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Chinese Vinegar and its Solid-State Fermentation Process

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 FOOD REVIEWS INTERNATIONAL
Vol. 20, No. 4, pp. 407–424, 2004

Chinese Vinegar and its Solid-State

Fermentation Process

Dengru Liu,1,* Yang Zhu,2 Rik Beeftink,1 Lydia Ooijkaas,1

Arjen Rinzema,1 Jian Chen,3 and Johannes Tramper1
1
Food and Bioprocess Engineering, Wageningen University and Research Centre,
Wageningen, The Netherlands
2
Applied Plant Research, Wageningen University and Research Centre, Horst,
The Netherlands
3
Southern Yangtze University, School of Biotechnology, Wuxi, China

ABSTRACT

China uses solid-state fermentation (SSF) processes on a large scale for products
such as vinegar, Chinese distilled spirit, soy sauce, Furu, and other national foods
that are consumed around the world. In this article, the typical SSF process is
discussed, with a focus on Chinese vinegars, especially those that are prepared
through solid-state fermentation. Six well-known types are discussed in detail.
Finally, possible ways to improve the traditional vinegar production process are
discussed. The article discloses Chinese information about solid-state vinegar
fermentation that otherwise would be inaccessible to Western scientists due to
language barriers.

Key Words: Chinese vinegar; Solid-state fermentation; Mixed culture;

Anaerobic and aerobic fermentation.

*Correspondence: Dengru Liu, Food and Bioprocess Engineering, Wageningen University

and Research Centre, P.O. Box 8129, 6700, EV Wageningen, The Netherlands; E-mail:
dengru.liu@wur.nl.

407

DOI: 10.1081/LFRI-200033460 8755-9129 (Print); 1525-6103 (Online)

Copyright & 2004 by Marcel Dekker, Inc. www.dekker.com
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408 Liu et al.

INTRODUCTION

Solid-state fermentation (SSF) refers to the growth of microorganisms on moist

solid substrate in the absence of free-flowing water (Moo-Yong et al., 1983). SSF
may be more suitable and practical than submerged fermentation (SmF) for low-
technology applications because (1) cheap unrefined agricultural products are used
as substrates; (2) capital investment and operating cost are moderate; and (3) aseptic
processing is less stringent. Other advantages and disadvantages of SSF over SmF
have been discussed extensively (Mitchell and Lonsane, 1992; Murthy et al., 1993;
Robinson et al., 2001).
During the last two decades, SSF has attracted interest from scientists and
industries due to its potential for the production of food and pharmaceuticals, but
large-scale application of SSF in Western countries is still limited because of scale-up
problems (Nagel, 2002). In Asian countries, however, SSF has been playing an
important role in food and beverage preparation for thousands of years. In China
alone, annual amounts of 4.0  109 kg of Chinese soy sauce, of 2.0  109 kg of
Chinese vinegar, and of 8.0  109 kg of Chinese distilled spirits were produced; more
than 80% of these materials were produced by SSF (Wei, 2001). Other important
SSF products include Chinese sufu, lobster sauce, Chinese distilled spirit, and
mushrooms.
Vinegar is consumed worldwide as a food condiment and preservative, especially
in the Chinese diet. Chinese vinegar has a history of more than 3000 years (Shi, 1999).
There are at least 14 main types on the market, including Zhenjiang aromatic
vinegar, Shanxi old mature vinegar, Jiangzhe Rose vinegar, and Sichuan bran
vinegar, whereas are all produced by SSF (Huang, 1998). Traditional Chinese vinegar
is made from different sorts of cereals, where as in more recent years many vinegars
prepared from fruits have appeared on the market. This article does not discuss fruit
vinegars because these are usually prepared by liquid fermentation.
The SSF process for Chinese vinegar comprises four successive stages, namely
koji preparation, saccharification of starch and alcohol fermentation, acetic-acid
fermentation, and maturation (Huang and Yin, 2000a). Figure 1 is a general
flowchart of Chinese vinegar SSF. Saccharification and alcohol fermentation may
either be SSF or SmF; other stages are always SSF (Huang and Yin, 2000a). The
entire process includes two aerobic fermentations (koji preparation and acetic-acid
fermentation) and an anaerobic fermentation (alcohol fermentation). Fungi, yeasts,
and bacteria are involved in the process; traditionally, all these microorganisms
come from the environment or from the substrate and more or less spontaneously
evolve throughout the process (Lei, 2000).
Although vinegar is produced and consumed worldwide, no publications in
English on vinegar production by SSF processes have appeared in the last 20 years.
A large number of Chinese publications are inaccessible to Western scientists due to
language barriers. Hubert (1976) reviewed vinegar history, but did not include
Chinese and Japanese SSF processes. The Japanese SSF processes are rooted in and
similar to relevant processes in China (Bao, 1988).
This article reviews Chinese vinegar production through SSF. First, the history
of Chinese vinegar production is presented briefly and general characteristics of the
process are discussed, followed by the raw materials and composition of Chinese
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Chinese Vinegar 409

