COMPOUND DETECTION AND TOXICITY EVALUATION OF ORGANIC EXTRACT OF THE ERYTHRINA CORALLOIDES

Castaeda Antonio D1, Rosales Herrera J.M.2, Blas Castillo J.3 Hernndez Aldana F.4, Jimnez Salgado T.1, Tapia Hernndez A.1 1Laboratorio de Microbiologa de Suelos, 2Facultad de Ciencias Qumicas, 3Colegio de Ingeniera Ambiental, 4Centro de Qumica. Benemrita Universidad Autnoma 103J 2. piso Centro en Investigaciones en Ciencias Microbiolgicas. C.U., Jardines de Sn Manuel Puebla, Pue. Tel. 01 222 2295677 castanedaantonio@hotmail.com de Puebla. Ed.

Keywords: Erythrina coralloides, alkaloid, gas chromatography mass spectrometry ABSTRACT Mexico counts with the Erythrina Coralloides whose components contain certain concentration of alkaloids which have poisonous properties and can induce paralysis disorders in the motor nerves either by oral ingestion or intravenous administration, in phytotherapy. In the present research, the acid and alkaline extraction of it`s flower and seed had been made, evaluating the composition of the extracts by gas chromatography/mass spectrometry and their toxicity under the FEUM criterion by systemic injection (MGA-DM 3083) with CD1 mice strains. The mixture obtained contains importance as an agonist: therabaine, oripavine, erysodine whose concentration values are estimated in abundance by the chromatogram obtained. The results shows that the seed is the part which contains a higher concentration of alkaloids followed by the corolla, and finally by the cup. The more efficient extractions were obtained in the acidic compounds, which could be used as precursors of more potent substances to treat pain, like morphine, by other simplified routes. The LD50 obtained is 3.99 mg/ 0.3 mL in mice, with an average weight of 20.70 g, by intravenous administration. The symptoms presented in the study group were paralysis of tail, hind limp prostration, convulsions, spasms and respiratory depression with hypothermia. At concentration of 266.38 mg/ml, the instantaneous death occurs.
INTRODUCTION In Mexico there are medicinal plant's ancestral phytotherapeutic applications allow an alternative treatment to allopathic medicine. However, we find that there are affectations listed parallel to cure the first instance, as in the case of some plants such as Erythrina coralloides whose components contain a certain concentration of alkaloids which are poisonous and paralyzing motor nerves by oral or intravenous intake. This plant has several medicinal uses in the center of the country, in the Distrito Federal is recommended to attend problems of measles and in Morelos is indicates in neuritis, in Puebla to cure kidney pain and is widely used for its properties antitussives, in Michoacn applied latex in snake bite cases and scorpion stings. Therefore it is necessary to identify and understand the components of such plants and to recognize the risk they are exposed.

Table 1 shows chromatography components of interest for phytotherapy

Table 1 components of interest for phytotherapy , corollas, calyx and seed of Erythrina coralloides
COMPOUND SYNONYMOUS CAS 612588 RETENTION TIME (min) 10.91 IDENTIFICATION m/z 143(999),115(389) 142(300),116(128) 144(110) 297(999),42(526) 298(196),296(193) 58 (78) 319(999),229(896) 138(813),162(719) 42(688) 268(999),299(584) 284(522),269(277) 215(169) 242(999),42(610) 171(470),198(420) 199(310)

3 methyl Quinoline sinnimo: 3- 3-methylquinoleine methylquinoleine Morphinan-3-ol, 6,7,8,14- Oripavine tetradehydro-4,5-epoxy-6methoxy-17 methyl (5a) Morphinan,7,8-didehydro-4,5Dimethyl morphine epoxy-3,6-dimethoxy-17-methyl(5 b) Erythrinan-16-ol, 1,2,6,7- Erysodine tetradehydro-3,15-dimethoxy-(3
) 5-phenyl-1,3 diazadadamantan-6- 5-phenyl-1,3 diazadadamantan-6one hydrazone one hydrazone

467049

19.71

2859167

19.82 19.82 19.57 20.37 19.93

7290031

125658039

The compounds found are opioid phenanthrene type to m and m2 receptors, which are : The 3methyl quinoline has not been reported as causal in affected human health and whether as a source of carbon and nitrogen, and energy storage of some bacteria2. Erysodine has been reported as a more potent inhibitor of cytisine binding to receptors 3H neuronal nicotinic acetylcholine1. Oripavine is used as raw material for the synthesis nor-buprenorphine is usually obtained from thebaine, requiring a step of o-dimethylation, common steps for getting started with morphine, thebaine, requiring a step of o-dimethylation. The derived dimethylmorphine opiate is used for its properties antitussive, and to a lesser degree than morphine. Nevertheless is often modified in the liver forming 10% morphine, hence its poor analgesic properties4.. The 5-Phenyl-1,3diazaadamantan-6-one Hydrazone, is reported in the listing of Chinese herbal medicine to prevent preterm birth , 7. It is confirmed that the seed is the part that contains a higher concentration of alkaloids followed by the Corolla, and finally by the cup, obtained greater efficiency in the acid extractions Biological testing : Injection systemic test used alcoholic extract at a pH of 6.3 sterilized at 121 C and 15 lb pressure for 20 minutes to lots of 6 mice CD1 strain , at the concentration of 266.38 mg / ml have died instantly, Are carried out with extract dilutions of 1:20 isotonic saline was injected intravenously (300ml) presenting an LD50 of 3.99 mg of the mixture withdrawn, symptoms are paralysis of tail, hind limb prostration, convulsions, spasms, respiratory depression with hypothermia.
Analysis of Extract
180.00

