Exploring the Q-marker of "sweat soaking method" processed radix Wikstroemia indica: Based on the "effect-toxicity-chemicals" study.

BACKGROUND: Radix Wikstroemia indica (RWI), named "Liao Ge Wang" in Chinese, is a kind of toxic Chinese herbal medicine (CHM) commonly used in Miao nationality of South China. "Sweat soaking method" processed RWI could effectively decrease its toxicity and preserve therapeutic effect. However, the underlying mechanism of processing is still not clear, and the Q-markers database for processed RWI has not been established. PURPOSE: Our study is to investigate and establish the quality evaluation system and potential Q-markers based on "effect-toxicity-chemicals" relationship of RWI for quality/safety assessment of "sweat soaking method" processing. METHODS: The variation of RWI in efficacy and toxicity before and after processing was investigated by pharmacological and toxicological studies. Cytotoxicity test was used to screen the cytotoxicity of components in RWI. The material basis in ethanol extract of raw and processed RWI was studied by UPLC-Q-TOF/MS. And the potential Q-markers were analyzed and predicted according to "effect-toxicity-chemical" relationship. RESULTS: RWI was processed by "sweat soaking method", which could preserve efficacy and reduce toxicity. Raw RWI and processed RWI did not show significant difference on the antinociceptive and anti-inflammatory effect, however, the injury of liver and kidney by processed RWI was much weaker than that by raw RWI. The 20 compounds were identified from the ethanol extract of raw product and processed product of RWI using UPLC-Q-TOF/MS, including daphnoretin, emodin, triumbelletin, dibutyl phthalate, Methyl Paraben, YH-10 + OH and matairesinol, arctigenin, kaempferol and physcion. Furthermore, 3 diterpenoids (YH-10, YH-12 and YH-15) were proved to possess the high toxicity and decreased by 48%, 44% and 65%, respectively, which could be regarded as the potential Q-markers for quality/safety assessment of "sweat soaking method" processed RWI. CONCLUSION: A Q-marker database of processed RWI by "sweat soaking method" was established according to the results and relationship of "effect-toxicity-chemicals", which provided a scientific evidence for processing methods, mechanism and the clinical application of RWI, also provided experimental results to explore the application of Q-marker in CHM.

Gastroprotective and anti-Helicobacter pylori potential of herbal formula HZJW: safety and efficacy assessment.

BACKGROUND: A traditional Chinese Medicine (TCM) formula, HZJW, has been applied in clinics in China for gastrointestinal disorders. However, the therapeutic mechanism underlying its efficacy and safety remained to be defined. The present investigation was undertaken to evaluate the formula HZJW for its gastroprotective potential, possible effect on Helicobacter pylori along with safety to justify its anti-ulcer action and safe clinical application. METHODS: The gastroduodenal cytoprotective potential was evaluated in rodent experimental models (HCl/Ethanol and NSAID-induced ulcer protocols). The anti-H. pylori property was assessed by agar dilution assay in vitro and analysis in vivo including rapid urease test, immunogold test and histopathology. For toxicity assessment, acute toxicity study was performed according to fixed dose procedure with a single oral administration of HZJW to mice. In the oral chronic toxicity, rats (80 males, 80 females) were administrated HZJW orally in 0, 1000, 2500, or 5000 mg/kg/day doses for 26 weeks (n = 40/group of each sex). Clinical signs, mortality, body weights, feed consumption, ophthalmology, hematology, serum biochemistry, gross findings, organ weights and histopathology were examined at the end of the 13- and 26-week dosing period, as well as after the 4-week recovery period. RESULTS: In the HCl/Ethanol-induced ulcer model, it was observed that oral administration with HZJW (260, 520 and 1040 mg/kg) and ranitidine (250 mg/kg) significantly reduced the ulcerative lesion index (116.70 ± 36.4, 102.20 ± 18.20, 84.10 ± 12.1 and 73.70 ± 16.70) in a dose-dependent manner, respectively, with respect to control group (134.10 ± 31.69). Significant inhibition was also observed in ulcerative index from aspirin-induced ulcer model, with decreases of 35.40 ± 5.93, 31.30 ± 8.08, 26.80 ± 8.27and 20.40 ± 6.93 for the groups treated with HZJW and ranitidine, in parallel to controls (41.60 ± 10.80). On the other hand, treatment with HZJW efficaciously eradicated H. pylori in infected mice in rapid urease test (RUT) and immunogold antibody assay, as further confirmed by reduction of H. pylori presence in histopathological analysis. In the in vitro assay, MICs for HZJW and amoxicillin (positive control) were 125 and 0.12 µg/mL respectively. The LD50 of HZJW was over 18.0 g/kg for mice. No drug-induced abnormalities were found as clinical signs, body weight, food consumption, hematology, blood biochemistry, ophthalmology and histopathology results across three doses. No target organ was identified. The No Observed Adverse Effect Level (NOAEL) of HZJW was determined to be 5,000 mg/kg/day for both sexes, a dose that was equivalent to 50 times of human dose. CONCLUSIONS: These results suggested the efficacy and safety of HZJW in healing peptic ulcer and combating H. pylori, which corroborated their conventional indications and contributed to their antiulcer pharmacological validation, lending more credence to its clinical application for the traditional treatment of stomach complaints symptomatic of peptic ulcer disease (PUD). HZJW might have the potential for further development as a safe and effective alternative/complementary to conventional medication in treating gastrointestinal (GI) disorders.