Fungi Koji preparation Koji

Step 1

Raw material treatment Saccharification Glucose solution

Step 2.1

Step 2.2 Yeast culture Alcohol fermentation

Step 3 Acetic-acid bacteria Acetic-acid fermentation

Maturation Chinese vinegar

Step 4

Figure 1. General flowchart of SSF for Chinese vinegar depicting the four steps.

vinegar. Then, six major types of SSF-prepared Chinese vinegars are reviewed.
Finally, opportunities to improve the traditional SSF processes and the possible
implementation of such innovations are discussed.

HISTORY

China probably has the oldest historical records about cereal vinegar in
the world. The book Ceremony Notes (about 800 b.c.) (Xiao, 2000) recorded that
vinegar played an important ceremonial role during the Zhou dynasty (1000 b.c.–
256 b.c.). Also, in The Analects of Confucius (450 b.c.), there are many records on
vinegar (Xiao, 2000). In Techniques of Qinese (533–544 a.d.), 23 methods for vinegar
preparation are described in detail, among which the SSF method with cereals, a
repeated successive fermentation process, was a unique technique for vinegar
preparation (Bao, 1985).
Until the late Qing Dynasty (1644–1911 a.d.), when industrial-scale vinegar
production appeared (Zhang, 2000), Chinese vinegar was only domestically
produced on a small scale. It is generally believed that the production process of
Shanxi old mature vinegar was invented by Wang Laifu during the Qing Dynasty,
between 1644 and 1661 a.d. (Yan, 1997). The fuming technique that characterizes
this process is still used widely by all old mature vinegar manufacturers (Yan, 1997).
The Zhenjiang Henshun vinegar plant was established in 1840 a.d. and still is a major
producer of aromatic rice vinegar, with an annual yield of 60 million kilograms
(Ren and Jian, 2000).
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410 Liu et al.

RAW MATERIALS AND COMPOSITION OF CHINESE VINEGARS

Raw Material

The main raw materials for production of traditional Chinese vinegars are
cereals and their bran (i.e., the outside layers of grains, containing about 50%
starch). In more recent years, however, many other starch and sugar-containing
materials, such as sweet potato and fruits, have found their way into vinegar
production, resulting in new types of vinegar. Table 1 lists the main raw materials
that are used for Chinese vinegars. The type of raw material is the basis for the
specific characteristics of different vinegar types (Zhu, 1991a). Wheat bran is the
main carrier of microorganisms in the SSF system; besides being a source of starch, it
also provides other nutrients, such as proteins, to microorganisms. Rice hull has less
nutritional value. It merely acts as a carrier of microorganisms and spacer to increase
porosity of the substrate mixture (Liu and Li, 1992).

Microorganisms

Microorganisms involved in Chinese vinegar fermentation include fungi, yeasts,

and bacteria. Although the SSF process for Chinese vinegar is not aseptic, specific
fungi, yeasts, and bacteria dominate the koji, alcohol, and acetic-acid fermentations
due to the highly selective materials and operational conditions. Nowadays, some
manufacturers add pure cultures of yeast and acetic acid bacteria, but traditional
Chinese vinegar fermentation is still a spontaneous mixed-culture fermentation
(Lin, 1985).
Aspergillus, Rhizopus, and Monascus are the main fungi in koji preparation;
Saccharomyces cerevisiae and Hansenula anomala are the main yeasts in alcohol

Table 1. Major raw materials for Chinese vinegar production.a

Carbohydrate Protein Lipid Ash

Name (%) (%) (%) (%) Example

Sticky rice 72.1 8.5 3.2 0.9 Zhenjiang vinegar,

Shanghai vinegar
Japonica rice 73.4 8.8 2.2 1.3 Jiangzhe vinegar
Nonsticky rice 74.5 8.2 2.3 1.3 Beijing vinegar
Oat 66.7 15.6 3.2 1.7
Highland barley 70.3 10.1 1.8 3.4
Sorghum 70.0 8.3 5.0 3.0 Shanxi vinegar
Foxtail millet 76.0 9.7 3.3 1.4
Corn 67.2 9.5 5.0 1.3
Sweet potato 76.7 6.1 0.5 2.4
Wheat bran 55.0 14.0 4.0 5.0 Sichuan vinegar
a
Data adapted from Huang and Cai (1999).
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Chinese Vinegar 411

fermentation (Huang and Yin, 2000a). In almost every Chinese vinegar SSF process,
several species of acetic-acid bacteria are present: Acetobacter pasteurianus is
the predominant species in Zhenjiang vinegar, due to the high initial ethanol
concentration (Huang and Cai, 1999). A. aceti dominates in Sichuan bran vinegar
production, whereas in Jiangzhe and Fujian vinegars, A. xylinum is the main
bacterium. A. xylinum forms a pellicle on the surface of the static liquid (Lei, 2000),
which is different from traditional wine vinegar production methods (e.g., the
Orleans method and the Balsamic method) where a foam rather than a pellicle forms
at the surface (Adams, 1985). Other bacteria, such as Gluconobacter and lactic-acid
bacteria, are commonly found as well and their metabolites contribute to the
characteristic flavor and aroma (Lei, 2000).

