1. Introduction
Complementary alternative medicine (CAM) has been widely used for a long time for the treatment of multiple diseases, despite the great advances in allopathic medicine. It is estimated that about 80% of the world population use some form of CAM.
CAM encompasses empirical knowledge and medical practice in which use is made of herbal medicinal plants, animals, minerals, manual therapy and exercise, alone or in conjunction for the treatment of diseases. In the early 1980's there emerged a strong interest in their study that has significantly influenced the pharmaceutical industry in developing technologies to identify new chemical entities and structures that are used for the synthesis of drugs. It has been shown that natural products play an important role in the discovery of compounds for drug development to treat multiple diseases.
Also, is important to recognize that use plants and their products have provided proven benefits to humanity, which falls into four areas: (i) food, (ii) essences and flavoring agents, (iii) perfumes and cosmetics, and (iv) biological and pharmaceutical agents [1]. Within the pharmaceutical area, the current outlook for natural products in drug discovery takes a central role, since at the beginning of this new millennium, only about 10% of 350,000 known species have been investigated from a phytochemical or pharmacology point of view [1].
A great examples of molecules that have hit the market as drugs by isolation from natural products metabolites are: taxol (1), an antitumor agent isolated from Taxus species [2] and camptothecin (2), isolated from the Chinese plant Camptotheca acuminate Decne (Nyssaceae), used to treat ovarian, breast and colorectal cancer, another example is ephedrine (3), which is isolated from the plant
Documentary research from 1981 to 2006 showed that natural products have been a source of 5.7% of drugs produced in those years. The derivatives of natural products are most of the times, chemical molecules synthetized from natural products and contributed to the 27.6% of the total of the new molecule.
2. Characterization of Geranium genus
The names come from Greek and refer to the form that its fruits acquire, likes beaks. Thus, the word "Geranium" comes from “geranos" meaning crane, and "Pelargonium" derived from "Pelargos" meaning stork [4].
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Within the classification of
Currently, in Hidalgo state, in Central Mexico, are classified 8 different species [5] and anyone has chemical or pharmacological studies.
2.1. Biological activities and compounds isolated from Geranium species
Some species of
One of the major components in
2.2. Different species of geraniums and its relevant compounds
The specie
Recently the extracts of
From
Constituents from the aerial parts of
3. Study of Geranium schiedeanum
3.1. Plant material
Specimen of
3.2. Extraction and purification
Air-dried aerial parts (1 kg) were extracted acetone-H2O 7:3 (20 L) by maceration for 7 days. Vacuum evaporation of dissolvent give a 5 L residue Filtration give a fatty solid residue (12g) and complete evaporation of water give the acetone-water extract (115 g).
Lots of 3 g of acetone-water extract were purified on a Sephadex LH-20 (25 g) column using H2O, H2O-MeOH (9:1, 4:1, 7:3, 3:2, 1:1, 2:3, 3:7, 1:4, 1:9 ) and MeOH, as eluents. Fractions of 300 mL of each polarity were collected and marked “A–K”. They were evaporated and analyzed by TLC and NMR. Fractions “B” gave 75 mg, and were purified over silica gel (10 g), using CHCl3, CHCl3-AcOEt(9:1, 4:1, 7:3, 3:2, 1:1, 2:3, 3:7, 1:4, 1:9 ) and AcOEt (10 mL of each polarity), as eluents and collecting fractions of 7 mL, fractions 13-16 give I 25 mg. Fractions “C” and “D” gave 56 mg, and were purified over silica gel (10 g), using CHCl3-MeOH (50:7.0, 48:7, 45:7, 40:7, 35:7 and 30:7, 40 mL of each), as eluents and collecting fractions of 7 mL, fractions 33-66 give II 2 mg, (these procedure was repeated ten times to obtain 18 mg of compound), Fractions “F-I” gave 1.8 g, a portion of 500 mg were purified over silica gel C-18 (5 g) using H2O, H2O-MeOH (9:1, 4:1, 7:3, 3:2, 1:1, 2:3, 3:7, 1:4, 1:9 ) and MeOH (20 mL of each polarity), fractions of 10 mL were collected fractions 2-4 gave 325 mg of III, fraction “K” gave 90 mg were added 5 mL of (40°C) pyridine and were placed a room temperature for 72 h, filtrated of mixture give a yellow needles 60 mg of IV (Figure 3).
