Hexane-ethyl Acetate Research Articles

Protective effect of Croton caudatus Geisel leaf extract against experimental visceral leishmaniasis induces proinflammatory cytokines in vitro and in vivo

In the present state of overwhelming emergence of drug-unresponsive phenotypes of Leishmania donovani and persistent severe toxicity in conventional anti-leishmanial therapy, in search for novel leads, the aim of this study has been fixed to identify the active extract(s) of Croton caudatus Geisel. var. tomentosus Hook effective against the parasitic protozoans in vitro and in vivo. C. caudatus Geisel. is often used by Chakma and Hmar community, the local tribes of north-east India for medicinal and veterinary purposes. Among the five semi-purified extracts tested, C. caudatus leaves, extracted in hexane and subsequently semi-purified in a column packed with silica gel (70–130 µM; mesh size 60 A°) using ethyl acetate-hexane solvent (9:1), was found to be the most effective growth inhibitor (JDHex) against the Leishmania promastigotes and amastigotes. JDHex significantly altered the biochemical parameters (protein, lipid and carbohydrates) in promastigotes followed by the morphological changes, DNA condensation and subsequent apoptosis in L. donovani. In consequent steps, it has been also proved that JDHex reduced the replication of intracellular amastigotes with concomitant release of nitric oxide and pro-inflammatory cytokines, IL-12 and TNF-α in vitro. Significantly, the 50% inhibitory concentration of JDHex was estimated much lower against the intracellular amastigotes (2.5 µg/mL) in comparison to promastigotes (10 µg/mL). JDHex was also found efficient in reducing parasite burden in spleen and liver when treated in vivo and increased the intracellular IFN-γ and decreased the IL-10 in CD4+ T cells in splenocytes of orally treated animals. The results of this study support the importance in exploration of novel anti-leishmanial leads from C. caudatus Geisel. var. tomentosus Hook. against the L. donovani (MHOM/IN/83/AG83) infection. Partial chemical characterization of JDHex revealed the presence of terpenoids. However, the further chemical investigation of JDHex is warranted.

Rethinking conventional approaches to the detection, management and amelioration of disease

The linear-leafed water primrose ( Ludwigia hyssopifolia ), with its distinctive yellow flowers and matted undergrowth, is easily recognized along paddy fields and tropical wetlands especially in South and South-East Asia. Also known as the seedbox because of its distinctive dimorphic seeds, it has the potential of being an invasive pest and is classed a ‘serious’ or ‘principal’ weed in many countries. It is used locally for composting and in the concoction of traditional remedies in an unscientific and informal manner. In common with other plants of the willow herb or evening primrose family, this weed could turn out to have immense and economically viable pharmaceutical potential. Das et al have reported moderate anti-tumor and antibacterial activities in extracts and in an alkaloid piperine from L. hyssopifolia and Mohammad et al demonstrated anti-­‐diarrheal properties in a methanol extract. It is noteworthy along the same line that the work by Luximon‐Ramma, et al on Mauritian Exotic fruits strongly advocated the use of the red and yellow Psidium cattleianum Sabine ‘Chinese guava’, in nutritional programs due to their exceptionally high antioxidant potencies attributed to their rich vitamin C and phytophenolic profiles. These plants are also generally considered as invasive causing havoc amidst the protected endemics of the island and sing an ecological conservation approach, indeed legitimate, to safeguard threatened plants from extinction, the systematic removal of Chinese Guava trees may deprive us of important sources of active nutritional biofactors. In a study that features in this issue of the Archives Of Medical And Biomedical Research, Scientists at the University Of Dhaka have further formalized the pharmacotherapeutic potential that this much-­‐derided weed may hold. They studied the effects of hexane, ethyl acetate and methanol extracts of whole plant parts of Ludwigia hyssopifolia on carrageenan-induced paw edema, acetic acid‐induced writhing, and diuresis in mice. The Hexane extract and ethyl acetate extract showed significant inhibition of experimental paw edema. All three fractions significantly inhibited writhing and, in comparison to furosemide, exhibited good diuretic activity.