Composition

The composition of Chinese vinegar has been investigated and reviewed

extensively (Cui, 1986; Huang and Yin, 2000b). A large number of substances are
present in Chinese vinegar, including organic acids, free amino acids, carbohydrates,
alcohols, esters, and various microconstituents. Table 2 outlines the general
composition of seven Chinese vinegars; more detailed data were presented by Cui
(1986). The complicated composition of Chinese vinegars is a reflection of their raw
materials and production processes. Some components (e.g., carbohydrates) were
already present in raw materials, whereas others emerge during the production
process, either (bio-)chemically or microbially.
Because of differences in composition, each type of Chinese vinegar has its own
distinct sensory characteristics. Traditional Chinese vinegar usually has a higher
concentration of organic acids (up to 12%, of which more than 99% is acetic acid)

Table 2. General composition of main Chinese vinegars (g/100 mL).

Rice Fumed Zhenjiang Shanxi Sichuan Jiangzhe

Components vinegar vinegar vinegar vinegar vinegar vinegar

Density 107.2 105.6 108.6 119.4 111.4 106.0

Acids 5.1 8.0 6.8 10.9 7.2 3.6
Solid 12.8 9.7 11.9 30.5 21.4 7.0
Salt 0.02 0.8 3.2 3.4 1.5 7.0
Total carbohydrate 3.9 0.8 1.8 12.8 7.3 3.7
Reduced sugar 2.0 0.6 1.5 11.3 4.5 2.5
Total nitrogen 0.3 0.6 0.7 1.2 1.3 0.2
Amino acids 0.7 1.7 1.1 2.3 2.6 0.4
Total aroma 0.5 0.6 0.7 1.1 0.7 0.4
volatile substance
Main aroma Propyl i-BuOH, i-PrOH 2,3-BuDiOH Furfural i-BuOH
components acetate furfural

Data adapted from Cui (1986) and Tang (1985).

i-PrOH, iso-propanol; i-BuOH, iso-butanol; 2,3-BuDiOH, 2,3-butanediol.
 ORDER REPRINTS

412 Liu et al.

than common vinegar. Xia et al. (1985) made a dynamic analysis of the formation of
main components in Chinese vinegar during SSF or traditional SmF. They showed
that during SSF, different components are formed continuously and simultaneously,
whereas in SmF, components are formed more sequentially. The traditional
SmF, however, is very different from modern SmF, as it uses mixed cultures like
in SSF; the quality of the traditional product is regarded as superior to modern
SmF vinegar (Liu, 1982). Similar traditional SmF processes are used in Europe to
produce high-quality wine vinegar (Tesfaye et al., 2002).

SOLID-STATE FERMENTATION OF CHINESE VINEGAR

Traditional production processes of Chinese vinegar are typically empirical and

craftlike, rather than scientific. Generally, SSF for Chinese vinegar includes four
stages: koji preparation, saccharification and alcohol fermentation, acetic-acid
formation, and maturation (Huang and Yin, 2000b).
In the past, processes were developed by trial and error, and handed down by
mouth and hands of the veterans in the plants. A great body of experience and
practical knowledge accumulated over time, but the underlying microbiological and
biochemical mechanisms were not investigated systematically. Consequently, many
different production processes exist and significant improvements have been scarce.
There are two basic SSF processes in Chinese vinegar production, based on the
treatment method of the raw material, namely the ‘‘cook method’’ and the ‘‘steam
method’’ (Huang and Cai, 1999). In the ‘‘cook method,’’ saccharification and
alcohol fermentation occur in the liquid state (i.e., SmF; Fig. 2a), whereas the acetic-
acid fermentation is SSF. In the ‘‘steam method,’’ the whole process takes place in
the solid state (SSF; Fig. 2b), where rice hull and part of the wheat bran are mixed
with the main raw material (grain) immediately after soaking and are steamed
together.

