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Candida tropicalis uses alkanes and fatty acids to produce long chain dicarboxylic acids. However, the yield can be affected by β-oxidation in peroxisomes. Pxa1p was a membrane protein of Saccharomyces cerevisiae peroxisomes. Pxa1p and Pxa2p form a dimer that is involved in transporting of long chain fatty acids into peroxisomes, but the similar transporting system of Candida tropicalis has not yet been reported. In this study, a ctpxa1 gene deletion strain named C. tropicalis 1798-pxa1 was constructed by homologous single exchange method using PCR fragment. The expression of ctpxa1 gene in C. tropicalis 1798, C. tropicalis 1798-pxa1 was detected by semi-quantitative RT-PCR, and the ratio of gray value was 2.03, implying that the expression of ctpxa1 in C. tropicalis 1798-pxa1 was weakened. After 144 h fermentation, the dodecanedioic acid production of C. tropicalis 1798-pxa1 was increased 94.3% than the former strain, the maximum yield was 10.3 g/L.
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OBJECTIVE:To study and compare the volatile constituents in Forsythia suspensa and old F. suspensa. METH-ODS:Steam distillation method was adopted to extract volatile oil. GC-MS was adopted to determine the content of volatile oil:the chromatographic conditions:the column was HP-5MS,inlet temperature was 200 ℃,the carrier gas was nitrogen,split ratio was 20 to 1,by programmed temperature,flow rate was 1 ml/min,the injection volume was 1 μl;MS conditions:ion source was electric bombardment ionization source,electron energy was 70 eV,the interface temperature was 280 ℃,ion source temperature was 230 ℃,quadrupole temperature was 150 ℃,electron multiplier voltage was 2 300 V,and mass scanning range was 29-350 amu. RESULTS:The extraction yield order of volatile oil was that F. suspensa heart was higher than F. suspensa,followed by F. suspensa shell and old F. suspensa;there were 18 common peaks in the volatile oil,and the content of β-pinene was the maxi-mum,followed by α-pinene,β-phellandrene,terpinen-4-ol and terpinene;in addition,4-methylene-1-cyclohexanone,3-cyclohex-ene-1-methanol,cumene formaldehyde,1,7-trimethyl-4-methylenedecahydro,palmitic acid,nutmeg aldehyde can only be detected in the volatile oil of old F. suspensa;while α-phellandrene,ocimene,α-terpineol,α-pinocarvone,cis-piperitol,bornyl acetate,α-cubebene and germacrene D can only present in F. suspensa. CONCLUSIONS:The main chemical components in the volatile oil of F. suspensa and old F. suspensa are basically the same,but there are also some differences. The discovery can provide some ba-sis for the material basic research in the pharmacodynamic differences of F. suspensa.
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Thioesterase catalyzes the hydrolysis of acyl-ACP and saturated fatty acyl chain. It plays a key role in the accumulation of medium chain fatty acids in vivo. In this study, to construct an engineering strain to produce MCFAs, the Arabidopsis acyl-ACP thioesterase gene AtFatA was amplified by PCR from cDNA of arabidopsis and double digested by EcoR I/Xba I, then linked to the plasmid digested with same enzymes to get the recombinant plasmid pPICZaA-AtFatA. We transformed the gene into Pichia pastoris GS115 by electroporation and screened positive colonies by YPD medium with Zeocin. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results showed that the recombinant enzyme had a molecular of 45 kDa band which was consistent with the predicted molecular mass and we constructed the expression system of gene AtFatA in fungus for the first time. Under shake-flask conditions, Gas Chromatograph-Mass Spectrometer-computer results indicated that recombinant strain produced 51% more extracellular free MCFAs than the wild and its yield reached 28.7% of all extracellular fatty acids. This figure is 10% higher than the control group. The result provides a new way to produce MCFAs.
