Bone turnover markers reference database in five Southeast Asian countries.

Osteoporosis is highly prevalent, particularly in developing countries. However, bone turnover marker reference ranges for management of osteoporosis in Asian population are yet to be explored and established. Thus, this study aims to develop a regional bone turnover markers (BTMs) reference database by combining country-specific reference database from five ASEAN countries: Malaysia, the Philippines, Singapore, Thailand, and Vietnam. We established a healthy reference population of 746 healthy premenopausal women aged 20 to 44 years old. Serum Procollagen 1 N-Terminal Propeptide (P1NP), Osteocalcin (OC), and Beta-Crosslaps (CTX) concentrations were measured using an automated immunoassay analyzer system, the cobas® modular analyzer systems (Roche Diagnostic Gmbh). The reference interval was defined as the central 95 % range. The estimated reference interval for CTX was 128 to 811 ng/L, OC was 9.0 to 33.0 µg/L, and for P1NP, the range was 22.8 to 96.5 µg/L. Comparison across countries showed that Singaporeans had the highest levels of median CTX along with Thais and Filipinos, who had significantly higher levels of P1NP and OC. Exploratory analysis on the associations with age showed that BTMs decreased with increasing age at 20 to 29 years old and plateaued after 30 years old. When excluding participants in their 20s, the reference interval estimated were CTX: 117-678 ng/L, P1NP: 21.6-85.8 µg/L and OC: 3.5-27.0 µg/L respectively. To the best of our knowledge, this is the first study to report BTMs reference intervals based on a healthy premenopausal Southeast Asian population which will contribute to the appropriate assessment and monitoring of bone turnover rate in the evaluation and management of osteoporosis in the Southeast Asian region. LAY SUMMARY: Osteoporosis is a common health issue, especially in developing countries. However, there is a lack of information on bone health markers specific to the Southeast Asian population. This study aimed to fill this gap by creating a reference database for bone turnover markers (BTMs) in Southeast Asian countries, including Malaysia, the Philippines, Singapore, Thailand, and Vietnam. The researchers studied 746 healthy women aged 20 to 44 years and measured blood markers related to bone health. The reference interval, representing the normal range, was determined. For example, the normal range for CTX was found to be 128 to 811 ng/L, for Osteocalcin was 9.0 to 33.0 µg/L, and for P1NP, the range was 22.8 to 96.5 µg/L. When excluding participants in their 20s, the reference intervals estimated were CTX: 117-678 ng/L, P1NP: 21.6-85.8 µg/L and OC: 3.5-27.0 µg/L respectively. Comparing the results across countries, Singaporeans, Thais, and Filipinos showed variations in their biochemical bone marker levels. Additionally, the study observed changes in the levels with age, with a decrease in BTMs observed after the age of 30. This groundbreaking study provides the first-ever reference intervals for BTMs in a healthy premenopausal Southeast Asian population. These findings will help in the proper assessment and monitoring of bone health, contributing to the management of osteoporosis in the Southeast Asian region.

Survey of Using Oral Anticoagulants in Patient with Non-valvular Atrial Fibrillation.