Root-derived auxin contributes to the phosphorus-deficiency-induced cluster-root formation in white lupin (Lupinus albus).

Formation of cluster roots is a typical morphological response to phosphorus (P) deficiency in white lupin (Lupinus albus), but its physiological and molecular mechanisms are still unclear. We investigated the role of auxin in the initiation of cluster roots by distinguishing the sources of auxin, measuring the longitudinal distribution patterns of free indole-3-acetic acid (IAA) along the root and the related gene expressions responsible for polar auxin transport (PAT) in different developmental stages of cluster roots. We found that removal of shoot apex or primary root apex and application of auxin-influx or -efflux transport inhibitors, 3-chloro-4-hydroxyphenylacetic acid, N-1-naphthylphthalamic acid and 2,3,5-triiodobenzoic acid, to the stem did not affect the number of cluster roots and the free-IAA concentration in the roots of P-deficient plants, but when these inhibitors were applied directly to the growth media, the cluster-root formation was greatly suppressed, suggesting the fundamental role of root-derived IAA in cluster-root formation. The concentration of free IAA in the roots was higher in P-deficient plants than in P-adequate ones, and the highest in the lateral-root apex and the lowest in the mature cluster roots. Meanwhile the expression patterns of LaAUX1, LaPIN1 and LaPIN3 transcripts related to PAT was consistent with concentrations of free IAA along the lateral root, indicating the contribution of IAA redistribution in the cluster-root development. We proposed that root-derived IAA plays a direct and important role in the P-deficiency-induced formation of cluster roots.

Relieving visceral hyperalgesia effect of Kangtai capsule and its potential mechanisms via modulating the 5-HT and NO level in vivo.

Kangtai capsule (KT) is one type of traditional Chinese medicine preparation derived from the proved recipe, which was frequently applied as an effective clinical treatment of IBS. However, there still lack the reasonable and all-round analytical approach and the scientific studies on its underlying mechanisms. Therefore, our study aimed to develop the novel method for evaluating its quality as well as to interpret the potential mechanisms. In our study, high performance liquid chromatography (HPLC) fingerprint was applied to provide a chemical profile of KT. The neonatal maternal separation (NMS) on Sprague-Dawley pups was employed to evaluate the therapeutic effect of KT by virtue of various parameters including visceral hyperalgesia, serum nitric oxide (NO) level, and tissue 5-hydroxytryptamine (5-HT) level. Consequently, a chromatographic condition, which was carried at 30°C with a flow rate of 0.5 ml/min on AQUA 3µ C18 column with mobile phase of acetonitrile and water-phosphoric acid (100:0.1, v/v), was established to give a common fingerprint chromatography under 254 nm with a similarity index of 0.963 within ten batches of KT samples. On the NMS model, KT markedly elevated the pain threshold of NMS rats. Furthermore, KT at three doses significantly decreased 5-HT content from distal colon of visceral hyperalgesia rats induced by NMS, while the significant decrease of 5-HT content in serum was only observed in the group with KT at high dose. However, compared with that in NMS rats without KT, there was no apparent difference of 5-HT level from brain issue in the rats with various doses. Besides, KT could substantially elevate the concentration of NO in the serum. The results showed our study developed the simple, rapid, accurate, reproducible qualitative and quantitative analysis by HPLC fingerprint for the quality control for KT. Data from the pharmacological investigation suggested that the curative effect of KT to the visceral hypersensitivity may be concerned with the level of 5-HT and NO in vivo, promising its potential in irritable bowel syndrome treatment.