Development and validation of one-step ultrasound-assisted extraction for simultaneous determination of multiclass fungicides in soils.

A rapid, efficient, and simple one-step ultrasound-assisted extraction (UAE) method was developed for the analysis of seven fungicides (cymoxanil, metalaxyl, mandipropamid, folpet, chlorothalonil, kresoxim-methyl, and famoxadone) in horticultural soils. Analytes in the samples were determined by HPLC with variable wavelength detection. Key parameters that influence the UAE procedure were optimized, such as the nature and volume of extraction solvent, number of sonication steps, and sonication time. The highest extraction efficiencies in the range of 61.1-87.8% were obtained by using only 7.5 mL of ethyl acetate-hexane (1+1, v/v) and sonicating for 10 min. At 0.5 and 2.0 μg/g fortification levels, satisfactory recoveries (>60%) with RSD<13% were obtained for each analyte, except for folpet (>52%). The method was linear over the range of 0.005 to 10 μg/g and the correlation coefficients (r2) obtained ranged from 0.9955 to 0.9992. The LODs (S/N=3) varied from 0.0015 to 0.006 μg/g. The proposed UAE procedure was compared to classical extractions (shake-flask and Soxhlet extraction) and showed satisfactory extraction efficiencies using shorter time and smaller amounts of organic solvents, thereby minimizing the costs of the analysis and the disposal of waste solvent.

Novel Design for Centrifugal Counter-Current Chromatography: VI. Ellipsoid Column

A novel ellipsoid column was designed for centrifugal counter-current chromatography. Performance of the ellipsoid column with a capacity of 3.4 mL was examined with three different solvent systems composed of 1-butanol–acetic acid–water (4:1:5, v/v/v) (BAW), hexane–ethyl acetate–methanol-0.1 M HCl (1:1:1:1, v/v/v/v) (HEMH), and 12.5% (w/w) PEG1000 and 12.5% (w/w) dibasic potassium phosphate in water (PEG-DPP) each with suitable test samples. In dipeptide separation with BAW system, both stationary phase retention (Sf) and peak resolution (Rs) of the ellipsoid column were much higher at 0° column angle (column axis parallel to the centrifugal force) than at 90° column angle (column axis perpendicular to the centrifugal force), where elution with the lower phase at a low flow rate produced the best separation yielding Rs at 2.02 with 27.8% Sf at a flow rate of 0.07 mL/min. In the DNP-amino acid separation with HEMW system, the best results were obtained at a flow rate of 0.05 ml/min with 31.6% Sf yielding high Rs values at 2.16 between DNP-DL-glu and DNP-β-ala peaks and 1.81 between DNP-β-ala and DNP-L-ala peaks. In protein separation with PEG-DPP system, lysozyme and myolobin were resolved at Rs of 1.08 at a flow rate of 0.03 mL/min with 38.9% Sf. Most of those Rs values exceed those obtained from the figure-8 column under similar experimental conditions previously reported.

Pharmacological effect of Ageratina pichinchensis on wound healing in diabetic rats and genotoxicity evaluation

