Angiotensin-converting enzyme (ACE) inhibitory potential of standardized Mucuna pruriens seed extract.

CONTEXT: Mucuna pruriens Linn. (Fabaceae) is a tropical legume, traditionally used for controlling blood pressure. Inhibition of angiotensin-converting enzyme (ACE) is one of the successful strategies for controlling hypertension. OBJECTIVE: The present study evaluated the ACE inhibition potential of the standardized extract of M. pruriens seeds. MATERIALS AND METHODS: Standardization of the extract and its fractions were carried out by RP-HPLC method [methanol and 1% v/v acetic acid in water (5:95 v/v)] using levodopa as a marker. The ACE inhibition activity of the extract and fractions was evaluated at different concentrations (20, 40, 60, 80, and 100 µg/mL) using the HPLC-DAD and the UV spectrophotometric method. The liberation of hippuric acid (HA) from hippuryl-L-histidyl-L-leucine (HHL) was estimated in the spectrophotometric method and RP-HPLC assay at 228 nm. RESULTS: Methanol extract and aqueous fraction showed a maximum activity with IC50 values of 38.44 ± 0.90 and 57.07 ± 2.90 µg/mL (RP-HPLC), and 52.68 ± 2.02 and 67.65 ± 2.40 µg/mL (spectrophotometry), respectively. DISCUSSION: The study revealed that the aqueous extract contains the highest amount of levodopa. Eventually the methanol extract showed highest ACE inhibition activity except levodopa alone. It was further observed that the inhibition was altered with respect to the change in the content of levodopa in the extract. Thus, it can be assumed that levodopa may be responsible for the ACE inhibition activity of M. pruriens seeds. CONCLUSION: It can be concluded that M. pruriens seed is a potential ACE inhibitor can be explored further as an effective antihypertensive agent.

RP-HPLC simultaneous estimation of betulinic acid and ursolic acid in Carissa spinarum.

Carissa spinarum is a well-known medicinal plant which has been reported for its anthelmintic, antipyretic, antiviral, antimicrobial and antitumour activities. In this study, a reverse-phase high-performance liquid chromatographic method was developed for the simultaneous estimation of betulinic acid (BA) and ursolic acid (UA) in the methanol extract of C. spinarum root. The method was further validated for linearity, limit of detection (LOD = 3.3σ/S), limit of quantification (LOQ = 10σ/S), precision, accuracy and ruggedness. The linear response was obtained using the equation, y = 511.5x+17603 (r(2) = 0.9920) and y = 2886x+6821 (r(2) = 0.9935) for BA and UA, respectively. The LOD and LOQ were found to be 0.268 ± 0.520 µg mL(-1), 0.878 ± 0.183 µg mL(-1) for BA (0.58% w/w) and 3.140 ± 0.36 µg mL(-1), 8.820 ± 0.85 µg mL(-1) for UA (1.09% w/w), respectively. The %RSD of precision and recovery of BA and UA was < 2.0%. The proposed method was simple, accurate, specific, precise and reproducible.

Anti-cholinesterase activity of the standardized extract of Syzygium aromaticum L.

BACKGROUND: Clove (Syzygium aromaticum) is a well-known culinary spice with strong aroma; contains a high amount of oil known as clove oil. The major phyto-constituent of the clove oil is eugenol. Clove and its oil possess various medicinal uses in indigenous medicine as an antiseptic, anti-oxidant, analgesic and neuroprotective properties. Thus, it draws much attention among researchers from pharmaceutical, food and cosmetic industries. OBJECTIVE: The aim of the present study was to determine the anti-cholinesterase activity of the methanol extract of clove, its oil and eugenol. MATERIALS AND METHODS: In vitro anti-cholinesterase activity of S. aromaticum was performed by a thin layer chromatography bio autography, 96 well micro titer plate and kinetic methods. Reverse phase high performance liquid chromatography (RP-HPLC) analysis was carried out to identify the biomarker compound eugenol in clove oil. RESULTS: Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibition study revealed that eugenol possess better inhibition of the enzymes than extract and oil. Clove extract, its oil and eugenol showed better inhibition of AChE than BChE. Polyphenolic compound eugenol was detected through RP-HPLC analysis. The content of eugenol in essential oil was found to be 0.5 µg/ml. Kinetic analysis of the cholinesterase inhibition study of the extract; clove oil and eugenol have shown that they possess mixed type of inhibition for AChE and non-competitive type of inhibition for BChE. CONCLUSION: These results might be useful in explaining the effect of clove as anti-cholinesterase agent for the management of cognitive ailments like Alzheimer's disease.

Cytochrome P450 inhibitory potential and RP-HPLC standardization of trikatu--a Rasayana from Indian Ayurveda.

