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The present study analyzed and identified the chemical constituents from ethyl acetate(EA) extract of Taxilli Herba with UPLC-Q-Exactive-MS and screened active xanthine oxidase(XO) inhibitors with HPLC. The analysis was performed on an Hypersil GOLD C_(18) reversed-phase column(2.1 mm×50 mm, 1.9 μm), with the mobile phase of water containing 1% formic acid(A) and methanol(B) under gradient elution, the flow rate of 0.3 mL·min~(-1), and the injection volume of 5 μL. ESI source was used for MS and the compounds were collected in positive and negative ion modes. Xcalibur 4.1 was used to analyze the retention time, accurate relative molecular weight, and fragmentation of the compounds. The inhibitory activity of some known compounds on XO was screened by HPLC. Thirty chemical constituents were identified, including phenolic acids and flavonoids by experimental data combined with information of standards, data reported previously, and databases, such as MzCloud and ChemSpider. The activities of 10 chemical components were screened. Gallic acid and naringenin chalcone had strong inhibitory activities on XO with IC_(50) of 57 μg·mL~(-1) and 108 μg·mL~(-1). UPLC-Q-Exactive-MS allows the accurate, rapid, and comprehensive identification of main chemical constituents from Taxilli Herba. Gallic acid and naringenin chalcone may be the active components of XO inhibitors.
Subject(s)
Acetates , Chromatography, High Pressure Liquid , Drugs, Chinese Herbal/chemistry , Tandem Mass Spectrometry , Xanthine OxidaseABSTRACT
Objective:To explore the acute toxicities and hepatotoxicities of aqueous extracts of Taxilli Herba from <italic>Morus alba</italic>， <italic>Toxicodendron</italic> <italic>trichocarpum</italic>， <italic>Camellia oleifera</italic>， <italic>Salix babylonica</italic>， <italic>Melia azedarach</italic>， and <italic>Nerium indicum</italic> against zebrafish model and the effect of different hosts on the toxicity of Taxilli Herba， hoping to provide a theoretical basis for the safe use of Taxilli Herba. Method:The normally developed AB zebrafish at 3-day post fertilization was selected for acute toxicity study. According to the results of preliminary toxicity experiments， the zebrafishes were treated with aqueous extracts of Taxilli Herba from different hosts at six doses， and their mortality was calculated 72 h later. GraphPad Prism 6.0 was used for plotting the dose-toxicity curve， followed by the calculation of their median lethal concentration （LC<sub>50</sub>） and 10% lethal concentration （LC<sub>10</sub>）. The gz15Tg/+（AB） liver fluorescent protein transgenic zebrafish with normal development at 4-day post fertilization was applied for the hepatotoxicity study. The zebrafishes were divided into the low-， medium-， and high-dose groups of aqueous extracts of Taxilli Herba from six hosts， the positive control （acetaminophen） group， and the blank （embryo amniotic fluid） group， and then treated with the corresponding drugs. Seventy-two hours later， the liver morphology and fluorescent area changes in zebrafish were observed. And the activities of alanine aminotransferase （ALT） and aspartate aminotransferase （AST） were detected. Result:The results of acute toxicity test demonstrated that the LC<sub>50</sub> values of water extracts of Taxilli Herba from <italic>M. alba</italic>， <italic>T.</italic> <italic>trichocarpum</italic>， <italic>C. oleifera</italic>， <italic>S. babylonica</italic>， <italic>M. azedarach</italic>， and <italic>N. indicum</italic> were 1.24， 0.94， 0.51， 0.38， 0.11， 0.09 g·L<sup>-1</sup>， respectively， and the LC<sub>10</sub> values were 0.70， 0.60， 0.35， 0.28， 0.08， 0.07 g·L<sup>-1</sup>， respectively. As revealed by hepatotoxicity test， compared with the blank group， the positive control group exhibited liver morphological changes， decreased fluorescent area （<italic>P</italic><0.01）， and elevated ALT and AST activities （<italic>P</italic>< 0.01）， suggesting that acetaminophen was hepatotoxic to zebrafish. However， there was no change in the liver morphology or fluorescent area of zebrafish in the low-， medium-， and high-dose groups of water extracts of Taxilli Herba from <italic>M. alba</italic>， and the ALT and AST activities were decreased. By contrast， the liver morphology and fluorescent areas in the medium- and high-dose groups of water extracts of Taxilli Herba from <italic>T.</italic> <italic>trichocarpum</italic>， <italic>C. oleifera</italic>， <italic>S. babylonica</italic>， <italic>M. azedarach</italic>， and <italic>N. indicum</italic> changed to varying degrees （<italic>P</italic><0.05， <italic>P</italic><0.01）. Besides， the activities of both ALT and AST were also enhanced. These indicated that Taxilli Herba from <italic>M. alba</italic> had no hepatotoxicity to zebrafish， while that from <italic>T.</italic> <italic>trichocarpum</italic>， <italic>C. oleifera</italic>， <italic>S. babylonica</italic>， <italic>M. azedarach</italic>， and <italic>N. indicum</italic> showed varying degrees of hepatotoxicity to zebrafish. Conclusion:The toxicity of Taxilli Herba is host-dependent. Taxilli Herba from <italic>M. alba</italic> has no hepatotoxicity， but that from the other five hosts shows varying degrees of hepatotoxicity. Standardizing the host source may be an important measure to realize the medication safety of Taxilli Herba.