Background: In Vietnam, there has been no survey conducted on the prescribing and monitoring practices of oral anticoagulants to ensure that patients with atrial fibrillation receive appropriate. Objective: Therefore, we conducted this research to clarify the aforementioned issue in our hospital. Methods: We carried out a cross-sectional study by reviewing outpatient electronic medical records at the University Medical Center in Ho Chi Minh City. Our study included 1087 patients aged 18 years or older diagnosed with non-valvular atrial fibrillation (NVAF), who visited the Cardiology clinic between January 1st, 2021, and June 30th, 2021. Results: Among the 1087 patients with non-valvular atrial fibrillation (NVAF), 1036 were eligible for anticoagulant therapy. However, only 847 (81.8%) received prescriptions for either Vitamin K antagonists (VKAs) (n=129, 15.2%) or Non-Vitamin K antagonist oral anticoagulants (NOACs) (n=718, 84.8%). NOAC prescriptions were more commonly found in patients aged 75 or older (adjusted odds ratio [OR]=2), those with health insurance coverage (adjusted OR=2.9), and in individuals with a history of hypertension (adjusted OR=2). On the contrary, patients with a farming occupation were less likely to be prescribed NOACs (adjusted OR=0.4). About 75% of patients adhered to the guidelines recommending close monitoring during anticoagulant treatment. Notably, inappropriate prescriptions were identified in 27.7% of cases, especially among those with no recorded body weight (which is necessary for precise dosing based on creatinine clearance), those without health insurance, those with undocumented CHA2DS2-VASc scores, or those who were concurrently using antiplatelet agents. Conclusion: A discrepancy persists between clinical guidelines and the actual practice in diagnosing and managing patients with non-valvular atrial fibrillation (NVAF). It is crucial to prioritize the regular reevaluation of thromboembolic risk scores at follow-up appointments, ensure strict adherence to clinical monitoring standards, and align anticoagulant medication prescriptions with established guidelines.

Microplastics contamination in water supply system and treatment processes.

Due to global demand, millions of tons of plastics have been widely consumed, resulting in the widespread entry of vast amounts of microplastic particles into the environment. The presence of microplastics (MPs) in water supplies, including bottled water, has undergone systematic review, assessing the potential impacts of MPs on humans through exposure assessment. The main challenges associated with current technologies lie in their ability to effectively treat and completely remove MPs from drinking and supply water. While the risks posed by MPs upon entering the human body have not yet been fully revealed, there is a predicted certainty of negative impacts. This review encompasses a range of current technologies, spanning from basic to advanced treatments and varying in scale. However, given the frequent detection of MPs in drinking and bottled water, it becomes imperative to implement comprehensive management strategies to address this issue effectively. Consequently, integrating current technologies with management options such as life-cycle assessment, circular economy principles, and machine learning is crucial to eliminating this pervasive problem.

Production of an O-desmethylated product, a major human metabolite, of rabeprazole sulfide by bacterial P450 enzymes.

Rabeprazole is a common type of proton pump inhibitor (PPI) used to treat various peptic disorders. Unlike most PPI drugs, rabeprazole is spontaneously reduced to rabeprazole sulfide (thioether) when it is given to patients. As a result, rabeprazole sulfide is considered one of the active metabolites of rabeprazole. Rabeprazole sulfide is mainly metabolized to desmethyl rabeprazole sulfide by CYP2C19 and CYP2D6 in people. However, the pharmacological efficacy and safety of desmethyl rabeprazole sulfide have not yet been investigated. Its usage is challenging due to the high cost associated with the drug. In this study, we found CYP102A1 mutants that can produce desmethyl rabeprazole sulfide as a major metabolite of rabeprazole sulfide. The chemical characteristics of the major product were confirmed using high-performance liquid chromatography, LC-mass spectrometry, and nuclear magnetic resonance spectroscopy. CYP102A1 mutants R47L/F87V/L188Q, R47L/F87V/L188Q/A335V/Q359R, and R47L/F87V/L188Q/I254V/D351E showed kcat values of 39, 93, and 88 min-1, respectively, for O-desmethylation of rabeprazole sulfide. Furthermore, the highest concentration of desmethyl rabeprazole sulfide product from 2 mM rabeprazole sulfide at optimal conditions was obtained in bacterial whole-cell biotransformation with the R47L/F87V/L188Q mutant, reaching 0.63 mM at 4-h incubation. In conclusion, we present a platform that facilitates the efficient and sustainable production of the desmethylated product from rabeprazole sulfide for use in the biopharmaceutical industry.

Cyclin B3 is a dominant fast-acting cyclin that drives rapid early embryonic mitoses.

