Development of UPLC Fingerprint with Multi-Component Quantitative Analysis for Quality Consistency Evaluation of Herbal Medicine "Hyangsapyeongwisan".

Hyangsapyeongwisan (HSPWS), known as traditional herbal medicine, has been used in the treatment of gastric disease. Standardization of HSPWS is a necessary step for the establishment of a consistent biological activity for the production and manufacturing of HSPWS herbal preparations. A simple, sensitive and accurate method using ultra-performance liquid chromatography (UPLC) fingerprinting with a diode array detector (DAD) was developed and validated for systematic quality evaluation of HSPWS. Separation conditions were optimized using a Halo C18 2.7 µm, 4.6 × 100 mm column with a mobile phase of 0.1% phosphoric acid and acetonitrile, and detection wavelengths of 215, 250 and 350 nm. Validation of the analytical method was evaluated by tests of linearity, precision, accuracy and robustness. All calibration curves of components showed good linearity (R(2) > 0.9996). The limit of detection (LOD) and limit of quantification (LOQ) were within the ranges of 0.004-0.134 and 0.012-0.406 µg/mL, respectively. The relative standard deviation (RSD) values of intra- and inter-day testing were within the range of 0.01-3.84%. The result of the recovery test was 96.82-104.62% with an RSD value of 0.14-3.84%. Robustness values of all parameters as well as the stability test of analytical solutions were within the standard limit. It showed that the developed method was simple, specific, sensitive, accurate, precise, reproducible and robust for the quantification of active components of HSPWS. Chromatographic fingerprinting with quantitative analysis of marker compounds in HSPWS prepared by different methods and commercial formulation was also evaluated successfully.

Evaluation of in vitro and in vivo biological activities of Cheilanthes albomarginata Clarke.

BACKGROUND: The Cheilanthes albomarginata Clarke (CA), a fern belonging to Pteridaceae family, is found mainly in India, Nepal, Pakistan and Bhutan at an altitude of 1300-2700 m. It grows mostly in the rock crevices on slopes. Juice from the rhizome of CA has been used to treat peptic ulcer. In this study, the biological activities (antioxidant, anti-inflammatory, anti-adipogenic and anti-obesity) of the extracts of CA were investigated. The total phenolic content of each extract was quantified. This is the first report regarding the study of biological activities on CA. METHODS: In the current study, the crude methanol and fractionated extract of the aerial part of CA were investigated for the antioxidant tests which were namely DPPH assay, hydrogen peroxide scavenging assay and nitrite scavenging assay. Their phenolic contents were measured by the Folin-Ciocalteu's method.In vitro anti-inflammatory and anti-adipogenic assays were evaluated against the RAW 264.7 macrophage cells and 3 T3-L1 cells respectively. The crude methanol extract and phenolic fraction (combination of ethyl acetate and butanol fraction) were studied for the in vivo anti-obesity test using male Sprague Dawley rats. RESULTS: The ethyl acetate fraction showed the strongest DPPH radical scavenging (82.54 ± 0.48%), hydrogen peroxide scavenging (3.41 ± 0.21 mg/ml) and nitrite scavenging activity (61.39%). The highest phenolic content was found in the ethyl acetate fraction followed by the butanol fraction. The ethyl acetate fraction showed the highest in vitro anti-inflammatory and anti-adipogenic activities. From the in vivo study on rats, the crude methanol extract and phenolic fraction showed plasma triglyceride lowering activity as well as reduction of weight of adipose tissue in high fat diet induced obese rats. CONCLUSION: The current study suggests that the ethyl acetate and butanol extracts of CA are potential source for antioxidant, anti-inflammatory and anti-adipogenic remedies. In addition to that the results of in vivo studies evidenced the possibility of CA as a source of anti-obesity drug remedies.