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Objective: To screen the extraction process of Wuzi Yanzong plus and minus Recipe (WYR) by pharmacodynamics and long-term toxicity test, and optimize it by orthogonal experiment to determine the best extraction process of WYR. Methods Long-term toxicity experiments were carried out on different processed WYR samples. The penile erection latency and sex hormone levels in rats with kidney-yang deficiency were used as pharmacodynamic indicators. The results of long-term toxicity and efficacy experiments were combined to screen out the optimal process plan. Using the content of ingredients and the rate of ointment as indicators, the orthogonal test was used to screen the optimal level of different extraction processes and verified. Results Long-term toxicity test results showed that all mice survived healthily and no obvious toxicity was observed. The results of pharmacodynamic experiments showed that the extracts of Morindae Officinalis Radix, Eucommiae Cortex, Taxilli Herba, Dipsaci Radix, and Glycyrrhizae Radix et Rhizoma Praeparata cum Melle were refluxed with 70% ethanol, and the remaining medicinal materials were extracted with 70% ethanol as the solvent to obtain the best extraction method of WYR. The suitable extraction process was as follows: Morindae Officinalis Radix, Eucommiae Cortex, Taxilli Herba, Dipsaci Radix, and Glycyrrhizae Radix et Rhizoma Praeparata cum Melle were extracted three times with 10 times 70% ethanol for 2.0 h each time; The remaining medicinal materials were percolated with 10 times 70% ethanol at 1.5 mL/min. The medicinal herbs were firstly soaked for 16 h before percolation. Conclusion WYR can significantly improve the penile erection latency and sex hormone levels in rats with kidney yang deficiency. The optimal process conditions are reasonable and feasible, which provides a basis for the follow-up development of WYR.
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AIM To observe the effects of Taxilli Herba from six different hosts (Morus alba L.,Salix babylonica L.,Camellia oleifera Abel.,Castanea mollissima B1.,Liquidambar formosana Hance and Nerium indicum Mill.) on lowering blood pressure of spontaneously hypertensive rats (SHR).METHODS SHR were randomly divided into 14 groups,captopril positive group (20 mg/kg),model group,and Taxilli Herba groups of 6 different hosts,and each Taxilli Herba group was further divided into high-dose group (5.9 g/kg) and low-dose group (1.48 g/kg);WKY,in addition,was the blank control.And the 20-day consective correspondence medication was applied to the groups,each with eight rats.The caudal arterial systolic blood pressure (SBP) was measured by tail-cuff method before the administration,and on the 10th day and 20th day of the administration.Anaesthesia was performed at the blood collection 12 h after the last administration;and thus final serum contents of nitric oxide (NO) and changes of superoxide dismutase (SOD) activity,plasmatic contents of angiotensin Ⅱ (Ang Ⅱ) and endothelin-1 (ET-1) were determined.RESULTS From the data before and after administration,an SBP drop among all SHR groups was observed on the 10th day of administration,among which the blood pressure lowering effect by high-dose Taxilli Herba from Morus alba L.was very obvious (P ＜ 0.01);remarkable SBP decrease on the 20th day of administration induced by Taxilli Herba from Salix babylonica L,Liquidambarformosana Hance and Camellia oleifera Abel,and high-dose Taxilli Herba from Morus alba L,low-dose Taxilli Herba from Castanea mollissima B1 were detected (P ＜ 0.01).No significant SBP variation was available between the model group and Taxilli Herba groups after10-day administration;all the Taxilli Herba groups exhibited obvious effect in lowering SBP except Taxilli Herba from Nerium indicum Mill,low-dose Taxilli Herba from Morus alba L.and high-dose Taxilli Herba from Castanea mollissimaBl.after 20-day administration,compared to the model group (P ＜ 0.05).Highdose Taxilli Herba from Morus alba L.and Salix babylonica L significantly decreased plasmatic Ang Ⅱ and ET-1 contents of SHR groups in comparison with the model group (P ＜0.01).High-dose Taxilli Herba from Morus alba L.,Salix babylonica L.and Liquidambarformosana Hance significantly increased serum NO release and superoxide dismutase (SOD) activity (P ＜ 0.01).CONCLUSION Taxilli Herba from the five different hosts,except Nerium indicum Mill,can lower blood pressure,and there exists an effect difference due to the host variation.