Ethnopharmacological relevanceAmong the main causes affecting the wound healing process, we find diabetes mellitus, which is due to the occurrence of a prolonged inflammation phase, defects in angiogenesis, and a diminution in fibroblast proliferation. The species Ageratina pichinchensis has been utilized in Mexican traditional medicine for the treatment of skin wounds. Pharmacological models have demonstrated that an extract obtained from this species improves wound healing and, through a clinical study, it was evidenced that the extract (in a pharmaceutical form) is effective in the treatment of patients with chronic venous ulcers. The 7-O-(β-d-glucopyranosyl)-galactin compound was recently identified as responsible for the pharmacological activity. The objective of the present study was to evaluate the wound healing activity of an aqueous extract and another hexane-ethyl acetate extract from Ageratina pichinchensis (both standardized in the active compound) in a diabetic foot ulcer rat model, as well as evaluating the possible genotoxic effects produced by the same species. Materials and MethodsRats with streptozotocin-induced diabetes were submitted (under anesthesia with pentobarbital) to a circular lesion on the skin (excisional) on the rear of the paw. All animals were topically treated daily until healing. 5-methyl-1 phenyl-2-(1H) Pyridone was used as a positive control treatment. Once the wound was healed, a skin sample was obtained and utilized for histopathological analysis. The possible genotoxic effects produced by the extract, in a model of spermatozoid viability and morphology, were evaluated. ResultsThe results showed that 100% of animals treated with Ageratina pichinchensis extracts presented wound healing between days 4 and 11 of treatment, while in the positive control group (treated with 5-methyl-1 phenyl-2-(1H) pyridone) and in the negative control group (vehicle), only 70% and 40%, respectively, exhibited wound healing at day 11. Histological analysis demonstrated evidences of an active regenerative process in animals that received the extracts, in addition to that in the study, the effects of the plant extracts that could be compatible with genotoxicity were not observed. ConclusionsAqueous and hexane-ethyl acetate extracts of the aerial parts of Ageratina pichinchensis (standardized in its content of 7-O-(β-d-glucopyranosyl)-galactin), consistently improve wound healing induced on the skin of rats with streptozotocin-induced diabetes. The capacity was evidenced of the extracts to promote histological tissue regeneration, without exhibiting genotoxicity.

EFFECTIVE ON-LINE COMBINATION OF HIGH SHEAR TECHNIQUE AND CONTINUOUS SAMPLE INJECTION IN HIGH-PERFORMANCE COUNTER-CURRENT CHROMATOGRAPHY FOR ISOLATION AND PURIFICATION OF POLYPHENOLS IN GREEN TEA

A new sample preparation method of high shear dispersing emulsifier (HSDE) coupled with continuous sample injection in high-performance counter-current chromatography (HPCCC) was successfully used for the extraction and online isolation of caffeine and two polyphenolic compounds from green tea. The lower phase of hexane-ethyl acetate–methanol–water (1:8:1:8, v/v/v/v) was used as both the HSDE solvent and HPCCC mobile phase. In one time sample loading, 1.5 mg of (-)-epicatechin-3-gallate, 5.0 mg of (-)-epigallocatechin-3-gallate, and 3.1 mg of caffeine with the purity over 95.0% determined by high performance liquid chromatography (HPLC), could be obtained from about 0.1 g of raw material of green tea. Ten time sample injections were realized in HSDE extraction−HPCCC separation process, and the quantity could be magnified about decuple, 13.8 mg of (-)-epicatechin-3-gallate, 46.3 mg of (-)-epigallocatechin-3-gallate, and 28.9 mg of caffeine could obtain from 1.0 g of raw material of green tea. The compounds were analyzed by HPLC and identified by their chromatograms and their retention time compared with that of authentic reference compounds. It was the first time combining HSDE with continuous sample injection in HPCCC for effective isolation and purification of active compounds from natural products. The mechanism of HSDE extraction was illustrated.

Apoptosis Induction in Colon Cancer Cell Lines and Alteration of Aberrant Crypt Foci in Rat Colon by Purple Rice (Oryza sativa L. var. glutinosa) Extracts