In many species, early embryonic mitoses proceed at a very rapid pace, but how this pace is achieved is not understood. Here we show that in the early C. elegans embryo, cyclin B3 is the dominant driver of rapid embryonic mitoses. Metazoans typically have three cyclin B isoforms that associate with and activate Cdk1 kinase to orchestrate mitotic events: the related cyclins B1 and B2 and the more divergent cyclin B3. We show that whereas embryos expressing cyclins B1 and B2 support slow mitosis (NEBD to Anaphase ~ 600s), the presence of cyclin B3 dominantly drives the ~3-fold faster mitosis observed in wildtype embryos. CYB-1/2-driven mitosis is longer than CYB-3-driven mitosis primarily because the progression of mitotic events itself is slower, rather than delayed anaphase onset due to activation of the spindle checkpoint or inhibitory phosphorylation of the anaphase activator CDC-20. Addition of cyclin B1 to cyclin B3-only mitosis introduces an ~60s delay between the completion of chromosome alignment and anaphase onset, which likely ensures segregation fidelity; this delay is mediated by inhibitory phosphorylation on CDC-20. Thus, the dominance of cyclin B3 in driving mitotic events, coupled to introduction of a short cyclin B1-dependent delay in anaphase onset, sets the rapid pace and ensures fidelity of mitoses in the early C. elegans embryo.

Malnutrition, Frailty, and Health-Related Quality of Life Among Rural Older Adults in Vietnam: A Cross-Sectional Study.

Purpose: Rural older adults are more likely to be malnourished than urban older adults, particularly those living in lower-middle-income countries like Vietnam. Therefore, this study aimed to address the prevalence of malnutrition and its association with frailty and health-related quality of life in older rural Vietnamese adults. Participants and Methods: This cross-sectional study was conducted on community-dwelling older adults (aged ≥ 60 years) living in a rural province in Vietnam. Nutritional status was determined using the Mini Nutritional Assessment Short Form (MNA-SF), and frailty was evaluated using the FRAIL scale. The 36-Item Short Form Survey (SF-36) was used to evaluate health-related quality of life. Results: Among the 627 participants, 46 (7.3%) were malnourished (MNA-SF score <8), and 315 (50.2%) were at risk of malnutrition (MNA-SF score: 8-11). Individuals with malnutrition had significantly higher rates of impairments in instrumental activities of daily living and activities of daily living than those without malnutrition (47.8% vs 27.4% and 26.1% vs 8.7%, respectively). The prevalence of frailty was 13.5%. Risk of malnutrition and malnutrition were associated with high risks of frailty, with odds ratios of 2.14 (95% confidence interval [CI]: 1.16-3.93) and 4.78 (1.86-12.32), respectively. Furthermore, the MNA-SF score was positively correlated with eight domains of the health-related quality of life among rural older adults. Conclusion: The prevalence rates of malnutrition, risk of malnutrition, and frailty were high among older adults in Vietnam. A strong association was observed between nutritional status and frailty. Therefore, this study reinforces the importance of screening for malnutrition and risk of malnutrition among older rural individuals. Further studies should explore whether early nutritional intervention reduces the risk of frailty among older adults and increase their health-related quality of life in the Vietnamese population.

3-OH Phloretin Inhibits High-Fat Diet-Induced Obesity and Obesity-Induced Inflammation by Reducing Macrophage Infiltration into White Adipose Tissue.

Phloretin and its glycoside phlorizin have been reported to prevent obesity induced by high-fat diet (HFD), but the effect of 3-OH phloretin, a catechol metabolite of phloretin, has not been investigated. In this study, we investigated the anti-obesity effects of phloretin and 3-OH phloretin in HFD-fed mice. The body weight gain induced by HFD was more inhibited by administration of 3-OH phloretin than by phloretin. The increases in fat mass, white adipose tissue (WAT) weight, adipocyte size, and lipid accumulation by HFD were also remarkably inhibited by 3-OH phloretin and, to a lesser extent, by phloretin. The HFD-induced upregulation of chemokines and pro-inflammatory cytokines was suppressed by 3-OH phloretin, preventing M1 macrophages from infiltrating into WAT and thereby reducing WAT inflammation. 3-OH phloretin also showed a more potent effect than phloretin on suppressing the expression of adipogenesis regulator genes, such as PPARγ2, C/EBPα, FAS, and CD36. Fasting blood glucose and insulin levels increased by HFD were diminished by the administration of 3-OH phloretin, suggesting that 3-OH phloretin may alleviate obesity-induced insulin resistance. These findings suggested that 3-OH phloretin has the potential to be a natural bioactive compound that can be used in the prevention or treatment of obesity and insulin resistance.