Koji Preparation

Koji may be regarded as an enzyme-containing starch material for starch

hydrolysis. Its preparation method is ancient and originated in China. Later, it found
its way to Japan and other Southeast Asian countries (Fukashima, 1989). In China,
koji is widely used in the production of traditional distilled spirit (Bai Jiu, in
Chinese), soy sauce, and vinegar. Historically, koji is a solid fermentative mixture of
substrate and microbial metabolites, especially saccharolytic enzymes. Generally,
koji is classified according to its preparation method: great koji, mini koji, bran koji,
wheat koji, herb koji, and red-rice koji. Herb koji is a special koji only used in
Sichuan vinegar production (see ‘‘Major Types of Chinese Vinegar’’). The dominant
microorganisms in koji are various moulds, such as Aspergillus, Rhizopus, Mucor,
and Penicillium (Huang and Yin, 2000a). Different types of koji have a different
microbial flora. The microorganisms in great koji, mini koji, herb koji, and red-rice
koji spontaneously originate from the natural environment, whereas bran koji and
wheat koji are usually based on cultures of Aspergillus or Rhizopus.
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Chinese Vinegar 413

sticky
rice soak cook cool mix

water koji,
yeast

saccharification acetic-acid
alcohol fermentation fermentation
(liquid) rice hull (solid)
wheat bran
parent Pei

maturation leach pasteurization vinegar

salt
Materials kg Products kg
Sticky rice 500 Aromatic vinegar
Koji 30 (6.5% acetate) 1450
Wheat bran 850 waste 2625
Rice hull 475
Water 2200
Salt 20

sorghum
grind soak mix steam

rice hull
wheat bran

saccharification acetic-acid
cool mix alcohol fermentation fermentation
(solid) (solid)
wheat bran
yeast, water
parent Pei

fuming leach maturation old

vinegar
salt fumed vinegar

Materials kg Product kg
Sorghum 500 Fumed vinegar
Wheat bran 350 (6% acetate) 400
Rice hull 350 or
Koji 300 Old mature vinegar
Water 2400 (>10% acetate) 120
Salt 25

B
Figure 2. Flowchart of solid-state fermentation of Chinese vinegar. A, Cook method
(Zhenjiang aromatic vinegar); B, steam method (Shanxi old mature vinegar). (Data from
Huang and Cai (1999).)
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414 Liu et al.

Barley 70% Grind Blend & Koji cakes

Pea 30% & mix configurate

Water

Pile in line Get moldy Aeration Fermentation

(2–4 d) (12 h) (12–15 d)

Maturation Great koji

(2 d)
Figure 3. Typical flowchart of preparation of Chinese great koji. Koji cakes are piled in line
in the ‘‘koji house.’’ At different stages, temperature is controlled by adjusting the spacing
among the cakes and opening or closing windows. The period indicated in the bracket may
vary a little between winter and summer. For more details, see text.

Great koji (Da Qu, in Chinese) and mini koji (Xiao Qu) are the most
traditional koji. Figure 3 is a chart of the typical preparation of Chinese great koji,
which was used to produce traditionally distilled spirit and vinegar (Huang and
Cai, 1999).
Every step of the process schematized in Fig. 3 must be carried out carefully to
ensure high-quality koji (Wang, 1999), and practical experience and hands-on
knowledge are very important. The fermentation actually consists of three stages,
each at a different temperature. As a result, specific fungi proliferate (Wang, 1997).
These different temperatures are obtained by controlling the heat transfer among the
cakes (by adjusting the spacing) and the heat transfer from the koji room to the
environment (by closing or opening the windows). The important processes are
metabolic heat production, heat convection, and water evaporation. Finally, most of
the water in the great koji evaporates and most microorganisms lose viability. Only
temperature-tolerant fungal enzymes remain and play an important role in the
subsequent saccharification (Wang, 1995).
The preparation of mini koji is similar to that of great koji, with the dominant
microorganism being Rhizopus rather than Aspergillus (Huang and Yin, 2000a).
Other kinds of koji are much simpler to prepare. Instead of great koji, more and
more companies use wheat koji, which is prepared from a pure culture of Aspergillus
sp. and wheat. Bran koji is another pure-culture product. The preparation of
pure-culture koji and the relevant reactors have been described elsewhere (Kitamoto,
2002). The preparation of koji is laborious and time consuming. Some
manufacturers tried to replace the saccharifying action of koji with enzyme
mixtures, but found that the quality of their products was negatively affected
(Hou et al., 1996).
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Chinese Vinegar 415

Saccharification and Alcohol Fermentation

In traditional processes, alcohol fermentation starts as soon as liquefaction and

saccharification of starch are under way (Hou et al., 1996). The process is initiated
by addition of koji and cultured yeast to the starch material. Many manufacturers
do not use cultured yeast, but rely on yeast that comes spontaneously from the
environment or is indigenous to the koji (Lei, 2000). Some manufacturers also use
fermented broth from a previous successful batch as inoculum to initiate alcohol
fermentation (repeated fermentation or ‘‘back slopping’’).
However, a few manufacturers, such as the Shanghai rice vinegar plant, replace
koji with enzyme mixtures. As a result, the starch material is liquefied and then
saccharified completely before alcohol fermentation is initiated by yeast addition
(Chen and Chen, 1996).
In both the cook method and the steam method, the temperature during
alcohol fermentation must be kept below 28 C to minimize evaporation of ethanol.
The average duration of saccharification and alcohol fermentation is 10 days
(Huang and Cai, 1999).

