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Direct acid hydrolysis of Dioscorea zingiberensis rhizomes for preparation of diosgenin is wildly used in the traditional industry， which uses a large amount of inorganic acid catalysts， with high wastewater discharge and serious environmental pollution. Therefore， exploring clean and efficient preparation methods and processes has become an inevitable choice to realize the sustainable development of industrial production of diosgenin. Herein， the author reviewed and analyzed the research progress and problems of enzymatic hydrolysis， microbial transformation and modified acid hydrolysis in the preparation of diosgenin from D. zingiberensis rhizomes during the last ten years， and their application prospects are analyzed. Enzymatic hydrolysis has mild reaction conditions， but the yield of diosgenin is low， the economic cost is high， and the purification process of active enzyme is complicated. Microorganism shows specific activity to the substrate and high efficiency for diosgenin production， and microbial transformation is clean and environmentally friendly， but microbial transformation is time-consuming and the metabolic intermediates are complicated. For the modified acid hydrolysis， two-phase acid hydrolysis can reduce the amount of acid catalyst， and sulfonic acid-functionalized ionic liquid displays good recyclable performance by replacing the traditional inorganic acid， however， the wastewater discharge should still be considered. Solid acid catalysts are non-corrosive and easy to be recycled， but the need to use ethanol as the reaction solvent has certain safety hazards， and the catalyst preparation process is cumbersome. In conclusion， exploring clean and efficient conversion methods is an important research trend for preparation of diosgenin from D. zingiberensis rhizomes. For the enzymatic hydrolysis， the key glycoside hydrolases in the bioconversion process should be explored in depth， the conversion pathway of enzymatic saponins and enzyme specificity should be fully elucidated， and efforts should be made to improve the efficiency of enzymatic hydrolysis. For the microbial transformation， we should accelerate its industrial application process based on selecting and breeding efficient transformation strains， and optimizing stable transformation systems and processes， and in-depth investigation of the mechanism of microbial transformation， fully elucidating the specific key hydrolases and its catalytic properties， and striving to improve the efficiency of microbial transformation. For the modified acid hydrolysis， novel acid catalytic system with simple structure， stable performance and good biodegradability should be explored and applied， which can effectively solve the problems of environmental pollution and production safety.
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ObjectiveTo investigate the intervention effect of Qufeng Gutong Babu ointment （QFGT） on rats with osteoarthritis （OA） with cold-dampness obstruction， and preliminarily clarify its mechanism. MethodSD male rats were divided into 6 groups， namely， the blank group， model group， positive control drug Huoxue Zhitong ointment （HXZTG） group （1.26 cm2·d-1）， and low， medium， and high-dose QFGT group （75， 150， 300 mg·d-1）. OA model was prepared by joint cavity injection of papain and L-cysteine. On the second day of modeling， climate factors were applied to establish an animal model of combination of disease and syndrome of OA rats with cold-dampness obstruction. Standard VonFrey fiber was used to evaluate the threshold of mechanical pain. Weight bearing difference score and joint function score of both hind limbs were recorded. Hematoxylin-eosin （HE） staining and safranine fixation green staining were used to observe the pathological changes and cartilage degeneration of rat knee joint. Immunohistochemistry （IHC） was used to detect the expression of interleukin-1β （IL-1β）， interleukin-8 （IL-8）， tumor necrosis factor-α （TNF-α）， matrix metalloproteinase-9 （MMP-9）， and cathepsin K （CTSK）. Western blot was used to detect the protein expression of kinase B （Akt）， phosphorylated protein kinase B （p-Akt）， phosphatidylinositol 3-kinase （PI3K）， nuclear factor 1 （NFATc1）， MMP-9， and CTSK in T cells. ResultCompared with the normal group， the model group showed significant mechanical pain sensitivity reaction after modeling （P<0.01）， and the weight bearing difference of both hind limbs and joint function score were significantly increased （P<0.05， P<0.01）. Compared with the model group， both the high-dose QFGT group and the HXZTG group significantly reduced the mechanical pain sensitivity， weight difference， and joint function score of rats （P<0.05， P<0.01）， and the medium-dose QFGT group also improved the joint function to a certain extent， and the degeneration of the knee joint cartilage of rats was significantly reduced （P<0.05， P<0.01）. QFGT and HXZTG both inhibited the protein expression of IL-1β， IL-8， TNF-α， MMP-9， CTAK， PI3K， p-Akt， Akt， and other related proteins in articular cartilage of rats with OA to a certain extent （P<0.05， P<0.01）. ConclusionQFGT can inhibit the release of inflammatory factors and matrix metalloproteinases by inhibiting the PI3K/Akt signal pathway in articular articular cartilage of rats with OA with cold-dampness obstruction， thus ultimately weakening local cartilage degeneration and improving joint function.
