Matrix Solid-Phase Dispersion Extraction and Quantification of Alpinetin in Amomum Seed using Validated HPLC and HPTLC Methods

Alpinetin is a flavonoidal constituent of seeds of Amomum subulatum Roxb., recently reported to possess vasorelaxant and antiHIV activities. Simple, accurate and precise HPLC and HPTLC methods were developed for the analysis of alpinetin in A. subulatum seed extracts and extraction technique was optimized to get maximum yield using conventional, ultrasonic and matrix solid phase dispersion extraction. HPLC was performed on a C18 column with methanol and water (70:30, v/v) as mobile phase at a flow rate of 1.0 ml/min whereas HPTLC on silica aluminum sheet (60F₂₅₄) using toluene, dichloromethane and ethyl acetate as solvent system. A sharp peak was obtained for alpinetin at a retention time (R_t) of 5.7 min by HPLC and retardation factor (R_f) of 0.48 by HPTLC. Both methods were validated as per the ICH guidelines and the content of alpinetin was estimated in different extracts. Matrix solid phase dispersion technique was found most suitable for extracting alpinetin as compared to other techniques. Validation data are indicative of good precision and accuracy and proved the reliability of the methods.     

Antidiabetic,antioxidant, molecular docking and HPTLC analysis of miquelianin isolated from Euphorbia schimperi C. Presl

The present study demonstrates the miquelianin or quercetin 3-O-glucuronide (compound 1) isolated from aerial parts of Euphorbia schimperi exhibited significant results for antioxidant and antidiabetic potential. The compound 1 along with kaempferol 3-O-glucuronide (compound 2) and quercetin 3-O-rhamnoside (compound 3) isolated from the same source were quantified by validated HPTLC method. Antioxidant activity was determined by chemical means in terms of ABTS radical cation and DPPH radical scavenging activity. Compound 1 showed significant scavenging activity in both ABTS and DPPH assays as compared to standard BHA. In ABTS method IC₅₀ values of compound 1 and standard BHA is found to be 58.90 ± 3.40 µg/mL and 28.70 ± 5.20 µg/mL respectively while in DPPH assay IC₅₀ values of Compound 1 and standard BHA is 47.20 ± 4.90 µg/mL and 34.50 ± 6.20 µg/mL respectively. Antidiabetic effect was studied through α-amylase and α-glucosidase inhibitory activity. The mechanistic approach through molecular modelling also support the strong binding sites of compound 1 which showed significant α-amylase and α-glucosidase inhibitory activities with IC₅₀ values 128.34 ± 12.30 and 89.20 ± 9.20 µg/mL respectively as compared to acarbose 64.20 ± 5.60 and 52.40 ± 4.60 µg/mL respectively. The results of validated RP-HPTLC analyses revealed the concentration of compound 1 found to be 16.39 µg/mg and for compound 2 and compound 3 as 3.92 and 14.98 µg/mg of dried extract, respectively.     

Hypericum perforatum: Influences of the habitat on chemical composition,photo-induced cytotoxicity,and antiradical activity

Context: Hypericin, isolated from Hypericum perforatum L. and about another 300 Hypericum species (Guttiferae), is one of the most powerful photosensitizers found in nature.

Objective: The aim of this study was to assess the variability of chemical composition and biological activities of four H. perforatum samples, collected at different altitudes in the South Apennine of Italy.

Materials and methods: MTT assay was used to evaluate the antiproliferative activity of different samples concentrations (0.6–100?µg/mL) after irradiation at 365?nm. The inhibition of nitric oxide production was evaluated after 24?h of incubation using the macrophage cell line RAW 264.7 and sample solutions ranging from 12.5 to 1000?µg/mL. Antioxidant activities were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay and β-carotene bleaching test (ranges were 12.5–1000 and 1–400?µg/mL, respectively). Chemical composition was evaluated through HPTLC, and different contents of hypericin and rutin have been observed.

Results: The most phototoxic sample was collected from Zumpano (no. 1 at 370?m), with IC₅₀ values of 24.61?±?0.02?μg/mL. Sample no. 1 showed also the best radical scavenging activity (IC₅₀ value of 9.18?±?0.03?μg/mL) and the best antioxidant activity (IC₅₀ value of 10.04?±?0.03?μg/mL after 30?min of incubation). Best activity of extract no. 1 was well in accordance with chemical data, including the phenolic total content and particular metabolome profile.

Discussion and conclusion: This paper confirms the usefulness in maintaining the exploration of H. perforatum activities, in order to confirm its potentiality as a multipurpose plant.     

