pressurized liquid
extraction and rocket
column HPLC
analysis method for
the determination of
one avonoid
(panasenoside), nine
saponins of
ginsenoside and two
polyacetylenes
(panaxydol and
panaxynol) in Folium
Ginseng and Radix
Ginseng.
The sample in the extraction cell

was extracted using pressurized
liquid extraction on a Dionex ASE
200 system under the optimum
conditions: methanol; particle size,
0.300.45 mm; temperature, 150
C; static extraction time, 15 min;
pressure 1500 psi; ush volume,
40%; static cycle, 1 and number of
extraction, 1.
Panax ginseng
To quantify and
separate
protopanaxatrial and
protopanaxadiol
saponins with
macroporous resins.
To determine nine
types of saponins
from Panax ginseng.
To compare the
extraction efficiency
of methods
pressurized liquid,
ultrasonication, and
Soxhlet extraction.
To demonstrate a
novel simpler and
faster three-stage
temperature gradient
ASE coupled with
HPCCC for the
systematic extraction
and online separation
of saponins with a
broad range of
polarity from the raw
plant materials.
PT
Table 9. Summary of studies focusing on saponins extraction using accelerated solvent

extraction (ASE)
Findings
Reference
Chen et al.
(2007a)
A rapid pressurized liquid extraction

(PLE) and high-performance liquid
chromatography coupled with diode
array detection and mass spectrometry
(HPLC-DADMS) method for the
simultaneous determination of one
avonoid (panasenoside), nine
saponins (ginsenoside Rg1, Re, Rf,
Rg2, Rb1, Rc, Rb2, Rb3 and Rd) and
two polyacetylenes (panaxydol and
panaxynol) in Folium Ginseng and
Radix Ginseng was developed.
Qian et al.
(2009)
The dried powder of P. notoginseng

(90 g) was placed into six 33 mlstainless steel extraction cells. In
brief, the optimized conditions
were: particle size, 0.30.45 mm;
solvent, methanol; temperature, 150
C; static time, 15 min; pressure,
6.895 10 3 MPa; static cycle, 1.
The adsorption characteristics of PTS

and PDS on four types of macroporous
resins, including D-101, DA-201, DM301 and DS-401, have been compared.
Among them, DS-401 resin showed
the best separation behaviors.
Wan et al.
(2008)
Dried powder was placed into an 11

ml stainless steel extraction cell.
The optimized conditions were:
particle size, 0.30.45 mm; solvent,
methanol; temperature, 150 C;
pressure, 6.895 103 MPa; static
time, 15 min; and one static cycle
and one extraction times.
Nine types of saponins have been

identified.
PLE has the highest extraction
efciency and repeatability, which
would be valuable on standardization
of sample preparation for quality
control of Chinese medicines.
Wan et al.
(2006)
The extraction cells with

notoginseng powder were placed
into the ASE 150 system and the
extraction conditions and process
were: rstly, static in 10 min,
followed by a ush elution with
60% volume, and followed by the
nitrogen purge of 250 s, and
extracted once.
The extraction temperature and the
extraction solvents were optimized
in the subsequent experiments.
More than nine saponins including

notoginsenoside R6, notoginsenoside
R1, ginsenoside Rb1, notoginsenoside
Spt1, ginsenoside F4, ginsenoside Rh4,
ginsenoside 20S-Rg3, ginsenoside 20SRs3 and ginsenoside Rk1 with the
corresponding extraction rates of 0.78
mg/g, 2.23 mg/g, 1.86 mg/g, 0.83
mg/g, 1.23 mg/g, 1.46 mg/g, 2.47
mg/g, 2.22 mg/g and 1.84 mg/g were
extracted and online isolated by ASE
coupled with HPCCC from the nature
Zhang et al.
(2013b)
AC
CE
P
TE
MA
NU
SC
RI
Four major saponins of escin Ia (17.3

