PHARMACOGNOSTICAL AND PHYSICOCHEMICAL EVALUATION OF THE PLANT LEUCAS ASPERA

ABSTRACTObjective: Leucas aspera (Willd.) Linn. is a herbaceous annual distributed throughout India from the Himalayas down to Ceylon. This study was aimedto investigate the pharmacognostical, phytochemical characteristics, and heavy metal content of the aerial parts of L. aspera.Methods: The aerial parts of the plant were subjected to macroscopical and microscopical characterization, followed by physicochemical evaluationsusing standardized procedures. Atomic absorption spectroscopy (AAS) of the dried plant powder was done to determine the metal and mineralcontent. Preliminary phytochemical screening and metabolite analysis of the hydroalcoholic extract of L. aspera (HAELA) were also performed toanalyze the various phytoconstituents.Results: In this study, the microscopic characteristics of L. aspera were found to be consistent with earlier reports. The total ash value was relativelyhigh (11.5% w/w). Alcohol soluble and water-soluble extractive values were found to be 6.5% and 9%, respectively. AAS indicated the plant tocontain insignificant quantities of heavy metals and considerable amounts of copper, sodium, potassium, and iron. Phytochemical analysis revealedthe presence of alkaloids, flavonoids, glycosides, lignins, phenols, saponins, sterols, and tannins in HAELA.Conclusion: The pharmacognosy - anatomical, physicochemical, heavy metal characteristics, and the preliminary phytochemical studies of the aerialparts of L. aspera have revealed the presence of phytoconstituents such as triterpenoids, phenolic compounds, flavonoids, and glycosides; insignificantquantities of heavy metals and significant content of phenolics and tannins in the plant.Keywords: Leucas aspera, Ash value, Heavy metal content, Atomic absorption spectroscopy

ANTIPROLIFERATIVE EFFECTS OF PSIDIUM GUAJAVA LEAF EXTRACT ON ORAL SQUAMOUS CARCINOMA CELLS

Objective: Cancer is the major cause of mortality affecting population irrespective of age. Oral cancer is one among the various cancers affecting major population in India. To overcome toxicity of chemotherapy and disfiguration by surgical procedures, researchers are targeting phytochemicals for their anticancer properties. This study evaluates the antiproliferative effects of Psidium guajava leaf extract against OSC cells. Methods: KB cells were purchased from NCCS, Pune. Extract from leaves of P. guajava was prepared with ethanol and evaluated with high-performance thin-layer chromatography (HPTLC). Antiproliferative effects of the extract were assessed with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl--tetrazolium bromide (MTT) assay and flow cytometry. Results: HPTLC revealed the presence of quercetin in the extract. MTT assay showed decreasing pattern in cell viability with the increasing dose of extract. Flow cytometry revealed the seizing of cycle by the extract. Conclusion: The study results conclude the presence of antiproliferative properties in the leaf extract of P. guajava

STABILITY INDICATING RP-HPLC METHOD FOR ESTIMATION OF REPAGLINIDE IN RABBIT PLASMA

Objective: A simple, selective and sensitive reverse-phase high-performance liquid chromatography (RP-HPLC) method to estimate repaglinide (REP) in rabbit plasma using rabeprazole (RAB) as an internal standard was developed and validated for various qualifications. Methods: The chromatographic separation was performed on C18 (2) analytical column (5 μ, 250×4.6 mm) using acetonitrile: 0.05% trifluoroacetic acid in water (55:45, v/v) as mobile phase at the flow rate of 1 ml/min. Validation of the analytical method was performed as per ICH guidelines. Results: The retention times of REP and RAB were found at ~4.3 and 5.1 min respectively, with adequate system suitability parameters (theoretical plates ≥3619, tailing factor ≤1.38, resolution factor 2.37). The method has linearity over a concentration range of 10 to 1000 ng/ml (r2=0.9987). The results of accuracy (≥98.17%), intra-, inter-day precision (≤2.9%), recovery (101.21±2.09%) and process efficiency (99.77±3.74%) found satisfactory with no matrix effect. The analyte in samples were found stable up to 6 h, 3 freeze-thaw cycles and not more than 2 mo corresponding to bench-top, short and long term stability studies respectively. Conclusion: The developed RP-HPLC method for estimation of REP in rabbit plasma was developed. The method was found to be rapid, cost-effective and accurate to estimate the REP from the sample matrix. The method can be a most useful tool for in vivo study of REP in the rabbit