Combined Gap-Polymerase Chain Reaction and Targeted Next-Generation Sequencing Improve α- and ß-Thalassemia Carrier Screening in Pregnant Women in Vietnam.

Vietnam has a high thalassemia burden. We collected blood samples from 5880 pregnant Vietnamese women during prenatal health checks to assess thalassemia carrier frequency using combined gap-polymerase chain reaction (gap-PCR) and targeted next-generation sequencing (NGS). Thalassemia carriers were identified with prevalence of 13.13% (772), including 7.82% (460) carriers of α-thalassemia (α-thal), 5.31% (312) carriers of ß-thalassemia (ß-thal), and 0.63% (37) concurrent α-/ß-thal carriers. Deletional mutations (368) accounted for 80.0% of α-thal carriers, of which, --SEA (Southeast Asian) (n = 254; 55.0%) was most prevalent, followed by the -α3.7 (rightward) (n = 66; 14.0%) and -α4.2 (leftward) (n = 45; 9.8%) deletions. Hb Westmead (HBA2: c.369C>G) (n = 53) and Hb Constant Spring (Hb CS or HBA2: c.427T>C) (in 28) are the two most common nondeletional α-globin variants, accounting for 11.5 and 6.0% of α-thal carriers. We detected 11 different ß-thal genotypes. Hb E (HBB: c.79G>A) (in 211) accounted for 67.6% of ß-thal carriers. The most common ß-thal genotypes were associated with mutations at codon 17 (A>T) (HBB: c.52A>T), codons 41/42 (-TTCT) (HBB: c.126_129delCTTT), and codon 71/72 (+A) (HBB: c.217_218insA) (prevalence 0.70%, 0.68%, and 0.2%, respectively). Based on mutation frequencies calculated in this study, estimates of 5021 babies in Vietnam are affected with clinically severe thalassemia annually. Our data suggest a higher thalassemia carrier frequency in Vietnam than previously reported. We established that combining NGS with gap-PCR creates an effective large-scale thalassemia screening method that can detect a broad range of mutations.

Detection of maternal carriers of common α-thalassemia deletions from cell-free DNA.

α-Thalassemia is a common inherited blood disorder manifested mainly by the deletions of α-globin genes. In geographical areas with high carrier frequencies, screening of α-thalassemia carrier state is therefore of vital importance. This study presents a novel method for identifying female carriers of common α-thalassemia deletions using samples routinely taken for non-invasive prenatal tests for screening of fetal chromosomal aneuploidies. A total of 68,885 Vietnamese pregnant women were recruited and α-thalassemia statuses were determined by gap-PCR, revealing 5344 women (7.76%) carried deletions including αα/--SEA (4.066%), αα/-α3.7 (2.934%), αα/-α4.2 (0.656%), and rare genotypes (0.102%). A two-stage model was built to predict these α-thalassemia deletions from targeted sequencing of the HBA gene cluster on maternal cfDNA. Our method achieved F1-scores of 97.14-99.55% for detecting the three common genotypes and 94.74% for detecting rare genotypes (-α3.7/-α4.2, αα/--THAI, -α3.7/--SEA, -α4.2/--SEA). Additionally, the positive predictive values were 100.00% for αα/αα, 99.29% for αα/--SEA, 94.87% for αα/-α3.7, and 96.51% for αα/-α4.2; and the negative predictive values were 97.63%, 99.99%, 99.99%, and 100.00%, respectively. As NIPT is increasingly adopted for pregnant women, utilizing cfDNA from NIPT to detect maternal carriers of common α-thalassemia deletions will be cost-effective and expand the benefits of NIPT.

