Screening and identification of bioactive compounds from citrus against non-structural protein 3 protease of hepatitis C virus genotype 3a by fluorescence resonance energy transfer assay and mass spectrometry

BACKGROUNDHepatitis C virus genotype 3a (HCV G3a) is highly prevalent in Pakistan. Due to the elevated cost of available Food and Drug Administration-approved drugs against HCV, medicinal natural products of potent antiviral activity should be screened for the cost-effective treatment of the disease. Furthermore, from natural products, active compounds against vital HCV proteins like non-structural protein 3 (NS3) protease could be identified to prevent viral proliferation in the host.AIMTo develop cost-effective HCV genotype 3a NS3 protease inhibitors from citrus fruit extracts.METHODSFull-length NS3 without co-factor non-structural protein 4A (NS4A) and codon optimized NS3 protease in fusion with NS4A were expressed in Escherichia coli. The expressed protein was purified by metal ion affinity chromatography and gel filtration. Citrus fruit extracts were screened using fluorescence resonance energy transfer (FRET) assay against the protease and polyphenols were identified as potential inhibitors using electrospray ionization-mass spectrometry (MS)/MS technique. Among different polyphenols, highly potent compounds were screened using molecular modeling approaches and consequently the most active compound was further evaluated against HCV NS4A-NS3 protease domain using FRET assay.RESULTSNS4A fused with NS3 protease domain gene was overexpressed and the purified protein yield was high in comparison to the lower yield of the full-length NS3 protein. Furthermore, in enzyme kinetic studies, NS4A fused with NS3 protease proved to be functionally active compared to full-length NS3. So it was concluded that co-factor NS4A fusion is essential for the purification of functionally active protease. FRET assay was developed and validated by the half maximal inhibitory concentration (IC₅₀) values of commercially available inhibitors. Screening of citrus fruit extracts against the native purified fused NS4A-NS3 protease domain showed that the grapefruit mesocarp extract exhibits the highest percentage inhibition 91% of protease activity. Among the compounds identified by LCMS analysis, hesperidin showed strong binding affinity with the protease catalytic triad having S-score value of -10.98.CONCLUSIONFused NS4A-NS3 protease is functionally more active, which is effectively inhibited by hesperidin from the grapefruit mesocarp extract with an IC₅₀ value of 23.32 µmol/L.     

Effects of Hybridized Synthetic Fibers on the Shear Properties of Cement Composites

The use of fibers in cementitious composites yields numerous benefits due to their fiber-bridging capabilities in resisting cracks. Therefore, this study aimed to improve the shear-resisting capabilities of conventional concrete through the hybridization of multiple synthetic fibers, specifically on reinforced concrete structures in seismic-prone regions. For this study, 16 hybrid fiber-reinforced concretes (HyFRC) were developed from the different combinations of Ferro macro-synthetic fibers with the Ultra-Net, Super-Net, Econo-Net, and Nylo-Mono microfibers. These hybrids were tested under direct shear, resulting in improved shear strength of controlled specimens by Ferro-Ultra (32%), Ferro-Super (24%), Ferro-Econo (44%), and Ferro-Nylo (24%). Shear energy was further assessed to comprehend the effectiveness of the fiber interactions according to the mechanical properties, dosage, bonding power, manufactured material, and form of fibers. Conclusively, all fiber combinations used in this study produced positive synergistic effects under direct shear at large crack deformations.     

Behavioral and electrophysiological responses of Aedes albopictus to certain acids and alcohols present in human skin emanations

