Acid Assay Research Articles

Gaseous and aerosol emissions from open burning of tree pruning and hedge trimming residues: Detailed composition and toxicity

Gaseous and PM10 samples were collected during the open burning of pruning residues (olive branches and garden waste) and characterised by distinct analytical techniques to obtain comprehensive chemical emission profiles. Oxidative potential (dithiothreoitol and ascorbic acid assays) and cell viability tests were also performed with the aim of evaluating aerosol toxicity. Emission factors (EFs) were as follows (g kg−1 biofuel, dry basis): 1537–1672 for CO2, 41.9–80 for CO, 2.74–6.6 for CH4, 0.89–3.51 for ethane, 0.79–1.78 for ethylene and 0.56–3.47 for formaldehyde. Emissions of PM10, organic carbon (OC) and elemental carbon (EC) were in the ranges 8–41, 3–18, and 0.4–1.5 g kg−1 biofuel, dry basis, respectively. OC accounted for 35–45% of the total PM10 mass, while EC contributed between around 3% and 5%. WSOC/OC ratios varied from 0.4 to 0.6, revealing that a substantial portion of the carbon emitted was hydrosoluble. Water soluble ions constituted around 8–21% of the PM10 mass, with potassium and chloride as the most abundant ions in all samples. Levoglucosan, widely used a reliable biomass burning tracer, was found in significant amounts in all samples (up to 1.2% of the PM10 mass). Retene, generally pointed out as a biomass combustion biomarker, was the predominant PAH. WSOC and some PAHs showed significant positive correlations with the intrinsic OP measured with the DTT assay, while the OPAA was significantly correlated with some trace metals, such as Fe or Ni. All samples significantly reduced the viability of alveolar epithelial cells.

Evaluation of antioxidant status of lens epithelial cells in cataract patients.

The main factor that causes cataracts is the increased oxidative stress and imbalance of an antioxidant defense mechanism, which leads to significant changes in the lens microarchitecture. Senile cataract is the most common type of acquired cataracts due to aging. We carried out a case-control study in the biochemistry department to examine the antioxidant status (catalase and total antioxidant capacity [TAC]) and lipid peroxidation marker, that is, malondialdehyde (MDA) in human lens epithelial cells (HLECs) of different grades of senile cortical, nuclear, and posterior subcapsular cataracts. We collected 150 samples from patients aged 50-90 years. These included 50 samples of cortical cataracts, 50 of nuclear cataracts, and 50 samples of posterior subcapsular cataracts. We measured catalase activity by the Beer method, TAC by the Benzie and Strain method, and protein by the Bradford method. We also estimated TAC in the aqueous extract of HLECs by the ferric reducing ability of plasma (FRAP) method and MDA by the thiobarbituric acid assay method. The results of this study showed that the level of catalase enzyme was higher in the first grade of nuclear, posterior subcapsular, and cortical cataracts than in other grades. This suggests that the catalase enzyme activity drops sharply in the second and third grades of these types of cataracts. The same pattern was observed for TAC, which was higher in the first grade of nuclear, posterior subcapsular, and cortical cataracts than in other grades. There were significant differences between catalase and TAC in different grades of cataracts, indicating that as the grading increases, both catalase and TAC decrease. The results of this study showed that the levels of MDA were higher and the levels of catalase and TAC were lower in patients with more severe cataracts compared to the healthy controls.

Nucleic acid binding affinity and antioxidant activity of N-m-Tolyl-4-Chlorophenoxyacetohydroxamicacid.

Hydroxamic acids represent a group of weak organic acids, both naturally occurring and synthetically derived, characterized by the general formula RC(= O)N(R'OH). In this study, we investigated the binding behavior of N-m-tolyl-4-chlorophenoxyaceto hydroxamic acid with calf thymus DNA (ct-DNA) and torula yeast RNA (t-RNA) through a combination of techniques including UV-visible spectroscopy, fluorescence emission analysis, viscometry, and computational simulations using AutoDock4 software. Our findings reveal that the mode of binding between the compound and the nucleic acids is consistent with intercalation. Competitive binding experiments demonstrated that the complex competes effectively with ethidium bromide (EB) for binding to ct-DNA/t-RNA, displacing EB from its binding sites. Additionally, the introduction of the compound into the DNA-EB system resulted in a quenching of fluorescence emission peaks. Analysis of absorption spectra indicated a red shift and hypochromic shift when the compound interacted with DNA, further supporting the intercalative binding mode. The calculated binding constant (Kb) value for the compound is 6.62 × 104M-1 and 5.40 × 103M-1 indicating a strong interaction with ct-DNA and t-RNA respectively. We determined the Stern-Volmer constants for ct-DNA and t-RNA as 9.96 × 104M-1 and 8.13 × 105M-1, respectively. The binding free energy values for ct-DNA/t-RNA were calculated to be -3.741 × 107 and -5.425 × 108kcal/mol, respectively. Viscometric studies corroborated the UV results, showing a continuous increase in relative viscosity of ct-DNA/t-RNA solutions with the addition of the optimal hydroxamic acid concentration. Furthermore, we assessed the antioxidant activity of the compound using DPPH-radical scavenging and β-carotene linoleic acid assays. Gel electrophoresis results demonstrated the compound's remarkable efficacy in preventing DNA damage. Collectively, all experimental evidence supports the conclusion that N-m-tolyl-4-chlorophenoxyaceto hydroxamic acid binds to ct-DNA/t-RNA through an intercalative mechanism, which is consistent with our molecular docking simulations.

Physiochemical Analysis, Antioxidant Activity, Antimicrobial Activity and Phenolic Profile of Ipomoea eriocarpa R. Br. Leaves.