Effects of sperm preparation techniques on sperm survivability and DNA fragmentation.

OBJECTIVE: This study was performed to determine the effect of swim-up (SU) and density gradient centrifugation (DGC) on sperm survival and DNA fragmentation. METHODS: Individual semen samples were analyzed before each was divided into two aliquots (half for SU and half for DGC) for calculation of sperm survival and the DNA fragmentation index (DFI). Sperm DNA fragmentation was determined using the sperm chromatin dispersion test. RESULTS: The DFI of the 63 semen samples processed using both procedures was lower than that of the fresh semen samples. The DFI was significantly lower for samples processed using the SU than DGC method. In the sperm survival test, the SU technique was associated with increased sperm motility and vitality following preparation. After 24 hours, however, the concentration and percentage of surviving sperm were significantly lower in the SU than DGC group. CONCLUSIONS: Both semen preparation techniques help to minimize sperm DNA fragmentation; however, when the DFI is <30%, the SU technique is more appropriate than DGC. While DGC may be superior for intrauterine insemination, the SU method may be preferable for in vitro fertilization or maturation.

Phthalates in the environment: characteristics, fate and transport, and advanced wastewater treatment technologies.

Phthalates are well-known emerging contaminants that harm human health and the environment. Therefore, this review aims to discuss about the occurrence, fate, and phthalates concentration in the various environmental matrices (e.g., aquatic, sediment, soil, and sewage sludge). Hence, it is necessary to treat sources containing phthalates before discharging them to aqueous environment. Various advanced wastewater treatments including adsorption process (e.g., biochar, activated carbon), advanced oxidation processes (e.g., photo-fenton, ozonation, photocatalysis), and biological treatment (membrane bioreactor) have been successfully to address this issue with high removal efficiencies (70-95%). Also, the degradation mechanism was discussed to provide a comprehensive understanding of the phthalate removal for the reader. Additionally, key factors that influenced the phthalates removal efficiency of these technologies were identified and summarized with a view towards pilot-scale and industrial applications.

A Novel Statin Compound from Monacolin J Produced Using CYP102A1-Catalyzed Regioselective C-Hydroxylation.

Statins inhibit the 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMG-CoA reductase), which is the rate-limiting enzyme in cholesterol biosynthesis. Statin therapy reduces morbidity and mortality in those who are at high risk of cardiovascular disease. Monacolin J is a statin compound, which is an intermediate in the lovastatin biosynthesis pathway, in the fungus Aspergillus terreus. It is also found in red yeast rice, which is made by culturing rice with the yeast Monascus purpureus. Monacolin J has a hydroxyl substituent at position C'-8 of monacolin L. Here, a new statin derivative from monacolin J was made through the catalysis of CYP102A1 from Bacillus megaterium. A set of CYP102A1 mutants of monacolin J hydroxylation with high catalytic activity was screened. The major hydroxylated product was C-6'a-hydroxymethyl monacolin J, whose structure was confirmed using LC-MS and NMR analysis. The C-6'a-hydroxymethyl monacolin J has never been reported before. It showed a greater ability to inhibit HMG-CoA reductase than the monacolin J substrate itself. Human liver microsomes and human CYP3A4 also showed the ability to catalyze monacolin J in producing the same product of the CYP102A1-catalyzed reaction. This result motivates a new strategy for the development of a lead for the enzymatic and chemical processes to develop statin drug candidates.

Chemoprevention with a tea from hawthorn (Crataegus oxyacantha) leaves and flowers attenuates colitis in rats by reducing inflammation and oxidative stress.