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Objective: To observe the effects of Gentianae Macrophyllae Radix (Qinjiao) in different combinations on the expression of MMP-3, TIMP-1, and ankle pathological changes in rheumatoid arthritis (RA) model rats, and to explore the relationship between medicinal properties-disease-efficacy and explore the mechanism of traditional Chinese medicine in the treatment of RA. Methods: Eighty SD rats were randomly divided into blank group, collage II model group, rheumatic fever model group, tripterygium group, single Qinjiao group, Qinjiao-Weilingxian (Clematidis Radix et Rhizoma) group, Qinjiao-Sangjisheng (Taxilli Herba) group, and Qinjiao-Fangji (Stephanlae Tetrandrae Radix) group. Rheumatic fever arthralgia model was induced by collage II and exposed in rheumatic fever environment. After modeling, each administration group was ig administrated with corresponding drug solution. The paw thickness was observed every 3 d, and swelling was calculated; The arthritis index was scored in the early, middle, and late stages. Rats were sacrificed on day 39. The expression of MMP-3 and TIMP-1 in the ankle joint of rats was detected by immunohistochemistry, and the pathological changes of ankle joint were observed by HE staining. Results: Compared with the blank group, the paw edema, AI, and expression of MMP-3 in collage II model group and rheumatic fever model group was significantly higher, the expression of TIMP-1 were significantly lower, the articular surface of two groups was rough and damaged, and the articular cartilage was severely damaged. A large number of granulation tissues proliferated, accompanied by a large number of inflammatory cell infiltration and neovascularization. After treatment, compared with rheumatic fever model group, the paw edema, AI, and expression of MMP-3 in all treatment groups were reduced to varying degrees, in which the Qin-Fang group were lower most significant; The serum levels of TNF-α and IL-1β in all treatment groups were significantly decreased in varying degrees, especially in the Qin-Fang group; The expression of TIMP-1 was significantly increased in varying degrees, especially in the Qin-Fang group. HE staining results showed that the area of articular cartilage destruction and the degree of destruction were reduced, the inflammatory cells and neovascularization decreased, the number of repair fibers increased, and the scar tissue increased. Conclusion: For the rheumatic fever arthralgia RA, the combined effect of mild and cold Chinese materia medica (CMM) is better than the combined effect of mild and warm CMM, the combined effect of mild CMMs. Experimental results are consistent with the principles of traditional Chinese medicine clinical treatment of "treating the hot diseases should use the cold medicine", the mechanism of the compatibility drug relieving rheumatic fever arthralgia RA may be related to its ability, which can reduce the expression of MMP-3, increase the expression of TIMP-1, reduce articular cartilage destruction, and reduce inflammatory cell infiltration and vascular proliferation.
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OBJECTIVE: To investigate the microbial contamination in Anemones Raddeanae Rhizoma and Taxilli Herba to provide a reference for microbiological examination of Chinese herbal medicines. METHODS: Anemones Raddeanae Rhizoma and Taxilli Herba were analyzed according to the Chinese Pharmacopoeia 2015 Edition (publicity draft) and the methods of National Institutes for Food and Drug Control for thermoduric bacteria and specified microoganisms. RESULTS: The microbial contamination in 23 batches samples of the two Chinese herbal medicines was serious. CONCLUSION: There is an urgent need to control the microbial contamination of Chinese herbal medicines and Chinese medicines to improve their quality.