Crude ethanol extracts (CEE) of purple rice was fractionated to obtain hexane soluble (HSF) and ethyl acetate soluble fractions (EASF). Total antioxidant capacity was higher in CEE than the HSF and EASF. However, HSF exhibited strong antiproliferation and apoptosis induction against colon cancer cell lines, both p53 wild-type (RKO) and mutant (SW620) strains. Then, the CEE was used to determine the effects on the progression of aberrant crypt foci (ACF), a preneoplastic lesion seen in colon carcinogenesis in rats. Male Wistar rats were subcutaneously injected of 40 mg/kg body weight dimethylhydrazin (DMH) once weekly for 2 wk. After 2 wk, rats were orally administered ethanol extract at 100 and 1000 mg/kg body weight, for 4 wk. Rats fed with only the high dose of CEE had significantly decreased numbers of ACF per rat (45.56%) and crypt multiplicity (AC/focus) (16.67%) compared to rats that received DMH alone. The result also demonstrated that CEE induced apoptosis in colonic epithelium cells of rat received colon carcinogen as detected the increasing of caspase-3 activity. This finding could be concluded that purple rice extracts inhibited aberrant colonic epithelial cell progression via apoptosis induction.

Cycloartane Triterpenoids from Euphorbia Macrostegia with their Cytotoxicity against MDA-MB48 and MCF-7 Cancer Cell Lines.

The dried plant was extracted with dichloromethane and after defatting with hexane, transferred repeatedly on silica columns using dichloromethane-hexane and ethyl acetate-hexane as mobile phases. Finally the fractions were purified by high performance liquid chromatography using a Pack-Sil column and hexane: Ethyl acetate as mobile phase. The structures of the isolated compounds included: cycloart-25-ene-3β, 24-diol (1), cycloart-23(Z)-ene-3β, 25-diol (2), cycloart-23(E)-ene-3β, 25-diol (3), and 24-methylene-cycloart-3β-ol (4) were elucidated by (13)C- and (1)H-NMR as well as IR and by the aid of mass fragmentation pattern and comparing with the literature. The biological effects of the compounds were done by the MTT assay on two different cancer cell lines including MDA-MB48 and MCF-7. Among these compounds, cycloart-23(E)-ene-3β,25-diol (3) was the most active compound on MDA-MB468 cell line (LD50 = 2.05 μgmL(- 1) ) and cycloart-23(Z)-ene-3β, 25-diol (2) was the most active compound on MCF-7 cell line (LD50 = 5.4 μgmL(- 1)).

Quantification of bioactive marker β-amyrin by validated high-performance thin-layer chromatographic- densitometric method in different species ofMaytenusgrown in Saudi Arabia

A simple and sensitive high-performance thin-layer chromatographic (HPTLC)-densitometric method was developed and validated for quantification of β-amyrin in the crude extracts of two species of Maytenus (Maytenus obscura and Maytenus parviflora) grown in Saudi Arabia. HPTLC-densitometry was performed on glass-backed silica gel 60 F254 TLC plates with the binary mobile phase hexane-ethyl acetate (3:1, v/v). The developed plate was derivatized with p-anisaldehyde, then scanned and quantified densitometrically at 550 nm. The system was found to give a compact spot for β-amyrin at RF value of 0.38 ± 0.01. The method was found to be satisfactory in terms of sensitivity, accuracy, precision, and recovery. The content of β-amyrin was estimated as 0.42% ± 0.01% and 0.88% ± 0.01% w/w in M. obscura and M. parviflora, respectively. The developed HPTLC technique can be very useful for the quantification of β-amyrin present in various medicinal plants.

ISOLATION AND PURIFICATION OF ALOIN, ISOALOERESIN D AND ALOENIN-2′-p-COUMAROYL ESTER FROM Aloe barbadensis MILL BY HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY

Aloin, isoaloeresin D, and aloenin-2′-p-coumaroyl ester were separated and purified from Aloe barbadensis Mill by one step of high-speed counter-current chromatography for the first time. The separation was performed with an optimized two-phase solvent system composed of hexane-ethyl acetate- acetone-water (0.2:5:1.5:5, v/v/v/v), using the lower phase as the mobile phase in the head-to-tail elution mode. The revolution speed of the separation column, flow rate of the mobile phase, and UV detection wavelength were 860 rpm, 1.5 mL · min−1, and 300 nm, respectively. In a one-step operation, 51.9 mg of aloin, 23.1 mg of isoaloeresin D, and 6.5 mg of aloenin-2′-p-coumaroyl ester could be obtained from 161.7 mg of dried acetone extract at purities of 99.2%, 98.5%, and 95.6%, respectively, as determined by HPLC using area normalization method. The partial separation of aloin A and aloin B was also obtained. The structures of these compounds were identified by high resolution mass spectroscopy (HRMS), UV, IR, and NMR.