Acetic-Acid Fermentation

In the cook method, the fermented broth or solids from the alcohol fermentation
are mixed with wheat bran and vinegar seeds (i.e., parent vinegar Pei from the last
batch of acetic-acid fermentation or, as an alternative, an inoculum of acetic-acid-
producing bacteria such as Acetobacter). For the steam method, wheat bran and rice
hull are mixed with the main starch materials before alcohol fermentation, so only
vinegar seeds need to be added after alcohol fermentation. Wheat bran is used as a
substrate and as a carrier for the bacteria, whereas rice hull is used both as a carrier
of the bacteria and a spacer increasing porosity, thus increasing the exchange area
and promoting oxygen uptake and heat discharge. The ratio of the main substrate to
the bran and rice hull is generally 1 :1.2 : 1.4 (Liu and Li, 1992). The manufacturers of
Sichuan bran vinegar use pure wheat bran as the only solid substrate.
The traditional acid fermentor is a clay jar (about 0.5 m3 capacity). Nowadays,
many manufacturers replace it with a concrete basin (about 8 m3), which in fact is a
deep-bed tray system with intermittent mixing. Rice hull is fed to the fermentor
during mixing. Compared with the thin-layer tray system, the surface area per unit
volume in deep-bed tray system is reduced and consequently, evaporation of ethanol
is reduced. At the same time, intermittent mixing and rice-hull feeding facilitate
oxygen diffusion. The most important variable at this stage is the temperature, which
is controlled by stirring and turning the substrate (Liu, 1982; Zhu, 1991b). If there
is sufficient oxygen, bacteria will propagate and metabolize rapidly and produce a
large amount of metabolic heat. When the temperature becomes too high, the
operator will ‘‘turn up’’ or mix, the substrate to disperse heat, and then press the
surface to reduce oxygen supply, thereby decreasing the rate of ethanol consumption
and heat production (Yu, 1985). The temperature is usually kept at 38 C to 40 C.
Generally, stirring and turning up the substrate is carried out once per day
(Guo, 2000). Some manufacturers use concrete basins with two bottoms (the upper
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416 Liu et al.

bottom is perforated, a so-called ‘‘false bottom’’) to collect the vinegar liquid,

which is then intermittently recycled onto the top surface of the solid substrate to
regulate the temperature (Xing, 1991). The recycled broth is used as a cooling agent
(in summer) or heating agent (in winter). Even so, the solid substrate needs to be
stirred and turned up at least three times during the whole period because the
substrate tends to agglomerate and clog after being showered with the recycling
broth.

Maturation

Most of the traditional production processes of Chinese vinegar feature a

maturation period (Zhu, 1992b) in which many flavor substances, such as esters, are
formed by chemical reactions. Its length varies with the type of vinegar. During
maturation, microbial activity comes to a halt. Salt is usually added to the maturing
vinegar to prevent microbial overoxidation of acetic acid to carbon dioxide.
Storage is common for maturation, but the exact method varies. For example,
manufacturers of Zhenjiang aromatic vinegar usually mature their vinegar by adding
salt and sealing the surface of the fermented solid substrate with clay; thus, the
maturation process is anaerobic (Guo, 2000). The manufacturers of Shanxi old
mature vinegar, however, use solarization (in the summer) and ice removal (in the
winter) to mature and to concentrate the vinegar broth; thus, the course is mainly
aerobic. In addition, this method brings about a concentration of the acetic acid
because water is removed as ice and water vapor.

MAJOR TYPES OF CHINESE VINEGAR

In the former National Industrial Standard of Vinegar, vinegar is classified into

three grades, depending on its concentration of acetic acid (3.5–4.5%, 4.5–6%, and
>6%, respectively) (Wei, 2001). More recently, a new National Standard Code of
Condiments was issued by the Chinese State Administration Bureau for Quality and
Technology, in which vinegar is classified as either brewed or formulated (acetic-acid
blended with other ingredients, such as flavors) (Qiao, 1999). Besides, each major
vinegar also has its own local quality criteria and grading system. Generally, brewed
vinegars are more popular on the Chinese market, especially those that are
prepared by SSF. In this article, only brewed vinegar is referred to as Chinese vinegar
(Table 3).
Chinese vinegars have specific local features. Every region has its own manufac-
turers, who produce vinegar in specific processes, using particular raw materials.
Consequently, each vinegar has its own taste, flavor characteristic, and market.
There are at least 14 types of traditional Chinese vinegar on the market. Most of
them are produced by SSF, but published data are quite scarce (Huang, 1998). Here,
we only discuss six well-known types of brewed vinegar.
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Chinese Vinegar 417

Table 3. The main types of Chinese vinegar.