Theoretical and experimental studies on 2-(2-methyl-5-nitro-1-imidazolyl)ethanol

In this work, we report a combined experimental and theoretical study on molecular structure, vibrational spectra, natural bond orbital (NBO) and UV spectral analysis of [2-(2-methyl-5-nitro-1-imidazolyl) ethanol] (Metronidazole-MTD). The FT-IR solid phase (4000-400 cm-1) liquid phase, and FT-Raman spectra (3500-50 cm-1) of MTD were recorded. The molecular geometry, harmonic vibrational frequencies and bonding features of MTD in the ground state have been calculated using the density functional method B3LYP with 6-311G(d,p) as basis set. The assignments of the vibrational spectra have been carried out with the help of the Gauss view program package. Stability of the molecule arising from hyperconjugative interactions and charge delocalization have been analyzed using natural bond orbital analysis. The results show that charge in electron density (ED) in the σ* and π* antibonding orbitals and second order stabilization energies E2 confirms the occurrence of Intramolecular Charge Transfer (ICT) within the molecule. The UV spectrum was measured in ethanol solution. The energy and oscillator strength calculated by Time-Dependent Density Functional Theory (TD-DFT) results complement the experimental findings. The calculated HOMO and LUMO energies show that charge transfer occurs within the molecule. The 1H and 13C nuclear magnetic resonance (NMR) chemical shifts of the molecule were calculated by the Gauge Independent Atomic Orbital (GIAO) method and compared with experimental results.  Finally the results of calculations were applied to simulate Infrared and Raman spectra of the title compound which show good agreement with observed spectra

Production and partial purification of β-galactosidase enzyme from probiotic Bacillus subtilis SK09

139-144In the present study, lactose hydrolyzing enzyme β-galactosidase was produced from a novel probiotic Bacillus subtilis SK09 and it was compared with commercially obtained, B. subtilis MTCC 2413. The probiotic Bacilli was initially screened for its ability to hydrolyze the X-gal followed by production of blue colonies. The production medium was formulated using varied nutrients including carbon, nitrogen, mineral salts and amino acid sources. The crude enzyme with an activity of 76.2 U/ml was achieved from our novel strain B. subtilis SK09, and subjected for partial purification by solvent precipitation, dialysis and ion exchange chromatography using DEAE cellulose (DEAE-C) column. Partially purified β-galactosidase enzyme showed 11.6 folds increase in specific activity of 137.02 U/ml. This partially purified enzyme was further characterized and it was observed that it is homogeneous in nature, with molecular weight of 43 kDa. When compared, the β-galactosidase activity of our novel strain was found five times higher than that of commercially obtained strain (27.6 U/ml). Hence, this characteristic of B. subtilis SK09 of increase enzymatic activity can make it a promising candidate for various industrial as well as biotechnological applications

Ghrelin and its Association with Nutritional and Inflammatory Status of Patients on Maintenance Hemodialysis in a South Indian Tertiary Care Hospital

Background: Malnutrition and inflammation are associated with morbidity and mortality in patients on maintenance hemodialysis (MHD). Ghrelin, an orexigenic peptide hormone, is speculated to be associated with nutritional and inflammatory status in MHD. Aim: To assess the serum total ghrelin levels and its possible relationship with inflammation and nutritional status in patients on MHD. Subjects and Methods: The study was conducted on 90 patients on MHD for 6 months and above (56 males, 34 females, mean age 52.6 [11.7] years; mean dialysis vintage 20.9 [12.1] months) and 70 healthy volunteers as control (5 males, 25 females, mean age 50.6 [9.7] years). Demographics were obtained for the study population, and dialysis‑related data were collected for cases. Anthropometry, biochemical parameters, serum total ghrelin and inflammatory markers tumor necrosis factor‑alpha (TNF‑α), and high‑sensitivity C‑reactive protein (hsCRP) were assessed for cases and control. Self‑reported appetite (five questions of appetite and diet assessment tool) and nutritional status (subjective global assessment‑dialysis malnutrition score) were assessed for cases. Results: Ghrelin (242.5 [62.3] pg/mL vs. 80.2 [19.6] pg/mL; P < 0.001), TNF‑α (39.8 [15.2] pg/mL vs. 6.5 [1.2] pg/mL; P < 0.001), hsCRP (10.2 [2.8] mg/L vs. 2.7 [0.54] mg/L; P < 0.001) were significantly elevated in cases versus control, anthropometry, and biochemical parameters were significantly decreased in hemodialysis patient. Of 90 cases, (13/90 [14.4%]) were well‑nourished, (28/90 [31%]) mild to moderately malnourished, and (49/90 [54.4%]) were moderate to severely malnourished. Appetite was very good for 14.4%, good and fair for 47.8%, poor and very poor for 37.8% patients. There was a significant difference in appetite with respect to nutritional status (P < 0.001). Ghrelin had positive correlation with inflammatory markers and negative correlation with nutritional status (P < 0.001). Conclusion: The study identified the association of ghrelin with appetite, nutritional, and inflammatory status of the patients on MHD.Keywords: Appetite, Ghrelin, Hemodialysis, Inflammation, Nutritional statu