Large-scale separation of alkaloids from Gelsemium elegans by pH-zone-refining counter-current chromatography with a new solvent system screening method

A rapid and simple solvent system screening method was developed for pH-zone-refining counter-current chromatography (CCC) separation, which was much easier to find a suitable solvent system than the traditional method. Using this method, an optimal solvent system composed of hexane–ethyl acetate–methanol–water (3:7:1:9, v/v) was selected for pH-zone-refining CCC separation of alkaloids from the stems of Gelsemium elegans, where 10mM triethylamine (TEA) was added to the upper organic stationary phase as a retainer and 10mM hydrochloric acid (HCl) to the aqueous mobile phase as an eluter. As a result, six oxindole alkaloids, 420mg 19-xo-gelsenicine, 456mg gelsemine, 723mg koumine, 379mg 11-methoxygelsemamide, 342mg gelsenicine and 318mg humantenine were successfully purified in one step from 4.5g crude extract with the purity of over 95%, respectively. The structures of the oxindole alkaloids were identified by ESI-MS, 1H NMR and 13C NMR.

Dynamic pH junction high-speed counter-current chromatography coupled with microwave-assisted extraction for online separation and purification of alkaloids from Stephania cepharantha

A simple and efficient dynamic pH junction high-speed counter-current chromatography method was developed and further applied to the online extraction, separation and purification of alkaloids from Stephania cepharantha by coupling with microwave-assisted extraction. Mineral acid and organic base were added into the mobile phase and the sample solution, respectively, leading to the formation of a dynamic pH junction in the column and causing focus of alkaloids. Selective focus of analytes can be achieved on the basis of velocity changes of the pH junction through appropriate selection of solvent systems and optimization of additive concentrations. The extract can be directly introduced into the HSCCC for the online extraction, separation and purification of alkaloids from S. cepharantha. Continuous separation can be easily achieved with the same solvent system. Under the optimum conditions, 6.0g original sample was extracted with 60mL of the upper phase of hexane–ethyl acetate–methanol–water (1:1:1:1, v/v/v/v) containing 10% triethylamine under 50°C and 400W irradiation power for 10min, the extracts were directly separated and purified by high-speed counter-current chromatography. A total of 5.7mg sinomenine, 8.3mg 6,7-di-O-acetylsinococuline, 17.9mg berbamine, 12.7mg isotetrandrine and 14.6mg cepharanthine were obtained with purities of 96.7%, 93.7%, 98.7%, 97.3% and 99.3%, respectively. The online method provides good selectivity to ionizable compounds and improves the separation and purification efficiency of the high-speed counter-current chromatography technique. It has good potential for separation and purification of effective compounds from natural products.

English

Seven isolates of&nbsp;Fusarium graminearum&nbsp;obtained from Seed and Plant Improvement Institute were grown on autoclaved barley grains and toxicity of their cultures to wheat germinating seeds was tested using their semi-purified phytotoxins. The extracts of isolates 167, 169, 173 and 179 did not have considerable inhibitory effects on wheat seed germination, whereas that of 161 (a = 0.05) and two other isolates 131 and 164 displayed a significant inhibitory effect on germination. However, no correlation was found between virulence and toxicity of the isolates examined. Also, the effect of light was studied in relation to wheat germling level of tolerance. Six isolates were investigated for&nbsp;their ability of deoxynivalenol synthesis. Thus, their crude extracts were prepared and partially purified, and subjected to thin layer chromatography with chloroform (acetone and hexane) ethyl acetate solvent systems on fluorescent silica gel plates (60 meshes) using a commercial standard deoxynivalenol (1 mg/ml, Sigma Co., USA) along with the samples. The spots of interest were detected and analyzed under ultraviolet (UV) lights (254 and 365 nm). The isolate 161 produced the most amount of deoxynivalenol compared to others. Also, no correlation was found between virulence and toxigenicity of the isolates. &nbsp; Key words:&nbsp;Phytotoxicity,usarium head blight,head scab, virulence, deoxynivalenol,&nbsp;mycotoxin, phytotoxin,Fusarium graminearum.