Main Sticky Sticky Sticky Wheat Rice

materials rice rice rice Sorghum bran
Saccharifying Enzyme Red koji Wheat koji Great koji Herb koji No
agent
Process type SSF LSF SSF SSF SSF LSF
Production 100 >1000 60 >500 >400 160
period (days)
Annual 3.0  105 6.0  107 2.1  108
yield (kg)
Refs. Hang and Huang and Hang and Huang and Hang and Hang and
Cai, 1999 Yin, 2000a; Cai, 1999; Yin, 2000a; Cai, 1999; Cai, 1999
Liu, 1982 Ren and Wang, 1997 Li, 1999
Jian, 2000

Shanxi Old Mature Vinegar

Shanxi old mature vinegar is the most famous vinegar in northern China. It has
a history of more than 300 years. In the province of Shanxi, there are more than 1000
vinegar manufacturers that produce mature vinegar, fumed vinegar, and old mature
vinegar. Only 120 producers sell their products outside the province (Wang, 1997).
Shanxi old vinegars use sorghum as the main raw material with a very large
dosage of great koji (about 60% of the raw materials) (Huang and Cai, 1999).
Production of Shanxi old mature vinegar takes about 18 months, although the acid
production itself takes less than 10 days. The most time-consuming stage is the
maturation process. Upon fermentation, half of the solid Pei is ‘‘fumed’’ (heated in a
jar with a lid at 70 C for 4 days) and then mixed with the remaining half and leached.
Flavor compounds are formed chemically during fuming. The filtrate is transferred
to a big jar and is exposed to the sun (solarized); in winter, surface ice is removed.
Solarization and ice removal entail a concentration increase of acetic acid and flavor
substances formed by chemical and enzymatic reactions.
Preparation of great koji is also lengthy, taking more than 1 month. One of the
major challenges is to improve the process in such a way that the total production
time is significantly reduced while preserving the typical flavor style of old mature
vinegar (Yan, 1997). Replacing part of the great koji with wheat bran and applying
new maturation methods (e.g., infrared vacuum concentration instead of natural
evaporation and maturation), together with modern formulation methods, might
increase productivity and consistency of quality greatly (Wang, 1997).

Zhenjiang Aromatic Vinegar

Zhenjiang aromatic vinegar is most famous in Southern China. Its main raw
materials are sticky rice and wheat koji. Henshun Group Co., Ltd., is the largest
manufacturer and its product, KingsanÕ vinegar, is sold in more than 43 countries
 ORDER REPRINTS

418 Liu et al.

(Ren and Jian, 2000). Zhenjiang aromatic vinegar is a typical example of the
so-called ‘‘cook method’’ for ethanol production. However, the acetic-acid stage is a
unique multilayer SSF with a fed-batch fermentation process in a series of two open
containers (big jar or basin). First, a certain amount of rice wine (about 14%
ethanol, v/v) is mixed with wheat bran, rice hull, and so-called vinegar seeds (i.e.,
starter culture) to form a semi-solid substrate in the first container (half volume).
The substrate is divided in 10 layers; every 24 h, the top layer is mixed with fresh rice
hull to increase porosity and transferred to the second container; after 10 days, the
first container is thus empty and the second one full (Chen and Chen, 1996). This
process is called ‘‘fed-batch fermentation by layers’’ (Guo, 2000). Ethanol oxidation
and acetic-acid formation mainly take place in the top layer of the second container,
which is exposed to the air. In the old days, feeding and mixing was done manually.
In more recent years, technicians of the Henshun plant modified the process by
introducing a concrete basin with mechanical mixing, thus reducing manual
labor input. As a result, productivity was greatly increased and the vinegar quality
remained unaffected (Zhou, 1990). These changes made Henshun the largest vinegar
producer in China; they changed the view that a traditional process could not be
altered without affecting the quality of the product.

