M. Rajani

Evaluation of antioxidant properties of root bark of Hemidesmus indicus R. Br. (Anantmul).

Hemidesmus indicus R. Br. (Asclepiadaceae) is a well known drug in Ayurveda system of medicine. In the present study, antioxidant activity of methanolic extract of H. indicus root bark was evaluated in several in vitro and ex vivo models. Further, preliminary phytochemical analysis and TLC fingerprint profile of the extract was established to characterize the extract which showed antioxidant properties. The in vitro and ex vivo antioxidant potential of root bark of H. indicus was evaluated in different systems viz. radical scavenging activity by DPPH reduction, superoxide radical scavenging activity in riboflavin/light/NBT system, nitric oxide (NO) radical scavenging activity in sodium nitroprusside/Greiss reagent system and inhibition of lipid peroxidation induced by iron-ADP-ascorbate in liver homogenate and phenylhydrazine induced haemolysis in erythrocyte membrane stabilization study. The extract was found to have different levels of antioxidant properties in the models tested. In scavenging DPPH and superoxide radicals, its activity was intense (EC50 = 18.87 and 19.9 microg/ml respectively) while in scavenging NO radical, it was moderate. It also inhibited lipid peroxidation of liver homogenate (EC50 = 43.8 microg/ml) and the haemolysis induced by phenylhydrazine (EC50 = 9.74 microg/ml) confirming the membrane stabilization activity. The free radical scavenging property may be one of the mechanisms by which this drug is effective in several free radical mediated disease conditions.

A rapid method for isolation of andrographolide from andrographis paniculata nees (kalmegh).

A simple and rapid method for isolation of andrographolide from the leaves of Andrographis paniculata is reported. This involves extraction of the leaf powder by cold maceration in a 1:1 mixture of dichloromethane and methanol and isolation of andrographolide directly from the resulting extract by recrystallisation. The identity of the compound was confirmed through IR, UV, mass and melting point, and co-chromatography with a reference standard on TLC. The purity of the compound was confirmed by TLC, UV absorption spectrum, HPLC and differential scanning calorimetry, the latter of which gave the melting point of andrographolide as 235.3°C.

Swertiamarin: a lead from Enicostemma littorale Blume. for anti-hyperlipidaemic effect.

We have investigated the hypolipidemic effects of swertiamarin an active lead isolated from a perennial herb Enicostemma littorale Blume. in high cholesterol fed rats. Swertiamarin (50 and 75 mg/kg) and atorvastatin (50 mg/kg) was given orally daily for seven consecutive day to the high cholesterol feed rats. Serum total cholesterol, triglycerides, low density lipoprotein and very low density lipoprotein were found to be markedly elevated in the high cholesterol fed control rats and these changes were significantly prevented in swertiamarin treated animals. However, there was no significant effect on serum high density lipoprotein level. The 3-hydroxy 3-methyl glutaryl Co A (HMG-Co A) reductase activity was significantly inhibited in swertiamarin and atorvastatin treated groups compared to high cholesterol fed control group. Swertiamarin was also found to increased excretion of fecal bile acid and total sterols compared to control animals. In conclusion our data suggest that swertiamarin possess high antiatherogenic potential and an effective cholesterol lowering agent and inhibition of HMG-Co A reductase may be one of the main mechanisms of hypolipidemic effect of swertiamarin.

A Rapid Method for the Isolation of Swertiamarin from Enicostemma littorale

We report a simple method for the isolation of swertiamarin, a secoiridoid glycoside, from the whole plant of Enicostemma littorale Blume. Methanol extract of defatted plant material when treated with diethyl ether gave a precipitate containing swertiamarin as one of the major components. Swertiamarin was separated from this precipitate by column chromatography over silica gel. The identity of the compound isolated was confirmed through infrared (IR), ultraviolet (UV), nuclear magnetic resonance (NMR), and mass spectra and melting point and co-chromatography with a reference standard on thin-layer chromatography (TLC). The purity of the compound was confirmed from the UV absorption spectrum, NMR, mass, high performance thin layer chromatography (HPTLC), and differential scanning calorimetry.

Phytochemical Standardization of Herbal Drugs and Polyherbal Formulations

The recent global resurgence of interest in herbal medicines has led to an increase in the demand for them. Commercialization of the manufacture of these medicines to meet this increasing demand has resulted in a decline in their quality, primarily due to a lack of adequate regulations pertaining to this sector of medicine. The need of the hour is to evolve a systematic approach and to develop well-designed methodologies for the standardization of herbal raw materials and herbal formulations. In this chapter, various methods of phytochemical standardization, such as preliminary phytochemical screening, fingerprint profiling, and quantification of marker compound(s) with reference to herbal raw materials and polyherbal formulations, are discussed in detail and suitable examples are given.

A rapid method for isolation of piperine from the fruits of Piper nigrum Linn.

A simple, rapid and efficient method has been developed for the isolation of piperine from the fruits of Piper nigrum. The method involves extraction of the fruit powder with glacial acetic acid, from which piperine is partitioned into chloroform and subsequently crystallized. The identity of the compound was confirmed by its melting point, comparison of UV, IR, and mass spectral data with those from a reference standard, and co-chromatography with the reference standard using thin-layer chromatography (TLC). The purity of the compound was ascertained by TLC, by recording UV absorption spectra at the start, middle, and end positions of the spot on the plate, and by differential scanning calorimetry (DSC).