Method Development andValidation for the Simultaneous Determination of Organochlorine and Organophosphorus Pesticides in a Complex Sediment Matrix

The Xochimilco area in the southeastern part of Mexico City has a variety of socioeconomic activities, such as periurban agriculture, which is of great importance in the Mexico City metropolitan area. Pesticides are used extensively, some being legal, mostly chlorpyrifos and malathion, and some illegal, mostly DDT. Sediments are a common sink for pesticides in aquatic systems near agricultural areas, and Xochimilco sediments have a complex composition with high contents of organic matter and clay that are ideal adsorption sites for organochlorine (OC) and organophosphorus (OP) pesticides. Therefore, it is important to have a quick, affordable, and reliable method to determine these pesticides. Conventional methods for the determination of OC and OP pesticides are long, laborious, and costly owing to the high volume of solvents and adsorbents. The present study developed and validated a method for determining 18 OC and five OP pesticides in sediments with high organic and clay contents. In contrast with other methods described in the literature, this method allows isolation of the 23 pesticides with a 12 min microwave-assisted extraction (MAE) and one-step cleanup of pesticides. The method developed is a simpler, time-saving procedure that uses only 3.5 g of dry sediment. The use of MAE eliminates excessive handling and the possible loss of analytes. It was shown that the use of LC-Si cartridges with hexane-ethyl acetate (75+25, v/v) in the cleanup procedure recovered all pesticides with rates between 70 and 120%. The validation parameters demonstrated good performance of the method, with intermediate precision ranging from 7.3 to 17.0%, HorRat indexes all below 0.5, and tests of accuracy with the 23 pesticides at three concentration levels demonstrating recoveries ranging from 74 to 114% and RSDs from 3.3 to 12.7%.

Validation of one-step cleanup and separation method of polychlorinated biphenyls, organochlorine pesticides, and polycyclic aromatic hydrocarbons from atmospheric gas- and particle-phase samples

A one-step cleanup method is described for the determination of PAHs, PCBs and OCPs in air (gas and particulate phase) samples. Analytes were extracted from ambient air samples using soxhlet extraction with a solvent mixture of dichloromethane and petroleum ether (1:4) for 24h. They were concentrated, separated and fractionated on a florisil and alumina column. The amounts of florisil (1g or 2g) with/without alumina were tested in the cleanup column. The study systematically investigated the effects of solvent types, and the amounts of florisil and alumina, on the performance of the cleanup process. The first fraction was eluted with 25mL hexane, and analyzed for PCBs. The second fraction was collected via 40mL hexane–ethyl acetate (1:1) solvent mixture, and analyzed for OCPs and PAHs. The optimized method yielded average recoveries between 88% and 99% for PCBs; 56% and 118% for PAHs; and 51% and 128% for OCPs. Other validation parameters were also investigated, such as MDL, LOQ, linear range, sensitivity (r2). An oven-program optimization and adjustment of GC–MS were performed. For internal quality control, surrogate recoveries and field blanks values were calculated. External calibration curves were prepared for PAHs, and internal calibration curves were preferred for OCP and PCBs.