3.3. Animals and treatment
Male adult Wistar rats 2 months old (200–220g) were obtained from UAEH Bioterio, and acclimated to our animal room for two weeks before use. Throughout these two weeks rats were supplied with food and water
3.4. Processing of samples
In order to clarify the sequential changes during the different stages of liver injury and the post-necrotic regenerative response, samples were obtained from control and at 24 and 48 h of TA intoxication in both Gs pre-treated or non pre-treated animals. Rats were sacrificed by cervical dislocation and samples of liver were obtained and processed as previously described. Blood was collected from hearts and kept at 4 °C for 24 h, centrifuged at 3000 rpm for 15 min, and serum was obtained as the supernatant.
3.5. Determination of AST
Enzymatic determination were carried out in serum in optimal conditions of temperature and substrate and cofactor concentrations. Aspartate aminotransferase (AST) activity were determined in serum. AST (EC 2.6.2.1) and was assayed following the method of Rej and Horder [38].
The activity of this enzyme was determined spectrophotometrically, by measuring the decrease in absorbance at 340 nm at 37 ° C, produced by the oxidation of NADH to NAD+ in the coupled reaction of reduction of oxaloacetate to malate, catalyzed by malate dehydrogenase, according to the following process:
3.6. General
IR spectra measured in MeOH on a Perkin Elmer 2000 FT-IR spectrophotometer. Optical rotations were determined in MeOH on a Perkin Elmer 341 polarimeter. NMR measurements performed at 400 MHz for 1H and 100 MHz for 13C on a VARIAN 400 spectrometer from CDCl3, CD3OH, DMSO-d6 solutions. Column chromatography (CC) was carried out on Merck silica gel 60 (Aldrich, 230-400 mesh ASTM) and sephadex LH-20 Sigma Aldrich.
3.7. Statistical analysis
The results were calculated as the means ± SD of four experimental observations in duplicate (four animals). Differences between groups were analyzed by an ANOVA following Snedecor F (α = 0.05). Students’ test was performed for statistical evaluation as follows: (a) all values against their control; b) differences between two groups Gs + TA versus TA.
3.8. Results
3.8.1. Active compounds of Geranium schiedeanum
One kg of the aerial part of
3.8.2. Aspartate aminotransferase
The acute liver injury induced by a necrogenic dose of thioacetamide (TA), a potent hepatotoxic agent, is characterized by a severe perivenous necrosis [39]. The necrosis develops as a consequence of the biotransformation of TA through the microsomal flavin-dependent monooxygenase [40]. The reactive metabolites responsible for TA hepatotoxicity are the radicals derived from thioacetamide-S-oxide and the reactive oxygen species derived as sub products in the process of microsomal TA oxidation, both of which can depleted reduced glutathione leading to oxidative stress [41, 42].
Liver damage induced by xenobiotics is characterized by the release in serum of hepatic enzymes due to necrosis of hepatocytes. AST is randomly distributed in the hepatic acinus, and is the enzyme activity used as marker of necrosis. Our results showed that
4. Conclusion
There is evidence that free radicals play a critical role in certain pathological conditions such as some cancers, multiple sclerosis, inflammation, arthritis and arterosclerosis [43]. For this reason, some research objectives directed toward the development or discovery of these compounds catchers of these radicals.
A large number of plant species, like
Also, in the present study TA-induced hepatotoxicity was used to investigate the effect of the pretreatment of
The pre-treatment with the crude extract in the model of thioacetamide-induced hepatotoxicity in rats, decreased and delayed liver injury by 66% at 24 h.
The data obtained indicate that the crude
Acknowledgments
The authors would like to thank Teresa Vargas for her valuable technical Assistance. Supported by Grant PROMEP-MEXICO UAEHGO-PTC-454.
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