Sichuan Bran Vinegar

Sichuan is a hilly province in Southwest China. Its climate, mild all year
round and misty in autumn and winter, is favorable for specific microorganisms
that are a basis of traditional SSF. Chinese spirit (Bai Jiu, in Chinese) and
bran vinegar are the main fermentation products in Sichuan. Both are produced
by SSF, and both are based on mixed cultures of natural origin. BaoningÕ vinegar
and ZigengÕ sun vinegar are the most famous Sichuan bran vinegars (Feng et al.,
2000).
The raw materials for Sichuan bran vinegar production process are herb koji
(including as many as 108 medicinal herbs), which is the liquid extract of smartweed
leaves (a marsh plant, Polygonum hydropiper, Laliao in Chinese) used to initiate
vinegar fermentation; and wheat bran, used both as carrier and substrate in the
fermentation process. Production of herb koji is rather selective for specific
microorganisms. Unlike most other processes, rice hull is not used in bran vinegar
production and neither is parent Pei (fermented bran as vinegar seed) (Li, 1999).
Instead, a special ‘‘vinegar mother’’ is prepared from steamed rice, herb koji, and a
liquid extract of the special smartweed. This mixture is fermented for 7 days, with
intermittent mixing of the semi-liquid broth to provide oxygen for propagation
(Huang and Yin, 2000a). The ‘‘vinegar mother’’ is then mixed with wheat bran to
finish the acetic acid fermentation. Enzymes, yeast, and bacteria from the ‘‘vinegar
mother’’ convert the bran starch into ethanol, and finally, acetic-acid. The 14-day
fermentation is a combined alcohol and acetic-acid fermentation. When the
fermentations finished, the vinegar Pei is stored in a closed jar for maturation
(about 1 year) before extraction.
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Chinese Vinegar 419

Shanghai Rice Vinegar

Shanghai rice vinegar is a relatively modern type of vinegar. Its producers

adopted modern techniques for vinegar production to increase productivity, decrease
costs, and improve product appearance. As a result, Shanghai rice vinegar now has a
large market share (Lin, 1985; Wang, 1997).
Shanghai rice vinegar is generally produced by SSF. However, the traditional
koji is replaced by amylase, and manual mixing is replaced by mechanical circulation
of the vinegar solution (Huang and Yin, 2000a; Zhu, 1991b). The maturation period
is relatively short due to infrared heating (Dong and Tian, 1990). Although Shanghai
rice vinegar shares its basic characteristics with other SSF vinegars, its taste and
flavor are not as good as more traditional vinegars.

Jiangzhe Rose Vinegar

The name Jiangzhe is a combination of the names of two Chinese provinces

(i.e., Jiangsu and Zhejiang). Production of Jiangzhe rose vinegar is a combination of
SSF and SmF. First, microorganisms grow spontaneously on steamed rice. After
about 10 days, pigmented microorganisms (mainly Monascus) predominate (‘‘rice
blossoming’’), performing a simultaneous saccharification and alcohol fermentation
(Huang and Cai, 1999). Water is then added to ‘‘blossoming rice,’’ and within 20
days, acid-producing microorganisms, such as Acetobacter, proliferate and form a
bacterial film at the surface. From that moment on, the broth is stirred every 2 days.
It takes another 3 to 4 months to complete the fermentation. The filtrate of the
fermented broth is then bottled and pasteurized without further maturation (Huang
and Cai, 1999).

Fujian Red Rice Vinegar

Fujian red rice vinegar is prepared by combined SSF and SmF. Red koji, in
which Monascus dominates, is used as the saccharifying agent and is produced by
SSF. Production of Fujian red-rice vinegar itself is a special type of repeated fed-
batch SmF (Liu, 1982). During saccharification and alcohol fermentation (about 70
days), water that has been boiled and then cooled is added to the mixture of steamed
sticky rice and red koji in three batches, thus initiating an acid fermentation that lasts
3 years. Acid formation is a static SmF with intermittent manual mixing. Generally,
50% of the three-year-old product is harvested as finished product; into its residue,
50% of two-year-old product is fed. Half of the one-year-old product is then fed into
the residue of the two-year-old batch; the other half is supplemented with a freshly
fermented alcohol solution (Liu, 1982).
Production of Fujian red-rice vinegar takes a long time because no inoculum
is used; all microorganisms arise spontaneously from the environment and it takes
a long time for the target microorganisms to proliferate and become dominant. In
addition, acid production occurs by static fermentation without forced aeration.
Hence, low concentrations of oxygen in the broth prevent rapid oxidation of ethanol.
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420 Liu et al.