Human skin emanations attract hungry female mosquitoes toward their host for blood feeding. In this study, we report the flight orientation and electroantennogram response of Aedes albopictus females to certain unsaturated acids and alcohols found in human skin. In the Y-tube olfactometer, odors of lactic acid and 2-methyl-3-pentanol attracted 54–65 % of Ae. albopictus females at all doses in a dose-dependent manner. However, at the highest dose (10^?2 g), the acids repelled 40–45 % females. Attractancy (ca. 62–68 %) at lower doses and repellency (ca. 30–45 %) at higher doses were recorded for 3-methyl-3-pentanol and 1-octen-3-ol, while 5-hexen-1-ol, cis-2-hexen-1-ol, and trans 2-hexen-1-ol odor repelled ca. 55–65 % of Ae. albopictus females at all doses. Antenna of female Ae. albopictus exhibited a dose-dependent EAG response up to 10^?3 g of l-lactic acid, trans-2-methyl-2-pentenoic acid, 2-octenoic acid, trans-2-hexen-1-ol and 1-octen-3-ol stimulations; however, the highest dose (10^?2 g) caused a little decline in the EAG response. EAG response of 9–10-fold was elicited by lactic acid, 2-octenoic acid, trans-2-hexenoic acid, and 3-methyl-3-pentanol, while cis-2-hexen-1-ol and trans-2-methyl pentenoic acid elicited 1–5-fold responses compared to solvent control. A blend of attractive compounds could be utilized in odor-baited trap for surveillance and repellent molecules with suitable formulation could be used to reduce the biting menace of mosquitoes.     

Natural compounds with xanthine oxidase inhibitory activity: A review

Hyperuricemia (HUA), a disease due to an elevation of body uric acid level and responsible for various diseases such as gout, cardiovascular disorders, and renal failure, is a major ground debate for the medical science these days. Considering the risk factors linked with allopathic drugs for the treatment of this disease, the debate has now become a special issue. Previously, we critically discussed the role of dietary polyphenols in the treatment of HUA. Besides dietary food plants, many researchers figure out the tremendous effects of medicinal plants‐derived phytochemicals against HUA. Keeping in mind all these aspects, we reviewed all possible managerial studies related to HUA through medicinal plants (isolated compounds). In the current review article, we comprehensively discussed various bioactive compounds, chemical structures, and structure–activity relationship with responsible key enzyme xanthine oxidase.     

A Study of Patient Experience Using a Blood Glucose Meter With an In-Built Insulin Dose Calculator

Background:Accurate calculation and adjustment of insulin doses is integral to maintaining glycemic control in insulin treated patients. Difficulties with insulin dose calculations may lead to poor adherence to blood glucose monitoring and insulin treatment regimes, resulting in poor metabolic control. The main objective of this study was to evaluate ease of use and user preference of a high specification touch screen blood glucose meter, which has an in-built insulin calculator, compared to patients’ usual method of testing blood glucose and deciding insulin doses.Methods:Patients with diabetes on a multiple daily injection insulin regime used the Test Meter without the insulin calculator and 1 of 3 comparator meters, each for a 7-day period. They then used the Test Meter with the in-built calculator for 10 days. Patients completed an ease of use questionnaire after each 7-day period, a preference questionnaire after the second 7-day period, and a questionnaire comparing the Test Meter with their usual method after the final 10-day period.Results:Of 164 patients who completed the study, 76% stated a preference for the Test Meter as a diabetes management tool compared to their usual method. A small number of patients preferred familiar methods and/or calculating insulin doses themselves. The log book function of meters was important to most patients.Conclusions:The Test Meter system with in-built insulin calculator supports people to better manage their diabetes and increases their confidence. Patients have different needs and preferences which should be acknowledged and supported in a patient centered health service.     

Impact of a Spatial Repellent on Malaria Incidence in Two Villages in Sumba,Indonesia

A randomized, double-blinded, placebo-controlled study was conducted to examine the effect of spatial repellent (SR) in households at risk of malaria in Indonesia. Following presumptive radical cure for malaria in 180 adult men representing sentinels of new infection in four clusters within two villages, all households were given either metofluthrin or placebo mosquito coils. Weekly blood smear screening and human-landing mosquito catches were done throughout the 6 months intervention. Malaria infections occurred in 61 subjects living in placebo households and 31 subjects living in SR coil households, suggesting a 52% protective effect of SR. Likewise, anopheles indoor human landing rates were 32% lower in homes receiving SR coils. Differences in the malaria attack rate between SR- and placebo-treated homes was significant when not accounting for the effects of clustering. When the analysis was adjusted for intra-cluster correlation, the differences between SR- and placebo-treated homes were not statistically significant. The findings provide evidence of SR public health benefit and support a larger trial statistically powered to detect those effects.