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Arabidopsis , Genetics , Arabidopsis Proteins , DNA, Complementary , Electrophoresis, Polyacrylamide Gel , Electroporation , Pichia , Metabolism , Plasmids , Polymerase Chain Reaction , Recombinant Proteins , Thiolester Hydrolases , Transformation, GeneticABSTRACT
Objective To observe the therapeutic efficacy and safety of recombinant tissue plasminogen activator (rt-PA) for treatment of patients with acute cerebral infarction and investigate the prognostic factors of thrombolysis. Methods The clinical data of 60 patients with acute cerebral infarction from January 2009 to November 2013 in Department of Emergency, Hangzhou Traditional Chinese Medicine Hospital were retrospectively analyzed;of them, 30 cases received intravenous rt-PA thrombolytic treatment, being in the thrombolytic group, 0.9 mg/kg rt-PA was given to the patient within 4.5 hours after the disease onset, the total dosage could not exceed 90 mg, in which 10%was intravenously injected and the rest 90%was intravenously dripped slowly within 60 minutes. That another 30 cases did not undergo thrombolytic therapy was assigned as the control group, and they took aspirin, etc anti-platelet aggregation routine treatment. Before and after thrombolytic treatment for 1 hour, 24 hours and 14 days, the National Institutes of Health Stroke Scale (NIHSS) score was evaluated in the two groups;before and after thrombolytic therapy for 0.5, 1.0, and 1.5 hours, the patient's scores of Barthel index (BI) were observed in the two groups. In thrombolytic group, the situations of hemorrhage transformation, symptomatic hemorrhage and modified Rankin scale (mRS) score in 3 months of the patients with different ages, complications and NIHSS scores were observed. Results Before treatment, no statistically significant differences were found in the NIHSS score (15.2±3.6 vs. 15.5±3.3) and BI score (45.0±8.8 vs. 44.1±7.6) between the control group and thrombolytic group (both P>0.05);after treatment with the extension of time, the NIHSS score was gradually reduced, reaching the lowest level on the 14th day after thrombolytic treatment, while the BI score was gradually increased, reaching its peak at 1.5 hours after thrombolytic treatment, the changes being more prominent in thrombolytic group (NIHSS score:9.7±2.6 vs. 12.8±4.2, BI score:82.6±7.8 vs. 69.6±9.8, both P80 years old was higher than that in cases≤80 years old (15 vs. 12); the age (years: 71.0±4.1 vs. 61.5±2.6), baseline NIHSS score (14 vs. 11) and bleeding conversion rate [37.50% (3/8) vs. 18.18% (4/22)] of cases with atrial fibrillation were higher than those not complicated with atrial fibrillation; the NIHSS score of cases with elevated international normalized ratio (INR) was lower than those without elevated INR (11 vs. 14);The bleeding conversion rate [16.67%(1/6) vs. 29.17%(7/24)] with NIHSS score≤4 were lower than those with NIHSS score>4;the mRS score in 3 months (4 vs. 2), and percentage of 3-month mRS score≤2 [11.1%(1/9) vs. 52.38%(11/21)] in cases with NIHSS score≥20 was higher than that in cases with NIHSS score<20 (all P<0.05). The fatality rate of two groups was 3.33%. Conclusions The intravenous rt-PA thrombolytic treatment can significantly promote the early recovery of neurologic impairment for patients with acute cerebral infarction. The therapy can improve the prognosis and its safety is relatively good. Meanwhile it is similarly effective for cases over 80, with complications such as atrial fibrillation, raised INR, and with different degrees of severity.
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Objective To evaluate the pulmonary protection of dexmedetomidine in combination with parecoxib in pa?tients undergoing thoracotomy with one-lung ventilation. Methods Eighty patients undergoing elective resection of esopha?geal or lung cancer, including both sex, aged 40-70 yr, ASAⅠ-Ⅲ, were randomly divided into four groups (n=20), dexme?detomidine group (D group), parecoxib group (P group), dexmedetomidine in combination with parecoxib group (DP group) and control group (C group). Dexmedetomidine 1μg/kg was infused in ten minutes and then continued infusion at the rate 0.6μg·kg-1·h-1 until the chest was closed in group D. Parecoxib 40 mg was infused 10 min before the induction of anesthesia in group P. DP group was given parecoxib 40 mg and parecoxib 40 mg 10 min before the induction of anesthesia. The equal volume of normal saline was given in group C. Blood samples were collected for determination of blood gas analysis and the serum concentration of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-8 immediately after the induction of anes?thesia (T1), 30 min (T2) and 60 min(T3) after one-lung ventilation, and at the end of the operation (T4). Oxygenation index (OI) was calculated. The serum levels of TNF-α, IL-6 and IL-8 were detected by ELISA. Results Compared with time T0, the serum concentrations of TNF-α, IL-6 and IL-8 (except IL-8 at the time T2 in DP group) were significantly increased, and OI was decreased in all groups at the time T2-4 (P 0.05). Conclusion Combination of dexmedetomidine and parecoxib can further mitigate inflammatory response, improve lung oxygenation dur?ing one-lung ventilation, and provide pulmonary protection in patients undergoing thoracotomy.