The purpose of the study was to determine the effects of a tea from the leaves and flowers of Crataegus oxyacantha in rats with colitis. Colitis was induced by administration of 2,4,6-trinitrobenzene sulfonic acid. Hawthorn tea (HT) (100 mg/kg) was given via gavage for 21 days and the mesalamine drug (100 mg/kg) was administrated during the period of disease onset. HT was rich in total phenolic compounds (16.5%), flavonoids (1.8%), and proanthocyanidins (1.5%); vitexin-2-O-rhamnoside was the main compound detected. Mesalamine and the HT diminished the length of the lesions formed in the colon, in addition to reducing the levels of myeloperoxidase and interleukin-1ß. Mesalamine was able to significantly reverse the body weight loss, while HT improved the activity of glutathione reductase and catalase. Histological scoring was not changed by the interventions, but it was highly correlated with the necrotic area. HT given at 100 mg/kg can be effective against colitis.

Enzymatic Production of 3-OH Phlorizin, a Possible Bioactive Polyphenol from Apples, by Bacillus megaterium CYP102A1 via Regioselective Hydroxylation.

Phlorizin is the most abundant glucoside of phloretin from the apple tree and its products. Phlorizin and its aglycone phloretin are currently considered health-beneficial polyphenols from apples useful in treating hyperglycemia and obesity. Recently, we showed that phloretin could be regioselectively hydroxylated to make 3-OH phloretin by Bacillus megaterium CYP102A1 and human P450 enzymes. The 3-OH phloretin has a potent inhibitory effect on differentiating 3T3-L1 preadipocytes into adipocytes and lipid accumulation. The glucoside of 3-OH phloretin would be a promising agent with increased bioavailability and water solubility compared with its aglycone. However, procedures to make 3-OH phlorizin, a glucoside of 3-OH phloretin, using chemical methods, are not currently available. Here, a biocatalytic strategy for the efficient synthesis of a possibly valuable hydroxylated product, 3-OH phlorizin, was developed via CYP102A1-catalyzed regioselective hydroxylation. The production of 3-OH phlorizin by CYP102A1 was confirmed by HPLC and LC-MS spectroscopy in addition to enzymatic removal of its glucose moiety for comparison to 3-OH phloretin. Taken together, in this study, we found a panel of mutants from B. megaterium CYP102A1 could catalyze regioselective hydroxylation of phlorizin to produce 3-OH phlorizin, a catechol product.

Liquid biopsy uncovers distinct patterns of DNA methylation and copy number changes in NSCLC patients with different EGFR-TKI resistant mutations.

Targeted therapy with tyrosine kinase inhibitors (TKI) provides survival benefits to a majority of patients with non-small cell lung cancer (NSCLC). However, resistance to TKI almost always develops after treatment. Although genetic and epigenetic alterations have each been shown to drive resistance to TKI in cell line models, clinical evidence for their contribution in the acquisition of resistance remains limited. Here, we employed liquid biopsy for simultaneous analysis of genetic and epigenetic changes in 122 Vietnamese NSCLC patients undergoing TKI therapy and displaying acquired resistance. We detected multiple profiles of resistance mutations in 51 patients (41.8%). Of those, genetic alterations in EGFR, particularly EGFR amplification (n = 6), showed pronounced genome instability and genome-wide hypomethylation. Interestingly, the level of hypomethylation was associated with the duration of response to TKI treatment. We also detected hypermethylation in regulatory regions of Homeobox genes which are known to be involved in tumor differentiation. In contrast, such changes were not observed in cases with MET (n = 4) and HER2 (n = 4) amplification. Thus, our study showed that liquid biopsy could provide important insights into the heterogeneity of TKI resistance mechanisms in NSCLC patients, providing essential information for prediction of resistance and selection of subsequent treatment.

Assessment of myocardial performance index in late-onset fetal growth restriction.