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OBJECTIVE To study the spectru m-toxicity relationship of in vitro hepatotoxicity of aqueous extract from Euodia rutaecarpa. METHODS The aqueous extract from 16 batches of E. rutaecarpa from different habitats were prepared. The fingerprints of aqueous extract from E. rutaecarpa were established by ultra high performance liquid chromatography （UPLC） method and Similarity Evaluation System of TCM Fingerprint （2012A edition ），and common peaks were identified and the similarity was evaluated. Using normal human hepatocytes L 02 as subject ，inhibitory effect of aqueous extract from 16 batches of E. rutaecarpa to them were investigated. The spectrum-toxicity relationship of UPLC fingerprint of aqueous extract from E. rutaecarpa with the hepatotoxicity of hepatocytes L 02 was analyzed by grey relational analysis （GRA）and partial least squares regression analysis （PLSR）. The corresponding compound of the chromatographic peak with the greatest correlation with the in vitro hepatotoxicity of E. rutaecarpa were isolated ，prepared and identified. RESULTS There were 27 common peaks in UPLC fingerprints of aqueous extract from 16 batches of E. rutaecarpa ，with similarity of 0.375-0.995. Totally 9 peaks were confirmed ，i.e. neochlorogenic acid （peak 5），chlorogenic acid （peak 9），cryptochlorogenic acid （peak 10），caffeic acid （peak 12），rutin （peak 16），hyperin（peak 17），dehydroevotarine（peak 19），evotarine（peak 24），rutecarpine（peak 25）. The aqueous extract from 16 batches of E. rutaecarpa showed significant inhibitory effect on the growth of L 02 cells（P＜0.05 or P＜0.01），and the inhibitory rate ranged from 6.68％ to 67.95％. GRA showed that there were 18 common peaks with correlation degree greater than 0.8，which were peak 8＞peak 3＞peak 23＞peak 7＞peak 4＞peak 9＞peak 12＞peak 2＞peak 19＞peak 6＞ 4928381。E-mail：799247687@qq.com peak 15＞peak 5＞peak 1＞peak 17＞peak 21＞peak 26＞ peak 20＞peak 14 in descending order of correlation degree. PLSR showed that there were 14 peaks with regression coefficient＞0 and variable importance projection value ＞1，and the order of regression coefficient was peak 8＞peak 3＞peak 23＞ peak 2＞peak 7＞peak 4＞peak 12＞peak 9＞peak 19＞peak 5＞peak 17＞peak 26＞peak 10＞peak 15. Peak 8 had the greatest correlation with in vitro hepatotoxicity，and the corresponding compound of this peak was identified as 6-O-trans caffeoyl gluconic acid. CONCLUSIONS The in vitro hepatotoxicity of aqueous extract from E. rutaecarpa is the result of multiple component interaction，among which 6-O-trans caffeoyl gluconic acid shows closest relation with in vitro hepatotoxicity.