1.4 mg/g), escin Ib (10.4 1.6 mg/g),
isoescin Ia (9.3 2.5 mg/g) and
isoescin Ib (5.9 0.8 mg/g) were
extracted from seeds of A. chinesis
Bunge using ASE.
The optimized ASE procedure
employed 70% methanol, 120 C, 7
min of static extraction time, 60% ush
volume resulting extraction recoveries
of the four compounds nearly to 100%
for two cycles.
80
ACCEPTED MANUSCRIPT
AC
CE
P
TE
MA
NU
SC
RI
PT
plant P. notoginseng.
81
ACCEPTED MANUSCRIPT
Table 10. Spectrophotometric method in quantifying total saponins from various plant materials
0.2ml of 5%
vanillin-acetic
acid + 1.2ml
perchloric acid.
70 C, 15 min
Ipomoea batatas tuber
0.5ml 8%
vanillin
in ethanol + 5
ml of 72%
suiphuric acid
in water.
60 C, 20 min
0 C, 5 min
Xanthoceras
sorbifolia Bunge
kernel
0.2ml 5%
vanillin-acetic
acid + 1.2ml
70% perchloric
acid
70 C, 20 min
Running water, 2
min
Bryonia Laciniosa
0.5ml 8%
vanillin + 5ml
72% sulphuric
acid
60 C, 10 min
Defatted rice bran
Allium nigrum L.
0.7% vanillin60% sulphuric

acid reagent
Tulbaghia violacea
soymilk
Selected
wavelength
(nm)
Oleanolic acid
Oleanolic acid
Type of
spectrophotometer
PT
Ganoderma atrum
Standard used
550
CE
Ice water bath, 15

min
MA
Oleanolic acid
References
5.11% (the highest using

MAE)
Chen et al.
(2007b)
544
200.01 mg/100g dry

weight
Dini et al. (2009)
550
Unico 2100
Spectrophotometer
(Shanghai Unico
Equipment Co. Ltd.,
China
The optimum
microwave-assisted
extraction parameters of
51 C, 7 min, 900 W, 32
ml/g, 42% ethanol and 3
cycles yielded the
highest extraction of
triterpene saponins
reached 11.62 0.37% of
defatted kernel.
Li et al. (2010b)
538
15 g/mg
Patel et al.
(2012b)
236.14 mg DE/g extract
Chan et al. (2013)
PT
ED
Oleanolic acid
Yield obtained
Double beam
RI
Cooling
conditions before
UV/vis
measurement
Running water, 2
min
SC
Conditions to allow
full colour
development
NU
Reagents
mixture
AC
Plant material
diosgenin
diosgenin
473
U-2001
Spectrophotometer
(Hitachi, Tokyo,
Japan)
Yields (mg/g dw):

Roots: 19.38
Bulbs: 15.65
Leaves: 10.48
Mostafa et al.
(2013)
250l of
vanillin reagent
(8g/100ml
ethanol) +
2.5ml of 72%
sulphuric acid.
60 C, 10 min
Ice water bath, 3-4

min
diosgenin
544
Yields (mg DE/ml):

Micropogated: 25.14
Outdoor grown: 8.94
Ncube et al.
(2011)
0.5ml of 8%
60 C, 10 min
Ice water
Soyasaponin
544
multidetection
For both unfermented
Lai et al. (2013)
82
ACCEPTED MANUSCRIPT
microplate reader
(SynergyTM HT,
BIO-TEK, GA,
USA).
Soyasaponin
Seeds of Achyranthus
aspara, Tribulus
terrestris, and Albizia
lebbeck
Rhizoma
anemarrhenae
Roots of Panax
quinquefolium and
Panax ginseng
544
0.25 ml of
vanillin reagent
(8%, w/v in
99.9% ethanol)
+ 2.5 ml of 72%
(v/v) sulphuric
acid.
60 C, 10 min
Ice water bath
Quillaja saponin
6.0ml of H2SO4
60 C, 30 min
Cool water
0.2ml 5%
vanillin-acetic
acid + 0.8ml
perchloric acid
60 C, 15 min
and fermented soymilk

extracted with 80%
methanol gave the
highest yields of 247.44
and 167.21 mg saponin/g
extract, compared to
50% acetone and water.
Black soybean soup:

76.60-108.50 mg/g;
adzuki bean soup:
103.18-129.49 mg/g;
nungbean soup: 32.8439.23 mg/g.
Song et al. (2013)
544
The total saponins

content of seeds in chaff
tree, gokhru and Siris
were 45.75, 25.65 and
48.26% (w/w),
respectively.
Goel et al. (2012)
sarsasapogenin
283
Purity 65.3%
Liu et al. (2012b)
ginsenoside
560
It was found that UAE of

ginseng saponins was
about three times faster
than the traditional
extraction methods.
Wu et al. (2001)
AC
CE
PT
ED
MA
NU
SC
Bian-Que bean soup

(consist of black
soybean, mung bean
and adzuki bean)
RI
PT
vanillin solution
(in ethanol), and
5mL of 72%
sulfuric acid.
83
ACCEPTED MANUSCRIPT
Table 11. Quantification of plant saponins using HPLC

Detector
Column
Caulophyllum
thalictroides
Waters
Alliance, 996
photodiode
array
Agilent 1100, -
Phenomenex LC18
guard
Solvent system
detection
Ammonium acetateacetonitrile
Zorbax SB C18
Acetonitrile-acetic acid
Agilent 1300,
DAD 1300
diode array
Agilent 1100,
UV-Vis diode
array
Zorbax SB C18
Agilent 1100
series, G1315B
photodiode
array
Waters
Associates, -
Gemini C18
HwelettPackard 1100
series,
Photodiode
SC
Chai et al. (2005)
Water-acetic
acid/methanolacetonitrile-acetic acid
Acetonile
/0.1%phosphoric acid
n-buthanol fraction yield:

11.761 mg/g extract.
Chan et al. (2013)
Escin Ia of 17.3 mg/g was the most abundant

saponin in the seeds of Aesculus chinensis
Bunge.
Chen et al. (2007a)
0.1%formic acid
solution/acetonitrile
Total of 17 types of saponins detected: 1278

g/ml.
Cheng et al. (2011)
Tribulus
terrestris L.
Eurospher 100 C18
0.025% acetic acidacetonitrile
The results revealed distinct dierences in the

content of these compounds depending on
region of sample collection, plant part studied
and stage of plant development.
The samples from Bulgaria, Turkey, Greece,
Serbia, Macedonia, Georgia and Iran exhibited
similar chemical prole and only some
quantitative dierence in the content with
protodioscin and prototribestin as main
components.
Dinchev et al.
(2008)
Ipomoea batatas
tubers
Hewlett-Packard HP5
n-BuOH/HOAc/H2O
HPLC-DAD:
Saponin1: 161.20 mg/100g dw;
Saponin2: 14.67 mg/100g dw
HPLC-MS:
Dini et al. (2009)
MA
PT
ED
Gynostemma
pentaphyllum
Sinochrom ODS-BP
C18
CE
Aesculus
chinensis
Avula et al. (2011)
53.78 mg/g of dipsacoside B was found in

L.hypoglauca.
45.65 mg/g, 46.22 mg/g, and 41.22 mg/g of
macranthoidin B were found in L.confusa,
L.macranthoides, and L.similes, respectively.
AC
Defatted rice
bran
Reference
RI
The quantities of saponins were found in the

range of 5 25 g/ml.
NU
Flos Lonicerae
Saponins yields
PT
Source
84
ACCEPTED MANUSCRIPT
array
Saponin1:163.78 mg/100g dw;

