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A quantitative analytical method of ultra-high performance liquid chromatography (UPLC) was developed for simultaneously determining twelve components in Tibetan medicine Zuozhu Daxi. SIMPCA 12.0 software was used a principal component analysis PCA) and partial small squares analysis (PLSD-DA) on the twelve components in 10 batches from four pharmaceutical factories. Acquity UPLC BEH C15 column (2.1 mm x 100 mm, 1.7 µm) was adopted at the column temperature of 35 °C and eluted with acetonitrile (A) -0.05% phosphate acid solution (B) as the mobile phase with a flow rate of 0. 3 mL · min(-1). The injection volume was 1 µL. The detection wavelengths were set at 210 nm for alantolactone, isoalantolactone and oleanolic; 260 nm for trychnine and brucine; 288 nm for protopine; 306 nm for protopine, resveratrol and piperine; 370 nm for quercetin and isorhamnetin. The results showed a good separation among index components, with a good linearity relationship (R2 = 0.999 6) within the selected concentration range. The average sample recovery rates ranged between 99.44%-101.8%, with RSD between 0.37%-1.7%, indicating the method is rapid and accurate with a good repeatability and stability. The PCA and PLSD-DA analysis on the sample determination results revealed a great difference among samples from different pharmaceutical factories. The twelve components included in this study contributed significantly to the quantitative determination of intrinsic quality of Zuozhu Daxi. The UPLC established for to the quantitative determination of the twelve components can provide scientific basis for the comprehensive quality evaluation of Zuozhu Daxi.
OBJECTIVE: To study the material composition of Tibetan medicine "Brag-zhun", and to provide the basis for interpretation of its source and reference for the establishment of quality standards.METHODS: Pharmacognosy routine method was used in macroscopic and microscopic identifications, inductively coupled plasma spectrometry was carried out to determine 26 kinds of mineral elements,and X-ray diffraction analysis was used for the crystalline phase detection of Brag-zhun. The content of calcium,organic matter, humic acid and fulvic acid was determined according to GB/T 14610-2008, "technical specification for soil analysis" , GB/T 11957-2001 and capacity titration, respectively. Moisture, ash content, acid insoluble ash content and water soluble extract were determined with reference to the Chinese Pharmacopoeia (2010 edition).
RESULTS: Animal feces were commonly found in Brag-zhun, the main crystal phase was SiO2, 26 kinds of mineral elements content in total was about 3%, 13 batches of medicinal materials in organic matter content was 29.03%-71.79%. Humic acid content was 28.37%-58.51%; fulvic acid content was 19.69%-41.43%; moisture was 5.2% -11.7%; total ash content was 17.82%-64.39%; acid insoluble ash content was 2.10%-39.09%; and water soluble extract was 28.39%-57.40%.
CONCLUSION: The traditional record of Brag-zhun from molten juice is lack of scientific basis. The mineral elements in Brag-zhun exist in amorphous. Brag-zhun contains a lot of organic matter including humic acid and fulvic acid, and inorganic matter mainly comes from sediment. The organic matter in rock has gone through a long geological evolution.
Zuotais regarded as the king of Tibetan medicine. However, the major starting material ofZuotais mercury, which is one very toxic heavy metal. This has aroused serious doubts on the biosafety ofZuotacontaining drugs. In this study, we quantified the Hg contents in fourZuotasamples, monitored the release of Hg in simulated gastric/intestinal juice and evaluated their cytotoxicity to Caco-2 cells. Our results showed that the Hg contents inZuotasamples were in the range of 566–676 mg/g. Fortunately, the release of Hg fromZuotasamples was very low in simulated gastric juice, and much lower in simulated intestinal juice. Direct contact ofZuotawith Caco-2 cells led to dose-dependent cytotoxicity, including activity loss and membrane leakage. The toxicity was closely related to apoptosis, because the caspase 3/7 levels of Caco-2 cells increased after the exposure toZuota. Interestingly,Zuotasamples inhibited the oxidative stress at low concentrations, but the toxicity could be relived by antioxidants. The possible toxicity should be attributed to the cellular uptake ofZuotaparticulates. Beyond the cytotoxicity, significant differences amongZuotasamples from different institutions were observed, suggesting that the preparation process ofZuotahad meaningful influence of its biosafety. The implications to the safety and clinical applications ofZuotaare discussed. [ABSTRACT FROM AUTHOR]