Highly efficient biocatalytic synthesis of L-DOPA using in situ immobilized Verrucomicrobium spinosum tyrosinase on polyhydroxyalkanoate nano-granules.

L-DOPA (3,4-dihydroxyphenyl-L-alanine) is a preferred drug for Parkinson's disease, and is currently in great demand every year worldwide. Biocatalytic conversion of L-tyrosine by tyrosinases is the most promising method for the low-cost production of L-DOPA in both research and industry. Yet, it has been hampered by low productivity, low conversion rate, and low stability of the biocatalyst, tyrosinase. An alternative tyrosinase TyrVs from Verrucomicrobium spinosum with more efficient expression in heterologous host and better stability than the commercially available Agaricus bisporus tyrosinase was identified in this study. Additionally, it was prepared as a novel nano-biocatalyst based on the distinct one-step in situ immobilization on the surface of polyhydroxyalkanoate (PHA) nano-granules. The resulting PHA-TyrVs nano-granules demonstrated improved L-DOPA-forming monophenolase activity of 9155.88 U/g (Tyr protein), which was 3.19-fold higher than that of free TyrVs. The nano-granules also exhibited remarkable thermo-stability, with an optimal temperature of 50 °C, and maintained more than 70% of the initial activity after incubation at 55 °C for 24 h. And an enhanced affinity of copper ion was observed in the PHA-TyrVs nano-granules, making them even better biocatalysts for L-DOPA production. Therefore, a considerable productivity of L-DOPA, amounting to 148.70 mg/L h, with a conversion rate of L-tyrosine of 90.62% can be achieved by the PHA-TyrVs nano-granules after 3 h of biocatalysis under optimized conditions, without significant loss of enzyme activity or L-DOPA yield after 8 cycles of repeated use. Our study provides an excellent and robust nano-biocatalyst for the cost-effective production of L-DOPA.

[Risk factors of hepatorenal syndrome in patients with acute on chronic liver failure].

OBJECTIVE: To identify the risk factors of hepatorenal syndrome in patients with hepatitis B virus (HBV)-related acute-on-chronic liver failure(ACLF). METHODS: A total of 726 hospitalized patients with HBV-ACLF were retrospectively analyzed. Data of demographic and clinical parameters (sex, age, family history, and presence of liver cirrhosis and diabetes), common complications (spontaneous bacterial peritonitis, pulmonary infection, hepatic encephalopathy, and upper gastrointestinal hemorrhage), and baseline biochemical parameters (albumin, globulin, total bilirubin, direct bilirubin, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, cholesterol, cholinesterase, K+, Na+, plasma thromboplastin antecedent, alpha-fetoprotein, HBV DNA, white blood cell count, hemoglobin, and platelet count) were collected from the medical records database. Univariate and multiple regression analyses were performed to determine the risk factors of hepatorenal syndrome. RESULTS: Multiple logistic regression analysis indicated that upper gastrointestinal hemorrhage [risk (R) = 1.313, relative hazard (RH) = 3.716, 95% confidence interval (CI): 2.156-6.404], hepatic encephalopathy (R = 1.120, RH = 3.065, 95% CI: 1.900-4.945), spontaneous bacterial peritonitis (R = 1.005, RH = 2.733, 95% CI: 1.379-5.417), pulmonary infection (R = 1.051, RH = 2.862, 95% CI: 1.783-4.592), and white blood cell count (R = 0.056, RH = 1.058, 95% CI: 1.010-1.107) were independent risk factors for hepatorenal syndrome development in patients with HBV-ACLF. CONCLUSION: Several risk factors were significantly associated with the development of hepatorenal syndrome in HBV-ACLF, including upper gastrointestinal hemorrhage, hepatic encephalopathy, spontaneous bacterial peritonitis, pulmonary infection, and elevated white blood cell count.

[Clinical characteristics of and factors related to relapse in chronic hepatitis B patients after nucleoside analogues withdrawal].

OBJECTIVE: To investigate the clinical characteristics of and factors related to relapse in chronic hepatitis B (CHB) patients who had previously achieved cessation criteria and had been withdrawn from nucleoside analogues treatment. METHODS: Sixty CHB patients who experienced relapse after nucleoside analogues withdrawal based on cessation criteria were enrolled in the study retrospectively. Each patient's data on biochemical, serological and viral characteristics corresponding to baseline (treatment initiation), withdrawal and relapse were collected. COX proportional hazard modeling was used to evaluate the factors related to relapse. RESULTS: The hepatitis B e antigen (HBeAg)-positive and -negative patients had similar median antiviral treatment times (38 months (range: 24 - 80) vs. 35 months (30 - 60); Z = -1.313, P more than 0.05). For all patients, the median follow-up time was 12 months (2 - 72), during which 49 (81.7%) patients developed virological breakthrough and 17 (28.3%) developed HBeAg recurrence. The patients who experienced virological breakthrough or HBeAg recurrence had significantly higher baseline levels of HBV DNA than those patients who remained disease-free (t = 2.15 and -2.54 respectively; P less than 0.05). The median relapse time of the HBeAg-positive patients was significantly longer than that of the HBeAg-negative patients (14 months (3 - 72) vs. 6 months (3 - 36); Chi-square test = 7.045, P less than 0.01). HBeAg status at baseline was identified as an independent factor associated with relapse (relative risk = 1.937, 95% confidence interval = 1.14-3.28, P less than 0.05). CONCLUSION: HBeAg-positive and-negative patients showed distinct clinical characteristics of relapse, with the latter being more prone to relapse soon after nucleoside analogues withdrawal. Prolonging the treatment course may be beneficial to HBeAg-negative patients, even if cessation criteria are achieved.