Saponin2: 15.60 mg/100g dw
Waters
Alliance, 996
photodiode
array
Luna C8(2)
Water-methanol
Seeds of
Vaccaria
segetalis Garcke,
Saponaria
Vaccaria L.,
Vaccaria
pyramidate
Phasedus
vulgaris L.
Agilent, UVVis diode array
Zorbax SB-C18
Acetonitrile-waterformic acid/acetonitrileformic acid
Agilent 110,
diode array
Zorbax SB-Aq
Ganzera et al.
(2004)
PT
ED
MA
NU
SC
RI
PT
Bacopa monnieri
Trifluoroacetic acid
solution/acetonitrile
CE
Platycodi Radix
Waters
Alliance,
photodiode
array
Hitachi L-6200,
-
Gl-ntnda et
al. (2007)
Hilum contained the highest concentration of

saponins compared to cotyledons or seed coats.
The saponins concentration in hilum increased
Guajardo-Flores et
al (2012)
2.3-fold after soaking from 455.95 mg/100g to

1063.62 mg/100g.
After the rst day of germination, the saponin
concentration in sprouts and cotyledons
AC
Ziziphus jujuba,
Z.jujuba var.
spinosa
The highest saponin yield of 42.2g/100g extract

was obtained by 80% methanol using
accelerated solvent extraction.
increased 1.9 and 2.1-fold, respectively.

Additional germination days decreased the
amount of the most abundant soyasaponins in
black bean sprouts.
Sunfire C18
Acetonitrile/0.2%
acetic acid
Zizyphus saponins I: 1.41 mg/g

Zizyphus saponins I: 4.52 mg/g
Guo et al. (2011)
Zorbax SB-Aq C18
Water/acetonitrile
Platycodin D: 2431 g/g

Polygalacin D: 2116 g/g
Platycoside E: 1088 g/g
Ha et al. (2006)
85
ACCEPTED MANUSCRIPT
Deapi-platycoside E: 744 g/g

Deapi-platycodin D: 738 g/g
J.T Baker C18
Water-acetonitrile
Total saponins (ng/mg/dw):

Root: 5924
Stem: 417
Leaf: 1064
Seedpod: 30.8
Seed: 991
Huhman et al.
(2005)
Panicum
virgatum L.
-, Thermo
Finnigan LCQ
ion trap mass
spectrometer
Betasil C-8
1%formic acidacetonitrile
Total saponin concentrations were highest in

leaves, lowest in the stems, and intermediate
for the combined leaf and stem in all four
varieties of Switchgrass.
Lee et al. (2009)
Dioscorea
zingiberensis
Shimadzu,
Photodiode
array
Shim-pack VP-ODS
C18
Water/acetonitrile
The content ranges (g/100 g) were:

22.985844.5988 (parvioside),
33.861257.7133 (protodeltonin),
0.16921.1125 (protodioscin),
1.99794.2469 (protogracillin),
4.291211.6437 (zingiberensis saponin),
7.028920.4885 (deltonin),
0.37831.3685 (dioscin),
0.15072.0500 (diosgenin diglucoside)
0.11171.1037 (trillin)
Li et al. (2010a)
Gleditsia
sinensis Lam.
Agilent 1100,
UV
ODS-2 Hypersil
Acetonitrile-0.1%
acetic acid/water-0.1%
acetic acid
Total of 19 saponins (g/g %): 4.137-11.479.
Lian & Zhang

(2013)
Xanthoceros
sorbifolia
Shimadzu 2010
series, -
Diamonsil C18
Methanol/0.05% formic
acid
Yields of sorbifoside A and B (g/g):

Husks: 157.0; 58.33
Twig bark: 9.84; 30.14
Twig xylem: tr; 17.84
Seed coats: -; Seed kernels: -; 26.44
Flowers: 21.53; Leaves: -; -
Ling et al. (2011)
Paris and
Agilent 1200,
Kromasil RP-C18
Water/acetonitrile
Saponins of Paris VI was found majorly in
Man et al. (2010b)
PT
-, photodiode
array
AC
CE
PT
ED
MA
NU
SC
RI
Medicago
truncatula
86
ACCEPTED MANUSCRIPT
Paris fargesii (19.520 mg/g), and Trillium