Conical coils counter-current chromatography for preparative isolation and purification of tanshinones from Salvia miltiorrhiza Bunge

Modern counter-current chromatography (CCC) originated from the helical coil planet centrifuge. Recently, spiral coils were found to possess higher separation efficiency in both the retention of stationary phase and solutes resolution than other CCC coils like the helical and toroidal coils used on type-J CCC and cross-axis CCC. In this work, we built a novel conical coil CCC for the preparative isolation and purification of tanshinones from Salvia miltiorrhiza Bunge. The conical coils were wound on three identical upright tapered holders in head-to-tail and left-handed direction and connected in series. Compared with helical and spiral coil CCC, conical coil CCC not only placed CCC column in a two-dimensional centrifugal field, but also provided a potential centrifugal force gradient both in axial and radial directions. The extra centrifugal gradient made mobile phase move faster and enabled CCC much higher retention of stationary phase and better resolution. As a result, higher efficiency has been obtained with the solvent system of hexane–ethyl acetate–methanol–water (HEMWat) with the volume ratio of 5:5:7:3 by using conical coil CCC apparatus. Four tanshinones, including cryptotanshinone (1), tanshinone I (2), 1,2-dihydrotanshinquinone (3) and tanshinone IIA (4), were well resolved from 500mg to 1g crude samples with high purity. Furthermore, the conical coil CCC can make a much higher solid phase retention, which makes it to be a powerful separation tool with high throughput. This is the first report about conical coil CCC for separation of tanshinones and it may also be an important advancement for natural products isolation.

Centrifugal Partition Chromatography (CPC) as an Effective Method for Natural Products Purification

Natural products purification has often been a challenging task to many scientists worldwide. Many techniques have been invented and upgraded to make this task easier. This presentation will depict a few examples of natural products purification by using Centrifugal Partition Chromatography (CPC), which demonstrates a fast and effective way of natural products isolation. One example is CPC was applied to fraction F1 (hexane-ethyl acetate, 95:5) of hexane extract from Mesua elegans (Clusiaceae). Two 4-phenylcoumarins; mammea A/BA and mesuagenin C, was obtained from the fraction, with purity of 93.2% and 94.3% in 500 mg injection in multi-dual mode compare to 82.3% and 79.4% in isocratic mode. Another example is the development of a versatile tool for fast screening and rapid detection of bioactive natural products from plant extracts: the on-line coupling of CPC-UV to HPLC-UV-MS, via a six position switching valve. This strategy offers the possibility to get instant HPLC fingerprint of fractions and structural information about separated molecules during the CPC fractionation step. This new approach was applied to the fractionation and purification of xanthones from Garcinia mangostana (Clusiaceae) pericarp. The combined CPC-HPLC-DAD-MS allows the simultaneous fractionation, detection and characterization of sixteen molecules. Ten molecules were identified based on their UV and MS spectra. Furthermore, this method has led to the isolation of α-mangostin and γ-mangostin at 98.0% and 98.5% purity in a very short time.

A rugged and accurate liquid chromatography–tandem mass spectrometry method for the determination of asunaprevir, an NS3 protease inhibitor, in plasma

Asunaprevir (BMS-650032) is a potent hepatitis C virus (HCV) non-structural protein protease inhibitor currently in Phase III clinical trials for the treatment of HCV infection. A rugged and accurate LC-MS/MS method was developed and validated for the quantitation of asunaprevir in rat, dog, monkey, rabbit and mouse plasma. A systematic method screening and optimization strategy was applied to achieve optimized mass spectrometry, chromatography, and sample extraction conditions. The validated method utilized stable-isotope labeled D9-asunaprevir as the internal standard. The samples were extracted by liquid-liquid extraction using 10% ethyl acetate in hexane. Chromatographic separation was achieved with gradient elution on a Waters Atlantis dC18 analytical column. Analyte and its internal standard were detected by positive ion electrospray tandem mass spectrometry. The standard curve, which ranged from 5.00 to 2000ng/mL for asunaprevir, was fitted to a 1/x(2) weighted linear regression model. The intra-assay precision was within ±3.6% CV, inter-assay precision was within ±4.0% CV, and the assay accuracy was within ±8.1% of the nominal values in all the species. The method was successfully applied to support multiple pre-clinical toxicokinetic studies in different species.