Stability Indicating HPTLC Determination of Meloxicam

A simple, selective, precise and stability-indicating high-performance thin layer chromatographic method of analysis of meloxicam both as a bulk drug and in formulation has been developed. The mobile phase selected was ethyl acetate:cyclohexane:glacial acetic acid (6.5:3.5:0.02% v/v/v). The calibration curve of the drug was linear in the range of 100-500 ng. The spectrodensitometric analysis was carried out in the absorbance mode at 353 nm. The mean (±RSD) values of slope, correlation coefficient and intercept were 3183.8±0.358, 0.9996±0.0321 and 13012±7.1 respectively. The system precision and the method precision studies were carried out with RSD of 0.83 and 1.89 respectively. The limit of detection and quantitation were 30 ng and 99 ng respectively. The mean percent recovery was found to be 100.3%. The method was used to analyze meloxicam from marketed tablet formulation in the presence of commonly used excipients.     

新疆4种地产大蒜的高效薄层氨基酸分析

目的:使用高效薄层(HPTLC)方法对新疆地产4种大蒜中的氨基酸进行分析,控制大蒜质量。方法:将鲜蒜置于-80℃低温冰箱冷冻20min,切碎,真空冷冻干燥箱冻干24h,制成大蒜冻干蒜粉。将5mL甲醇加入1g冻干蒜粉中振摇1min,过滤,滤液点样20μL;展开剂∶正丁醇∶正丙醇∶冰醋酸∶水=3∶1∶1∶1;喷以0.2%茚三酮试剂,日光灯下检视,500nm处扫描吸收光谱。结果:得到大蒜中氨基酸类薄层色谱图,斑点清晰、分离度较好。结论:通过峰面积比较,分析不同产地大蒜中各类氨基酸成分有差异。     

Expression of Neutral Glycosphingolipids in the Brain and Spleen of Mice Lacking TNF Receptor 1

We investigated the expression of neutral glycosphingolipids (GSLs) in the brain and spleen of mice lacking the gene for the tumor necrosis factor‐α receptor p55 (TNFR1). Neutral GSLs of the ganglio‐, globo‐, and neolacto‐series were determined in the tissues of homozygous (TNFR1 ?/?) and control heterozygous (TNFR1 +/?) animals by high performance thin layer chromatography (HPTLC) overlay immunostaining with specific antibodies. The spleen of homozygous TNFR1 knockout mice lacked glucosylceramide substituted with palmitic acid, GlcCer(C16), and showed severe reduction in the expression of GlcCer(C24). In addition, gangliotetraosylceramide substituted with palmitic acid, Gg4Cer(C16), and globotetraosylceramide, Gb4Cer, were down‐regulated in the TNFR1 ?/? spleen in comparison with the heterozygous control. The brain of both groups of animals (TNFR1 ?/? and TNFR1 +/?) did not express detectable levels of Gg4Cer, Gb5Cer and Gb4Cer, but the brain of TNFR1 knockout mice expressed abundant globotriaosylceramide, Gb3Cer, compared to no expression in control heterozygous mice. nLcCer(C24) had slightly higher (1.4 fold) expression in the brain of TNFR1 ?/? mice compared with the control animals. This study provides in vivo evidence that TNF signaling via the TNFR1 is involved in the acquisition of a divergent GSL assembly in the brain, an immunologically privileged organ, and the spleen, typical secondary lymphoid organ.     

Standardization of a Traditional Polyherbo-Mineral Formulation-Brahmi Vati

The present study deals with standardization of an in-house standard preparation and three marketed samples of Brahmi vati, which is a traditional medicine known to be effective in mental disorders, convulsions, weak memory, high fever and hysteria. Preparation and standardization have been done by following modern scientific quality control procedures for raw material and the finished products. The scanning electron microscopic (SEM) analysis showed the reduction of metals and minerals (particle size range 2–5 µm) which indicates the proper preparation of bhasmas, the important ingredient of Brahmi vati. Findings of EDX analysis of all samples of Brahmi vati suggested the absence of Gold, an important constituent of Brahmi vati in two marketed samples. All the samples of Brahmi vati were subjected to quantitative estimation of Bacoside A (marker compound) by HPTLC technique. Extraction of the samples was done in methanol and the chromatograms were developed in Butanol: Glacial acetic acid: water (4.5:0.5:5 v/v) and detected at 225nm. The regression analysis of calibration plots of Bacoside A exhibited linear relationship in the concentration range of 50–300 ng, while the % recovery was found to be 96.06% w/w, thus proving the accuracy and precision of the analysis. The Bacoside A content in the in-house preparation was found to be higher than that of the commercial samples. The proposed HPTLC method was found to be rapid, simple and accurate for quantitative estimation of Bacoside A in different formulations. The results of this study could be used as a model data in the standardization of Brahmi vati.