tschonoskii (11.770 mg/g).
Allium nigrum L.
-, Diode array
AQUASIL SS-1251120
0.005% trifluroacetic
acid buffer (TFA)
Folium ginseng,
Radix ginseng
Prevail C18
Water-acetonitrile
Yucca gloriosa
L.
Agilent Series
1200, Diode
array
Agilent 1100,
UV
Waters Atlantis C18
Maesa
lanceolata
Agilent 1100,
Diode array
300 monomeric
C18
Asparagus
Waters Alliance,
Mediterranea Sea18
Among different plant organs, the highest

methiin content was detected in the bulb (0.17
mg/g fw), whereas a trace amount of methiin
was detected in the leaf (0.10 mg/g fw) and root
(0.08 mg/g fw).
PT
ELSD
Mostafa et al.
(2013)
SC
RI
Trillium plants
Qian et al. (2009)
Water-0.05%
triflouroacetic
acid/acetonitrile-0.05%
triflouroacetic acid
The dried extract obtained more than 25% w/w

(236.81 mg/g) of steroidal glycosides saponins.
Skhirtladze et al.
(2011)
0.05%HCO2H/CH3CN0.05%HCO2H
The greenhouse grown M.lanceolata has been

reported the highest percentage of 4.9% in total
saponins.
Theunis et al.
(2007)
Water-1%formic
acid/acetonitrile1%formic acid
Green spears contain from 0.024 mg/100g fw at

the top of the spears to 2.5 mg/100g fw at the
bottom cuts.
White spears content ranged from 1.4 to 5
mg/100g fw depending on the cultivar and
portion of the spear.
The total content of saponin detected in Hutor
asparagus ranged from 1.09 to 2.73 mg/100g
fw.
Vzquez-Castilla
et al. (2013)
AC
CE
diode array
PT
ED
MA
NU
Saponins content was found in the range of

59.14-111.55 mg/g for both ginseng varieties.
Polygala
japonica
Shimadzu, ELSD
Discovery C18
Acetonitrilemethanol/0.05%
trifluoroacetic acid
Total saponins found : 4.45-27.32 mg/g
Wang et al. (2007)
Panax
notoginseng
Dynamic
Extractions
Spectrum,
photodiode
Water SunfireTM C18
Acetonitrile/0.05%
phosphoric acid in
water
notoginsenoside R6: 0.78 mg/g

ginsenoside R1: 2.23 mg/g
ginsenoside Rb1: 1.86 mg/g
notoginsenoside Spt1: 0.83 mg/g
ginsenoside F4: 1.23 mg/g
Zhang et al.
(2013b)
87
ACCEPTED MANUSCRIPT
ginsenoside Rh4: 1.46 mg/g

ginsenoside 20S-Rg3: 2.47 mg/g
ginsenoside 20S-Rs3: 2.22 mg/g
ginsenoside Rk1: 1.84 mg/g
Water/acetonitrile
Agilent 1100,
UV
Zorbax SB C18
Acetonitrile/0.001%
formic acid
Agilent 1100,
ELSD
Kromasil RP-C18
The contents of ginsenosides Rg1, Re, Rb1, Rd,

and notoginsenoside R1 in most of the raw
samples were found to be higher than those in
the corresponding steamed samples.
Lau et al. (2003)
Content of saponins (%):

Notoginsenoside R1: 0.56-2.47
Ginsenoside Rg1: 2.15-5.00
Ginsenoside Rb1: 2.10-4.51
Ginsenoside Rg2: 0.05-0.31
Ginsenoside Rh1: nd-0.10
Ginsenoside Rd: 0.27-1.11
Li et al. (2005)
Total saponins (mg/g) found for:

Paris polyphylla var. yunnanensis=19.62
Paris polyphylla var. chinensis=1.63
Zhang et al. (2010)
RI
Waters Symmetry
C18
CE
Water-0.1% formic
acid/acetonitrile
AC
Paris polyphylla
var. yunnanensis,
Paris polyphylla
var. chinensis
PT
ED
MA
NU
SC
Agilent 1100,
photodiode
array
PT
array
88
ACCEPTED MANUSCRIPT
Highlights
PT
RI
SC
NU
MA
D
TE
Plant materials contain saponins and its pharmaceutical properties are highlighted.
Presented an overview of extraction techniques employed in different field of research
focus.
Review on conventional and green extraction techniques used in saponins extraction.
Spectrophotometric and chromatographic quantification methods for saponins are
synthesized.
AC
CE
P
89

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Food Research International

Choon Yoong Cheok, Hanaa Abdel Karim Salman, Rabiha Sulaiman*

Faculty of Food Science and Technology

Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia

* Corresponding author: Tel.: +60-3-89468520; Fax: +60-3-89423552;

Email address: rabiha@upm.edu.my

to design experiment to suit their particular objective in swift.

Keywords: Saponins; conventional extraction; green extraction technologies; quantification;

Plant materials contain saponins ........................................................................................................... 6

Pharmaceutical properties of saponins .................................................................................................. 7

Applications of saponins in foods ....................................................................................................... 10

Extraction techniques .......................................................................................................................... 12

Conventional extraction techniques ............................................................................................ 14

Green extraction technologies .................................................................................................... 16

Spectrophotometric method ........................................................................................................ 20

Chromatograhic method ............................................................................................................. 23

plant kingdom is reviewed in detail by Vincken et al. (2007).

anti-inflammatory, antifungal or antiyeast, antibacterial or antimicrobial, antiparasitic, antitumor,

triterpene saponins in prevention and treatment of metabolic and vascular disease.

provide a quick reference in future experimental design.

2. Plant materials contain saponins

the basis of EI-MS (electrospray ionization-mass spectra), 1H and

C NMR (nuclear magnetic

3. Pharmaceutical properties of saponins

adjuvants to increase specific immune responses (Estrada et al., 2000).

meal exhibited antibacterial activities against Staphylococcus aureus, Salmonella Typhimurium

antioxidant (Silva et al., 2013).

structure-function of saponins influenced the antitumor mechanism. Saponins derived from

property and suggested to use in the treatment of diabetes.

4. Applications of saponins in foods

beverage applications (Yang & McClements, 2013).

Based on the importance of saponins as pharmaceutical properties especially in countering

Conventional extraction techniques

and efficacy in obtaining significant saponins yields.

5.1.1. Maceration extraction

The maceration extraction is a solid-liquid extraction where the bioactive compound

2012) to shorten the extraction time.

5.1.2. Reflux and Soxhlet extractions

5.1.3. Subsequent extraction

Green extraction technologies

Ngunge, Finnie, & Staden, 2011).

5.2.1. Ultrasound-assisted extraction (UAE)

The phenomenon of ultrasound in creating cavitation bubbles in the solvent by acting as a

5.2.2. Microwave-assisted extraction (MAE)

5.2.3. Accelerated solvent extraction (ASE)

precious value of the product.

spectrophotometric and chromatographic methods. The difference quantitative expression

chromatographics quantifies specific saponins compound.

The spectrophotometric technique has become a popular method in the quantification of

The procedure to quantify total steroidal sapogenin is by weighing 0-40 g of crude

Haemolytic method is another spectrophotometric method uses to quantify saponins

measurable colour for spectrophotometer. The saponins concentrations in bittergourd varieties

concentrations (0.25%) compared to green varieties (0.67%).

antiinammatory and antiulcerogenic properties from the bulbs of Allium ampeloprasum

var. porrum. Fitoterapia, 82, 1175-1180.

Food Chemistry, 47, 3193-3196.

Research International, 43, 1710-1718.

and evolution of triterpenoid saponins. Phytochemistry, 72, 435-457.

methods for determination of magnoorine and saponins from roots of Caulophyllum

and Biomedical Analysis, 56, 895903.

Fitoterapia, 83, 234-240.

Medicago arabica L. Journal of Agricultural and Food Chemistry, 52, 1095-1099.