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Objective To investigate the relationship between the changes in intracranial excitatory amino acid(EAA) levels and the incidence of postoperative cognitive dysfunction (POCD) in eldedy patients after general anesthesia. Methods Forty ASA Ⅰ-Ⅲ patients of both sexes aged 65-79 yr weighing 48-76 kg undergoing elective non-cardiac surgery under general anesthesia were studied.Anesthesia was induced with midazolam,fentanyl,etomidate and succinylcholine and maintained with continuous iv infusion of propofol,remifentanil,isoflurane inhalation and intermittent iv boluses of vecuronium.The right internal jugular vein was cannulated with a catheter which was advanced cephalad until jugular bulb.The jugular bulb venous blood samples were taken before (T0,baseline),at the end of (T1) and at 24,48 and 72h(T2-4) after operation for measurement of plasma concentrations of glutamate (Glu),aspartate (Asp) and glycine (Gly) by RP-HPLC.The cognitive function was evaluated by mini-mental state examination (MMSE) at T0 and T4.Results Eleven of the 40 patients developed POCD (28%).The plasma Gh,Asp and Gly concentrations were significantly increased after operation as compared with the baseline values in POCD patients and were significantly higher than in non-POCD patients.The plasma concentrations of Glu and Asp were negatively correlated with MMSE score(Glu:r=-0.86.P<0.01;Asp:r=-0.99,P<0.01).Conclusion Elderly patients may develop POCD after operation performed under general anesthesia through increase in intracranial EAA levels.
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The effect of cultivation conditions on the optical purity of L(+)-lactic acid produced by Rhizopus oryzae HZS6 was investigated. The isomeric composition of lactic acid was influenced by the supplementation of L(+)-lactic acid to fermentation medium. L(+)-isomer increased with the dosage, no D(-)-lactic acid was observed when the concentration of supplemented L(+)-lactic acid in matrix was > or = 1.5 g/L. However, the L(+)-lactic acid yield, biomass and glucose conversion rate decreased with the dosage. With the same method, the supplementation of L(+)-lactic to substrate had no influence on isomeric composition of lactic acid by Lactobacillus and Escherichia coli.
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Culture Media , Chemistry , Fermentation , Lactic Acid , Chemistry , Rhizopus , Metabolism , StereoisomerismABSTRACT
Objective To establilsh the quality standard for Qingfei Yihuo tablet. Methods TLC was performed to identify Sophora flavescens Ait, Peucedani and Cortex Phellodendri. HPLC was used to determine Baicalin. ODS column was used. The mobile phase consisted of methanol-water-phosphoric acid (47∶53∶0.2). The flow rate was 1.0 mL/min and column temperature was at 25 ℃. The detecting wave length was at 280 nm. Results The characteristic for identification by TLC was distinct and hightly specific. Baicalin could be determined by HPLC. The linearity of Baicalin was good in the range of 0.163～0.975 ?g (r=0.999 9). The average recovery of Baicalin was 97.43% and RSD was 1.97% (n=6). Conclusion The methods of identification and quantification is simple and reproducible. It can be used effectively for the quality control of Qingfei Yihuo table.
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AIM: To establish a determination for ephedrine hydrochloride and pseudoephedrine hydrochloride in Suxing Capsules(Herba Ephedrae, Semen Armeniacae Amarum, Fructus Perillal, etc.). METHODS:HPLC with Agilent ZORBAX Extend C_(18) (250 mm?4.6 mm, 5 ?m) column was used. Acetonitrile-0.2% H_3PO_4(5∶95) was used as a mobile phase. The flow rate was 1.0 mL?min~(-1). The detection wawelength was at 210 nm. RESULTS:The methodological evaluation showed that this method had a good repeatability and reproducibility; The average recovery of ephedrine hydrochloride was 99.65% and RSD was 1.6%. The average recovery of (pseudoephedr)ine hydrochloride was 98.90% and RSD was 2.3%. CONCLUSION: This method is simple, quick, accurate and with good reproducibility and can be used for the quality control of Suxing Capsules.
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0.05). Intrathecal administration of D-serine (0.025, 0.05, 0.1 ng), antagonized analgesia induced by isoflurane in tail-immersiontest and the early phase of formalin test (P0.05). Intrathecal administration of D-serine (0.05 ng) antagonized and the isoflurane′s suppression to the expression of Fos-protein in the spinal cord in mice(P