Determination of ten pesticides of pyrazoles and pyrroles in tea by accelerated solvent extraction coupled with gas chromatography-tandem mass spectrometry

An effective method was developed and applied to determine the residues of ten pesticides of pyrazoles and pyrroles in tea by accelerated solvent extraction coupled with gas chromatography-tandem mass spectrometry (ASE-GC-MS/MS). The samples were extracted with ethyl acetate-hexane (1:1, v/v) for 5 min at 1.03 x 10(7) Pa and 100 degree C for one cycle. Then, they were purified by Envi-Carb/PSA column, and eluted by ethyl acetate-hexane (1:1, v/v). The analytes were determined by GC-MS/MS and quantified by external standard method. The limits of quantification were 0.003 mg/kg for fenpyroximate, 0.001 mg/kg for fipronil-sulfide, 0.002 mg/kg for fipronil, 0.005 mg/kg for fipronil-sulfone, 0.002 mg/kg for chlorfenapyr, 0.006 mg/kg for flusilazole, 0.001 mg/kg for difenzoquat, 0.001 mg/kg for pyraflufen-ethyl, 0.000 3 mg/kg for tebufenpyrad and 0.005 mg/kg for tolfenpyrad. The results show that the proposed method is sensitive and accurate for the determination of the ten pesticide residues.

Studies on Antimicrobial Activity and Phytochemical Profile of Mangifera Indica Leaf Extract

The present study deals with Studies on antimicrobial activity and photochemical profiles of Mangifera Indica (kottukonamvarika) leaf extract. The leaf of mangifera indica; studies with the aim of determined the following photochemical parameters for these species microscopial characters. Leaf contents, photochemical content, about 6 extract (methanol, acetone, hexane, ethyl acetate, hexane-ethyl acetate, aqueous extraction) from the leaves of Mangifera indica was extracted. Hexane-ethyl acetate was characterized by Gas Chromatography- Mass Spectroscopy. The antimicrobial activity of different extract was tested against human and plant pathogenic bacteria. Hexane-ethyl acetate extract showed significant role on inhibiting almost all tested pathogenic organisms. I. Introduction: The plant materials were collected from the village dihiya (District Rewa m.p.). In the month of January 2013. The plant was identified by local people of that village; A herbarium specimen of the plant was preserved in the Department of Environmental biology in Biotechnology Lab from APS University Rewa m.p. of our Institute for reference. The leaves use of medicinal plants as herbal remedies to prevent and cure several ailments differs from community to community Traditionally used medicinal plants have recently attracted the attention of the biological scientific communities. This has involved the isolation and identification of secondary metabolites produced by plants and their use as active principles in medicinal preparations plants have limitless ability to synthesize aromatic secondary metabolites, most of which are phenols or their oxygen-substituted derivatives. Extraction methods involve separation of medicinally active fractions of plant tissue from inactive/inert components by using selective solvents and extraction technology. Solvents diffuse into the solid plant tissues and solubilize compounds of similar polarity. Mango (Mangifera indica L.) is one of the most important tropical plants in the world. It grows in the tropical and subtropical regions and its parts are commonly used in folk medicine for a wide variety of remedies. Many phenolic compounds have been detected in mango peels mango bark 8, mango puree concentrate mango pulps and seed kernels. Several pharmacological activities of mango extracts have been reported including anti-inflammatory 6, antioxidant antiallergic and antihelmintic and antiamoebic. In the present study, certain works such as phyto-chemical characterization, antimicrobial activity of extract, histochemical studies, and cytotoxic effects of hexane-ethyl acetate extract against cell lines. II. Materials And Methods:- Plant Materials :-