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Objective To report the experiences on the diagnosis and treatment of duplication of kidney ureter and bladder. Methods Nine cases of duplication of kidney ureter and bladder from 1996 to 2008 were reviewed.Six cases of duplicated kidney ureter occurred in the left side and 2 cases in the right side,1 case bilateral kidney ureteral duplication.Of 8 cases with unilateral duplicated,duplicated bladders were incompleteness.And the patient with bilateral duplicated,whose duplicated bladder was completeness,was diagnosed with duplication of urethra,uterine,bilateral ovary and oviduet tubes,and also suffered from duplicated uterine prolapse Ⅱ,vaginal anterior wall bulging and duplicated vesicocele.There were two cases whose duplicated kidney losed function because of severe hydronephrosis,and 7 cases existed kidney secretion function. Excision of duplicated kidney ureter and bladder were performed on 2 cases with non-functional duplicated kidney.6 cases had undergone duplicated bladder excision and duplicated ureteral bladder replantation.The special case had undergone duplicated urinary bladder urethra uterine and bilateral annexes excision,and duplicated ureteral bladder replantation. Results The operation was successful in all paients without leakage and ureter stump syndrome.Three months after operation,ureter bladder imaging showed no ureteral reflux in 7 cases of ureteral bladder replantation.IVU were reviewed 12 months after operation:2 cases undergoing duplicated kidney excision showed that the function of residual kidney were normal,7 cases of replantation that the shape and function of sick side kidney and duplicated kidney were good. The patient who suffered from duplicated uterine simultaneously got pregnancy 1.5 years after operation. Conclsions Image examinations may help to diagnose the duplication of kidney ureter and bladder. The main treatment is surgery. Understanding the function of duplicated kidney and the shape of kidney ureter and bladder should be considered before operation. The goal of surgery should be relieving pain,protecting the function of duplicated kidney and minimizing the risk of infection.
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Objective To study the expression of kallikrein 7 (KLK7) in different prostate tissues and its clinical significance. Methods KLK7 mRNA levels in normal prostate epithelia (5 cases), benign prostat(ic) hyperplasia (BPH) epithelia (13 cases), prostate cancer and prostate cancer cell lines (8 cases) were analyzed by using semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Western blot was used to analyze the protein levels of human kallikrein 7 (hK7) in benign prostate epithelia and prostate cancer cell lines, hK7 expressions were examined in 20 normal prostate tissue specimens, 50 BPH specimens and 103 prostate cancer specimens by immunohistochemical staining.Results The mRNA levels of KLK7 in normal prostate, BPH and prostate cancer were 0.59, 0.52 and 0.02 respectively, mRNA levels of KLK7 were significantly different among the three groups (F=13.03, P＜0.01). mRNA levels of KLK7 were decreased in prostate cancers compared with that in benign hyperplastic prostate epithelial cells (P＜0.01) and in normal prostate epithelial cells (P＜0.01). No significant difference of KLK7 mRNA levels was found between normal prostate and BPH. The protein levels of KLK7 in normal prostate, BPH, DU145, LNCaP, PC3,22RV1 and BPH1 was 0.22, O. 40, 0.01, 0.05, 0, 0.03 and 0.14 respectively, hK7 protein level was down-regulated in prostate cancer cell lines compared to benign prostate epithelial cells. The expression of bK7 was observed in benign prostate epithelial cells, whereas little or no staining was observed in prostate cancer cells in immunohistochemical study, hK7 protein was detected in 13 of 20 (65%)normal prostate specimens, 38 of 50 (76%) BPH specimens and 18 of 103 (17.5%) prostate cancer specimens. The difference between the normal prostate and prostate cancer was significant (Z=-4.43, P＜0.01). The difference between BPH and prostate cancer was significant (Z=-7.77,P＜0.01) as well. However, no significant difference of hK7 protein level was found between normal prostate and BPH (Z=-1. 52, P＞0.05). Conclusions KLK7 expression level is down regulated in prostate cancer. KLK7 may play an important role in prostate cancer progression.