Sensitive high-performance thin-layer chromatographic method for the estimation of diospyrin, a tumour inhibitory agent from the stem bark of Diospyros montana Roxb.

Diospyrin, a tumour inhibitory agent from the stem bark of Diospyros montana was isolated and characterised. A sensitive high-performance thin-layer chromatographic (HPTLC) method was developed for the estimation of diospyrin. The method was validated for precision (intra- and inter-day), repeatability and accuracy. The method was found to be precise, with the RSDs for intra-day in the range of 0.72-1.85% and RSDs for inter-day in the range of 1.06-2.95%, for different concentrations. Instrumental precision and repeatability of the method were found to be 0.086 and 0.937 (% CV), respectively. Accuracy of the method was checked by performing the recovery study at two levels and average percentage recovery was found to be 97.87%. The developed HPTLC method was adopted for the estimation of diospyrin content of the stem bark of D. montana from different regions, which varied from 0.35 to 0.47% (w/w) in the samples.

Beneficial effects of swertiamarin on dyslipidaemia in streptozotocin-induced type 2 diabetic rats.

Dyslipidaemia is one of the major risk factors for cardiovascular disease in diabetes mellitus. Lipid changes associated with diabetes mellitus are attributed to increases in free fatty acid flux, secondary to insulin resistance. In the present study, we have investigated the beneficial effects of swertiamarin on dyslipidaemic conditions associated with type 2 diabetes in streptozotocin‐induced type 2 diabetic rats. Swertiamarin (50 mg/kg, i.p.) administered once a day for 6 weeks resulted in significant (p < 0.001) reductions in serum triglycerides, cholesterol and low‐density lipoprotein levels in diabetic animals as compared with diabetic control animals. Serum fasting glucose was significantly (p < 0.05) decreased, moreover, the insulin sensitivity index was significantly (p < 0.05) increased in swertiamarin treated animals. Overall the data suggest that swertiamarin has beneficial effects on diabetic associated complications such as dyslipidaemia. Copyright

A sensitive HPTLC method for estimation of swertiamarin in Enicostemma littorale Blume, Swertia chirata (Wall) Clarke, and in formulations containing E. littorale

Swertiamarin is a secoiridoid glucoside present in members of the Gentianaceae family, including Swertia chirata (Wall) Clarke, S. japonica Makino, S. angustifolia Buch.-Ham.ex D. Don, and Enicostemma littorale Blume. It has antidepressant and anticholinergic activity and can thus be used as a biomarker. We have developed a simple HPTLC method for quantification of swertiamarin which can be used for analysis of plant materials and formulations to determine swertiamarin content. The method was validated for precision, repeatability, and accuracy and found to be precise; intraand inter-day relative standard deviations (RSD) were in the ranges 0.68 to 0.85 and 0.71 to 1.03, respectively, for two different concentrations. Instrumental precision and repeatability of the method were 0.95 and 0.69 (%CV), respectively. The accuracy of the method was checked by determination of recovery at two different levels and the average recovery was found to be 100.13%. The method was used for estimation of the swertiamarin content of whole plants of E. littorale and S. chirata and of herbal formulations containing E. littorale as an ingredient. The method requires no clean-up of sample extracts before TLC and swertiamarin was well resolved from other components of the extracts. The method is simple, precise, specific, sensitive, and accurate and can be used for routine quality control of raw materials and herbal formulations. The method is suitable for quantification of swertiamarin in samples containing amounts ranging from 0.15 to 7.7% (w/w). E. littorale whole plant was found to be a rich source of swertiamarin (7.7% w/w).

Evaluation of the Antimicrobial Activity of Ammoniacum Gum from Dorema ammoniacum

Antimicrobial activity of the dichloromethane-methanol (1 : 1) extract of ammoniacum gum (from Dorema ammoniacum D. Don) was evaluated against 14 microorganisms which included seven Gram-positive bacteria (Bacillus cereus, Bacillus pumilus, Bacillus subtilis, Micrococcus luteus, Staphylococcus epidermidis, Staphylococcus aureus and Streptococcus faecalis), four Gram-negative bacteria (Escherichia coli, Pseudomonas aereuginosa, Klebsiella pneumoniae and Bordetella bronchiseptica), one yeast (Saccharomyces cereviseae) and two fungi (Aspergillus niger and Candida albicans). The extract of ammoniacum gum exhibited a of broad spectrum antimicrobial activity by inhibiting all the seven Gram-positive bacterium, one Gramnegative bacterium, one yeast and one fungus, with a minimum inhibitory concentration (MIC) of 40µg/ml. To overcome the solubility problem often faced when herbal extracts are added to aqueous medium, we employed a modified broth method where the broth cultures were agitated at 150 rpm in an orbital shaking incubator. This method reduced the MIC of the extract considerably, to 5-20µg/ml, against B. bronchiseptica, S. aureus and S. epidermidis.