OBJECTIVE: Recognize the components of organic solvent extraction and gas chromatographic analysis / mass of Erythrina coralloides and assess their toxicity. MATERIAL AND METHODS COLLECTION OF PLANT MATERIAL Erythrina coralloides that work was in the city of Puebla in urban areas, mainly of the species found at coordinates 19 0'5 .06 N and 98 12'2 .97''W in November-December period. ANIMALS 60 mice were used CD1 strain rate from the production unit and laboratory animal experimentation of CINVESTAV. Were handled in accordance with the Guide for the Care and Use of Laboratory Animals as promulgated adopted by the U.S. and National Institutes of Health
y

Figure 1. Biosynthetic scheme with intermediate steps for morphine(J. Nat. Prod. 2005) 5

mg/ml Considered value

160.00 140.00 120.00 100.00 80.00 60.00 40.00 20.00 0.00

Corola acida corola alcalina caliz acido caliz alcalino frijol alcalino frijol acido Corola acida corola alcalina caliz acido

3,4dihydroisoquinoli ne,

Preparation of extracts Extracts were obtained from approximately 300 g of seed and flower corolla divided into the cup, because the commonly consumed are the corollas (Lozoya and Lozoya, 1982) each party in triplicate, the extract was carried out on wet basis to avoid having separate losses and determining the moisture of the ground mixture of flower and seed, were extracted by cold maceration for 5 days, and Branson ultrasonic bath 300 watts of power, with and without maceration with heat at 121 C for 20 minutes in an autoclave, were well alkaline and acid extracts having a precedent that the presence of alkaloids are not affected by pH. The solvents used were water and methyl alcohol GRA. The extracts were collected and concentrated in a rotary evaporator for analysis. Chromatographic analysis:
The extracts obtained with organic compounds of the flower and the seed of Erythrina coralloides were analyzed by gas chromatography HP 6890/column HP-5MS mass spectrometry coupled to Agilent Technologies 5973 Network with the following conditions: 56 C increasing 12 C to 194 C / 1 min, continuous increase of 10 C to 280 C/15 min, injector temperature Split / splittles 250 C, transfer line at 280 C, flow of helium with constant flow 1.0mL/min. NIST02 Library

compounds

Figure 1. Assessment of recovery efficiency of the treatment compounds

CONCLUSIONS The resulting mixture contains alkaloids whose effect is not sufficient for analgesic use and whether sedation, respiratory depression and decrease the cough reflex, oral syrup recommended dose for this purpose is 10 4 4 mg/5mL . or 28.7-mg tablets have a ceiling effect of 400-450mg doses of codeine , the toxic effects of this extract should be considered, since without a purification of the mixture at a concentration (92 mg / mL) intravenously caused death in the study group. In many plants is the frequent presence of alkaloids because to serve as regulators of growth, nitrogen storage metabolized, bug deflector for its bitter taste, people who use these plants without reference to its toxicity for the compounds are part of a high statistical number of reported cases of poisoning or even deaths in which the diagnosis was mistaken, especially in rural area. these plants for their high-agonists could be considered a raw material for obtaining potential compounds against pain, modifying its structure in synthetic form in one or even two steps to obtain the most used drugs such as morphine (Figure 1), can even be modified to obtain compounds with lower dependency.
BIBLIOGRAFA 1. MW. Decker y col. (1992). Eur. J.Pharmacol. jun23;280(1)79-89 2. Schach Sy col.(1993). Biol Chem Hoppe Seyler. 1993 Mar;374(3):175-81. 3. Arango A. (2008).Facultad de qumica farmacutica. Medelln Colombia. 4. www. El ergonista.com/fitoterapa/opioextraccin.htm consultado 20 de enero de 2011 5. Frick S., Kramell R., Schmidt J., Fist J. and Kutchan T. 2005 Comparative Qualitative and Quantitative determination of alkaloids in Narcotic and Condiment Papaver somniferum cultivars . J. Nat. Prod. 2005, 68, 666-673 6.Gmez R. (1998). La toxicidad de las plantas ornamentales. CENEGARD. Espaa Lozoya X., Lozoya M., 1982 Flora Medicinal de Mxico. 1. Plantas Indgenas. Instituto Mexicano del Seguro Social, IMSS. D.F. Mxico. 309p 7.Snchez Herrera S., Soto Hernndez R., Kite G., Garca Mateos M.R. 2001. Identificacin de alcaloides en las inflorescencias de Erythrina americana Miller. Revista Chapingo Serie Horticultura 7(1): 37-4 8. Snchez Herrera S., Soto Hernndez R., Kite G., Garca Mateos M.R. 2001. Identificacin de alcaloides en las inflorescencias de Erythrina americana Miller. Revista Chapingo Serie Horticultura 7(1): 37-4

Biological assay: LD50 was determined under the criteria of the FEUM Systemic Injection MGA-DM 3083 with CD1 strains of mice weighing between 20-23 g from UPEAL-CINVESTAV. RESULTS AND DISCUSSION The evaluation of compounds of different extracts provides the extract temperature in acidic medium by autoclaving is more efficient in the recovery of organic compounds obtained as shown in Figure 1.

y y

codeine methylether

quinoline 3methyl

Oripavine

Erisodine

5-phenyl