RESEARCH ON SSF PROCESS

Solid-state fermentation involves growth of microorganisms on moist solid

substrates in the absence of free-flowing water (Murthy et al., 1993). In the past two
decades, SSF has gained the attention of Western scientists because of its promising
applications for the food and pharmaceutical industries. Many articles on SSF have
been published in the last 10 years, but large-scale application of SSF in Western
industries is still limited due to scale-up problems (Abiose et al., 1982; Aidoo et al.,
1982; Mitchell and Lonsane, 1992).
In China and Southeast Asian countries, however, SSF plays a central role in
indigenous fermented food production. Most Asian SSF processes are maintained
empirically. Such SSF processing cannot be applied to Western products without
modifications because there are too many uncertainties and misunderstandings
about Asian SSF processes. Scientific research is urgently needed to improve and
update the fundamental knowledge on traditional SSF processes.
For traditional vinegar production, Chen (1988), Cheng (1992), Huang (1998)
and Zhu (1991b, 1992a, 1992b) discussed options for process innovation. They
considered it necessary to increase productivity and yield on raw materials, and to
mechanize the whole process, because the processes require large residence times and
are quite laborious. The most important period of the process is the acid formation
stage. At this stage, many other components besides acetic acid must be produced by
the mixed microorganisms; a pure culture cannot fulfill this task. Also, minimizing
ethanol losses by evaporation at this stage is a bottleneck in increasing yield. At the
same time, sufficient oxygen must be supplied for the acetic-acid bacteria and
metabolic heat must be removed to prevent overheating.
The production period cannot be shortened significantly without sufficient insight
in the underlying biological and physical phenomena (Guo, 2000). Until now,
companies tried to supplement with enzymes or pure cultures to speed up the process.
Amylase, for example, is supplemented to accelerate saccharification (Dong, 1993;
Lei, 2000), whereas yeast cultures may be supplemented to speed up alcohol fermenta-
tion and Acetobacter to increase acetic acid productivity (Chen and Chen, 1996). In
some big plants, manual mixing and turning have been replaced by mechanical devices.
The traditional Chinese SSF system represents a unique ecosystem that is
regulated by environmental conditions. Besides microbiological reactions, many
(bio-)chemical and physical phenomena take place in the system. Traditional
Chinese vinegar contains a plethora of components that are produced in a wide
range of reactions. The taste of vinegar is based on the collective sensing of tongue,
mouth, and nose, and is determined by the components and their ratio in the vinegar
(Tesfaye et al., 2002). Until now, no SmF process or formulation technology has
been found that can completely replace the SSF process to produce traditional
Chinese foods and drinks of the same quality (Huang, 1998). However, the full
potential of the traditional SSF process is not yet completely explored due to
insufficient fundamental research (Suo and Zhang, 1989).
To apply the traditional SSF process more effectively, more in-depth research
and precise data are needed. The lab-scale studies carried out more recently by
Western scientists, and especially the scientific thought and methods used in their
studies, could certainly advance the traditional SSF processes. At the same time,
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Chinese Vinegar 421

studying the Chinese traditional SSF processes may provide a foundation for
their large-scale industrial applications. The deep-bed tray system with intermit-
tent mixing, which is used in Chinese vinegar production, may have a potential
application for flavor production by aerobic fungi.
For the Chinese vinegar SSF process, koji preparation most closely resembles
the typical SSF system because it involves fungi. Acetic-acid fermentation involves
acetic-acid bacteria, altering the process a little. Bacteria, unlike fungi, can resist
mixing. So, at the stage of acetic-acid fermentation, the macrogradients of
temperature, nutrients, and product concentrations can be minimized by mixing.
Mixing also helps in dispersing metabolic heat and improves oxygen transfer, but
also speeds up evaporation of ethanol. Further research on the production processes
is required to better understand aspects such as pH development, oxygen diffusion
during nonstirring period, loss of bacterial viability, and loss of volatile components
through evaporation (e.g., ethanol).

CONCLUSION

In China and other Asian countries, solid-state fermentation is an established

and ancient technique to produce indigenous foods. Chinese vinegar is a typical
product of SSF from cereals. Because Chinese restaurants are ubiquitous, Chinese
vinegars are consumed worldwide. This article discussed the characteristics of the
traditional SSF process for Chinese vinegar and six famous Chinese vinegars. The
traditional Chinese SSFs were developed mainly by trial-and-error means and during
this course a great deal of practical knowledge was accumulated. This knowledge
can be very useful for scale-up of new lab-scale SSF processes under investigation.
However, a lot of research work is needed to clarify the uncertainties of the
traditional SSFs and to improve their performance.

NOMENCLATURE

Koji A solid enzyme-rich mixture, produced by solid-state fermen-

tation by fungi and normally used as a starter culture for
starch hydrolysis
Vinegar seed An inoculum obtained from a previous fermentation and
containing high numbers of viable acetic-acid bacteria
Vinegar Pei A moistened solid mixture of alcohol, wheat bran, rice hull,
water and vinegar seeds
LSF Liquid-surface fermentation
SSF Solid-state fermentation
SmF Submerged fermentation

ACKNOWLEDGMENT

This study was funded by TNO Nutrition and Food Research Institute.
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422 Liu et al.

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Received April 2, 2004

Accepted July 19, 2004
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