ETHNOPHARMACOLOGICAL RELEVANCE: Trikatu is a very well known 'Rasayana' in Ayurveda and widely used as a polyherbal ayurvedic formulation in India. It consists of three well known plants, viz., Piper longum (PL), Piper nigrum (PN) and Zingiber officinale (ZO) in equal ratio. Trikatu has been prescribed for cough, cold, fever, asthma, respiratory problems and improvement of digestive disorders. The aim of the present study was to investigate the effect of individual ingredients of trikatu namely PL, PN, and ZO and formulations [Marketed formulation (MF) and laboratory formulation (LF)] on drug metabolizing enzymes (CYP3A4 and CYP2D6), to assess its herb-drug interaction potential through cytochrome P450 inhibition assays. Further this work was aimed to develop an RP-HPLC method for the identification and quantification of piperine and 6-gingerol in the crude drug trikatu. MATERIALS AND METHODS: Enzyme inhibition effect of LF, MF, PL, PN and ZO was explored through CYP450-CO complex assay using rat liver microsomes (RLM) and a fluorescence screening method using individual isoenzymes (CYP3A4 and CYP2D6). The RP-HPLC method was developed for the identification and quantification of piperine and 6-gingerol in LF, MF and individual plant materials at the concentration of 1mg/mL. RESULTS: RP-HPLC analysis confirmed the presence of piperine and 6-gingerol in LF and MF [Piperine: 7.89±2.12% (w/w) (MF), 6.70±2.13% (w/w) (LF)]; [6-gingerol: 5.3±1.21% (w/w) (MF), 4.95±2.34% (w/w) (LF)]. Inhibitory potential of MF and LF in CYP450-CO complex assay was found to be 37.54±3.12% (MF) and 35.12±2.31% (LF) and against CYP2D6 and CYP3A4 was estimated to be IC50 251.30±3.98 and 245.23±1.92µg/mL and IC50 225.50±1.02 and 223.254±0.92µg/mL respectively. CONCLUSIONS: Different concentrations of the trikatu formulation and its individual components showed significantly (p<0.001) less inhibitory activity on individual isoenzymes as compared to the positive control. The crude drug exhibited inhibitory potential against the CYP450 enzymes in a concentration dependent manner. Outcome of the present study demonstrated that trikatu has less interaction potential with drug metabolizing enzymes.

Cholinesterase inhibition activity of Marsilea quadrifolia Linn. an edible leafy vegetable from West Bengal, India.

Maesilea quadrifolia Linn. (Marsileaceae) is a leafy vegetable well known in India. The current study aims to explore the phytochemical profile of M. quadrifolia and investigate its anti-cholinesterase potential. The methanol extract of the plant was subjected to qualitative and quantitative phytochemical screening (total alkaloidal content, saponin content and phenol content) and its anti-cholinesterase potential was tested by TLC bioautography and other screening methods using acytylcholinesterase (AChE) and butyrylcholinesterase (BChE). The study revealed that the extract contains various classes of phytoconstituents including steroids, saponins, alkaloids and other polyphenols. Total alkaloid, phenolic and saponin contents were found to be 19.3 mg g⁻¹ and 158.5 ± 1.02 mg g⁻¹ as gallic acid equivalents and 2.63 mg g⁻¹ of the extract, respectively. The TLC bioautography method exhibited the inhibition of both enzymes. In a microtiter plate assay, the IC50 values of the extract for AChE and BChE were found to be 51.89 ± 0.24 µg mL⁻¹ and 109.43 ± 2.82 µg mL⁻¹, respectively. These findings suggest that M. quadrifolia is a potential lead as an AChE and BChE inhibitor, which may be useful in the management of Alzheimer's disease.

Anticholinesterase activity of standardized extract of Illicium verum Hook. f. fruits.

Illicium verum is a well known spice in traditional Indian system for its therapeutic potential. The present study was aimed to evaluate the acetylcholinesterase (AChE) and butyrylcholinesterase inhibitory (BChE) activity of standardized extracts of I. verum and its oil. Present study confirmed that anethole contributed to the anticholinesterase activity of I. verum, with more specificity towards AChE. IC(50) for AChE and BChE inhibitory activity of anethole was 39.89±0.32 µg/mL and 75.35±1.47 µg/mL, whereas for the oil, 36.00±0.44 µg/mL and 70.65±0.96 µg/mL respectively. Therefore I. verum can be a good lead as anti-cholinesterase agent from natural resources.

Exploring the potential of Nelumbo nucifera rhizome on membrane stabilization, mast cell protection, nitric oxide synthesis, and expression of costimulatory molecules.

Immunomodulatory activity of Nelumbo nucifera rhizome was evaluated for its standardized extract (NNRE) with respect to betulinic acid. Various key parameters including erythrocyte membrane stabilization, inhibition of histamine release, reduction in nitric oxide production and depletion of expression of costimulatory molecules of macrophages were estimated. The result displayed that NNRE stabilized erythrocyte membrane significantly at 10 (42.05%) and 100 microg/mL (44.31%). Although considering the protection of mast cells from degranulation, NNRE showed 38.66% (100 microg/mL) and 69.66% (10 microg/mL) degranulation against compound 48/80 (C 48/80). NNRE at 1 and 5 microg/mL inhibited lipopolysaccharide (LPS)-induced activation of macrophages by decreasing the expression of costimulatory molecules. Expression of CD40, CD80, and CD86 by NNRE was seen significantly at 5 microg/mL compared to LPS-treated group. The extracts also inhibited the nitrite concentration at 1 and 5 microg/mL compared to LPS-treated group.