Investigation of phytoconstituents of Cardiospermum halicacabum and its efficacy as a potential anti-cancer drug candidate

On the basis of the traditional knowledge of treating solid tumor using Cardiospermum halicacabum gained attention to carry out the present work. The preliminary screening for determination of cytotoxicity of successive extract of C halicacabum (hexane, chloroform, ethyl acetate and ethanol) was performed through brine shrimp lethality assay and yeast growth rate cytotoxicity assay. Among the extracts, chloroform extract of C halicacabum exhibited highest cytotoxicity which was further investigated for anti-proliferative and cytotoxicity property in A-549 lung cancer cell line. Correspondingly, dose dependent effects on reduction of cell proliferation with prominent morphological abnormalities of A-549 cells were observed under treatment with chloroform extract. Hemocompatibility assessment of chloroform extract, by hemolysis assay, revealed its compatibility towards RBC which in turn may prevent heamolytic anemia (myelosuppression) the most adverse effects of cancer chemotherapy.  Phytoconstituents of chloroform extract responsible for the cytotoxicity, anti-proliferative and hemocompatibiltiy was assessed by several chromatographic methods such as TLC, Column chromatography and HPLC which revealed the presence of flavonoids in chloroform extract of C halicacabum. Apparently the isolated flavonoids, in consistence with chloroform extract, exhibited similar effect on inhibition of proliferation of tumor cell line A-549. Therefore it could be evidenced from the current study that the C halicacabum, with its predisposed flavonoids, possesses anti-cancer property with least adverse effect on heamolysis. However detailed investigation on regulation of tumor cell proliferation and hemocompatability is required to bring C halicacabum as a potential candidate for cancer therapy.    Keywords: C halicacabum, Anti-cancer, Toxicity, Brine Shrimp Lethality Assay, Hemolysis

Identification of Patala (Stereospermum colais and Stereospermum suaveolens roots) by pharmacognostic parameters - A plant drug in Dasamula

Stereospermum colais and Stereospermum suaveolens are known as "Patala" in Ayurveda and also a constituent of Dasamula. It is difficult to distinguish between the two species and hence it is difficult to identify genuine Patala. Both Stereospermum species looks alike in morphology with the exception of flower-color. So collection of the plant material from wild source as well as differentiating from the marketed sample is a challenge for the herbal industries to identify the genuine Patala. Hence, an attempt was made to compare the pharmacognostical and phytochemical parameters of the roots of S. colais and S. suaveolens. Macroscopy, microscopy, physico-chemical analysis and elemental analysis were carried out to standardize the roots. The salient diagnostic features identified to distinguish the plant species are heterocellular periderm and calcium oxalate druses in S. colais and multitype (Rhytidome) periderm and calcium oxalate raphides in S. suaveolens. Qualitative and quantitative phytochemical analysis and comparative HPTLC fingerprint analysis of various extracts of roots revealed their phytochemical composition. The standardization parameters developed here can be used as reference standard for correct identification of the plant. Further, it will act as a tool to detect adulterants and substituents consequently maintains the quality, reproducibility and efficacy of the plant material.

Free radical scavenging activity of Lantana aculeata root extract in hyperlipidemic rats.

Abstract: Lantana aculeata is a common weed that grows abundantly in many parts of India. The aerial part of the plant is reported to be toxic while the roots were found to be non-toxic when tested in albino rats. The alcoholic extract of the roots showed a significant hypolipidemic activity in normal rats. Hence the roots were studied for their free radical scavenging potential in hyperlipidemic animals by administering the alcoholic extract (LAR) in doses of 25, 50 and 100 mg/kg for 30 days. The levels of LPO, non-enzymatic antioxidant (TRG) and enzymatic antioxidants viz. SOD, CAT and GPx that showed changes in diseased condition were reverted back to near normal values by LAR extract treatment of plasma, liver and heart tissues. The presence of flavonoids besides oleanolic acid in large amounts might have caused the observed effect

The human genome as the 'common heritage of humanity' : a human rights approach to international legal governance of the human genome

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