The aim of this study is to determine whether the myocardial performance index (MPI) is increased in fetal growth restriction (FGR) fetuses and if increased MPI is related to adverse outcomes of FGR. This is a prospective cross-sectional study. Seventy-three late-onset FGR fetuses and 97 gestational-age matched control fetuses were enrolled in this study. Fetal blood flow parameters including MPI values were measured and compared between the two groups. For the effect of severity of growth restriction on MPI value, they were also compared with < 3rd and 3rd - 10th centile groups. FGR fetuses were divided into two groups by favorable and adverse outcome and ultrasound parameters were compared between these two groups. Moreover, significant factors related to adverse outcomes by univariate analysis were analyzed by multivariate logistic regression analysis. Pulsatility index of umbilical arterial flow (UA-PI), MPI and amniotic fluid index in the FGR were significantly different from the control fetuses. However, no significant difference between < 3rd and 3rd - 10th centile groups was detected in MPI and UA-PI. The increased levels of MPI and UA-PI were independently related with adverse outcome of late-onset FGR pregnancy. In conclusion, MPI values were increased in late-onset FGR pregnancy, and the higher level of MPI could predict adverse outcome as well as the measurement of UA-PI. Clinicians should consider cardiac dysfunction in FGR through increased MPI.

Effects of Feed Mixed with Lactic Acid Bacteria and Carbon, Nitrogen, Phosphorus Supplied to the Water on the Growth and Survival Rate of White Leg Shrimp (Penaeus vannamei) Infected with Acute Hepatopancreatic Necrosis Disease Caused by Vibrio parahaemolyticus.

This study aimed to evaluate the growth, survival rate, and resistance to acute hepatopancreatic necrosis disease (AHPND) of white leg shrimp (Penaeus vannamei) by using Lactobacillus plantarum, Lactobacillus fermentum, and Pediococcus pentosaceus mixed with feed, and at the same time supplying CNP in a ratio of 15:1:0.1 to the water. As a result, the treatments that shrimp were fed with feed containing lactic acid bacteria (LAB), especially L. plantarum, have increased shrimp growth, total hemocyte cells, granulocyte cells, and hyaline cells significantly (p < 0.05) in comparison to the control group. The supply of CNP to the water has promoted the intensity of V. parahaemolyticus effects on shrimp health and significantly decreased total hemocyte cells, granulocyte cells, and hyaline cells by 30-50% in the period after three days of the challenge, except in L. plantarum treatment, which had only a 20% decrease compared to other treatments. In CNP supplying treatments, the AHPND infected rate and mortality of shrimp were higher than those in other treatments. In summary, the supply of CNP had significantly reduced the shrimp's immune response and promoted the susceptibility of shrimp to AHPND in both cases of use with and without LAB-containing diets.

Screening for pancreatic lipase natural modulators by capillary electrophoresis hyphenated to spectrophotometric and conductometric dual detection.

The search for novel pancreatic lipase (PL) inhibitors has gained increasing attention in recent years. For the first time, a dual detection capillary electrophoresis (CE)-based homogeneous lipase assay was developed employing both the offline and online reaction modes. The hydrolysis of 4-nitrophenyl butyrate (4-NPB) catalyzed by PL into 4-nitrophenol and butyrate was monitored by spectrophotometric and conductimetric detection, respectively. The assays presented several advantages such as economy in consumption (few tens of nanoliters for online assays to few tens of microliters for offline assays), no modification of lipase, rapidity (<10 min) and versatility. Tris/MOPS (10 mM, pH 6.6) was used as the background electrolyte and the incubation buffer for enzymatic reactions. We confirmed that in the conditions of the study (small substrate 4-NPB, 37 °C, pH 6.6), the PL was active even in the absence of dipalmitoylphosphatidylcholine (DPPC) vesicles, generally used to mimic the lipid-water interface. This was confirmed by the maximum velocity (Vmax) and the Michaelis-Menten constant (Km) values that were the same order of magnitude in the absence and presence of DPPC. The developed method was used to screen crude aqueous plant extracts and purified compounds. We were able to identify the promising PL inhibition of hawthorn leaf herbal infusions at 1 mg mL-1 (37%) and PL activation by fresh and dry hawthorn flowers (∼24%). Additionally, two triterpenoids purified from extracts of oakwood were identified for the first time as potent PL inhibitors demonstrating 51 and 58% inhibition at 1 mg mL-1, respectively.