A PETAR method for risk assessment of human health and environment on the regional scale.

Risk assessments are necessary to effectively reveal the state of the degradation of living environments on a regional scale. However, risk assessments are often limited by time, cost, and technology, which make conducting effective evaluations difficult. Thus, in this study, the procedure for ecological tiered assessment of risk (PETAR) method was used to analyze the human health and environmental risks in Daye, China. This method first used the United States Environmental Protection Agency's risk assessment approach to qualitatively determine the risk sources, pressures, receptors, and effect endpoints and constructed a conceptual model of threats to the human living environment. Each risk-prone subregion was then evaluated using the fuzzy logic method. Next, a quantitative assessment was conducted for the subregions with the most serious environmental degradation. Finally, quantitative analyses were performed to verify the original hypotheses. The results showed that the high-risk areas were distributed in the industrial regions of Daye, wherein mining and processing clusters and mining settlements are widespread and confirmed the locations of the particular subregions with the most serious human health and environmental risks. This study also validated the practicality of the PETAR method for human health risk assessments in mining areas with large-scale, multifactor, and multihazard paths. Integr Environ Assess Manag 2023;19:239-253. © 2022 SETAC.

Rapid Screening and Characterization of Triterpene Saponins from the Root of Phytolacca acinosa Roxb by High-Performance Liquid Chromatography Coupled to Electrospray Ionization and Quadrupole Time-of-Flight Mass Spectrometry.

Triterpene saponins (TSs) are important bioactive constituents with structural diversity widely distributed in many plants. The root of Phytolacca acinosa Roxb (RPa) has been used as a traditional Chinese medicine. However, TSs as the main active ingredients in RPa have not been fully characterized. Here, we profiled TSs from RPa by high-performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS/MS). We tentatively identified 29 TSs, including 13 that had not been reported previously from this plant. This study indicates that HPLC-ESI-QTOF-MS/MS is an effective and rapid method for the characterization of complicated TSs in herbal extracts.

[The impact of early rapid virological response on the outcomes of hepatitis B associated acute on chronic liver failure during antiviral treatment].

OBJECTIVE: To investigate the impact of early rapid virological response on the outcomes of hepatitis B associated acute on chronic liver failure during antiviral treatment. METHODS: 106 acute on chronic liver failure patients in our hospital from January 2008 to July 2010 were enrolled in present study retrospectively. Besides internal medicine therapy, all patients received lamivudine (100 mg/d) or entecavir (0.5 mg/d) treatment. The profile of liver biochemistry, prothrombin time activity and viral load were detected at baseline and week 4, respectively. The patients were divided into HBV DNA negative group and HBV DNA positive group according to the viral load at week 4. The clinical features and treatment outcomes were compared between groups. Frequency variables were compared by x2 test or Fisher's exact test. Continuous variables were compared using independent samples T-test. The factors that impact on the treatment outcomes were determined using stepwise multivariate logistic regression analysis. RESULTS: At the week 4, the TBil and PTA in HBV DNA positive group [(261.6+/-205.6)mumol/L and 44.7%+/-19.7%, respectively] were significantly different from those in HBV DNA negative group [(160.1+/-173.4) mumol/L and 56.8%+/-23.1%, respectively] ( t = 2.190 and -2.077, respectively, P less than 0.05). The non-effective rate of HBVDNA positive group (50%, 9/18) was significantly higher than that of HBV DNA negative group (14.8%, 13/88) (x2 = 9.235, P less than 0.01). By using stepwise multivariate logistic regression analysis, the disease stage and HBV DNA undetectable at week 4 were the independent factor. The OR values of disease stage and HBV DNA undetectable were 6.559 and 0.209, respectively, and 95% CI was 2.316~18.576 and 0.058~0.747, respectively. CONCLUSION: The rapid suppression of viral load by nucleotide analogue may improve the efficacy of hepatitis B associated acute on chronic liver failure treatment. The early rapid virological response within first 4 weeks may contribute to the prediction of the treatment outcomes.