Many active substances found in plants are essential in the treatment of transmissible and non-transmissible ailments. To enhance their well-being and capabilities, most people around the globe use medicines from plants and herbal origin. The goal of the current study was an assessment of physiochemical properties as well as measuring the effect of various solvents (ethanol, chloroform, and water) on the phenolic profile and on free radical scavenging potential. The standard procedures were used for the preliminary phytochemical, physiochemical and fluorescence analysis. Folin-Ciocalteu and colorimetric methods were used to determine the quantitative phytochemical analysis, respectively. The free radical scavenging potential was determined through 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′′-casino-bis (3-ethylbenzothiazoline-6-sulphonic) acid (ABTS) assays. The antimicrobial activity was assessed using the disk diffusion method on a range of bacterial and fungal strains. The finding showed that ethanol was the best solvent for extracting polyphenols. The presence of alkaloids, carbohydrates, flavonoids, glycosides, phenolic compounds, and tannins was observed in the plant. Additionally, fluorescence characteristics and physiochemical parameters provided valuable data that can be used to establish standards for this plant. Results of quantitative phytochemical estimation revealed that the ethanol extract had a higher phenolic content (71.82 mg/g GAE) and flavonoid content (82.7 mg/g QE), while the aqueous extract had the lowest phenolic content (27.05 mg/g GAE) and flavonoid content (18.93 mg/g QE). The I. eriocarpa ethanolic extract had the maximum scavenging potential against DPPH (32.36 µg/mL) and ABTS (55.54 µg/mL). The results of the antimicrobial activity assessment also indicate that the ethanol extract exhibited a greater inhibition zone when tested against both bacterial strains. Results of thecurrent study suggested that the ethanolic extract has significant antioxidant potential and has the capability of being a notable source of organic antioxidants for the formulation of functional foods.

Analysis of neuraminidase activity of human parainfluenza viruses using enzyme-linked lectin assay and BTP3-Neu5Ac assay.

Human parainfluenza viruses (hPIVs) are causative agents of upper and lower respiratory tract infections and they have four serotypes. The virion surface displays hemagglutinin-neuraminidase (HN), having hemagglutinating (HA) and neuraminidase (NA) activities in a single molecule. The HA activity binds the virion to sialic acid on the viral receptor on host cells and the NA releases the progeny viruses from the cell surface. There are several methods for assaying viral NA activity, such as the thiobarbituric acid assay, 4-methylumbelliferyl-N-acetyl-α-d-neuraminic acid assay, NA-Star assay, and enzyme-linked lectin assay (ELLA). However, these are mainly used for influenza viruses and not for hPIVs. A fluorescent-based cytochemical NA assay using BTP3-Neu5Ac as the substrate was recently developed and used for orthomyxo- and paramyxoviruses, including types 1 and 3 hPIVs. In this study, we used the ELLA, and BTP-Neu5Ac assay for 14 field isolate strains of hPIVs including all four serotypes. The reaction in ELLA at pH 6.5 using peanut agglutinin (PNA) as a lectin was very low for all tested viruses except a type 3 virus strain with the maximum reaction at pH 6.5 and the acidic conditions did not enhance the reaction. ELLA with another lectin, Erythrina cristagalli agglutinin exhibited significant and stronger reactions than with PNA in some strains of types 1 and 3 viruses. The BTP3-Neu5Ac assay showed a fluorescent signal on cells infected with all the viruses except the hPIV1/Sendai/713/2018 strain in LLC-MK2 and/or MNT-1. The signal was detected in cell-free virus, as well, in all the viruses except the hPIV4a/Sendai/3935/2003 strain. The strength of the signal varied among viral strains but it was stronger in the reaction at pH 4.0 than pH 7.0 and strongest in type 2 hPIVs.

Floral Fusion: Unravelling the Potent Blend of Ixora coccinea and Rhododendron arboreum for Health and Safety Benefits.

Ixora coccinea and Rhododendron arboreum are known for their traditional medicinal uses due to their diverse phytochemicals and pharmacological effects, which have attracted the interest of many researchers. This study aims to evaluate the antioxidant, anti-inflammatory, and cytotoxic effects of their combined extracts. In vitro antioxidant activity against reactive oxygen species (ROS) was measured using the ferric-reducing ability of plasma (FRAP), nitric oxide (NO), and 2,2-azino-bis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS) assays, while anti-inflammatory effects were assessed via the membrane stabilization method. Docking studies were performed to evaluate the interaction of phytochemicals - anthocyanins, quercetin, and ursolic acid, which are present in these plants - with nuclear factor kappa B (NF-κB), cyclooxygenase-1 (COX-1), and cyclooxygenase-2 (COX-2). Standard protocols were used to evaluate embryotoxicity using the brine shrimp model and cytotoxicity using the zebrafish model, which is crucial for determining safe clinical dosages. The analysis revealed diverse bioactive compounds, including anthocyanins, quercetin, and ursolic acid. The formulation effectively inhibited ROS production at lower concentrations (inhibitory concentration 50%, or IC50 value ~2.8 µg/mL), indicating their potential for managing oxidative stress. Quercetin demonstrated the strongest interaction with all tested proteins, particularly NF-kB. Cytotoxicity and embryotoxicity assays revealed a dose-dependent effect (lethal concentration 50%, or LC50 value 82.4 µg/mL), with no adverse effects on developing embryos at the tested doses (5-80 µg/mL), suggesting the extracts are safe for clinical use, even during pregnancy. The combined extracts of I. coccinea and R. arboreum exhibit potent antioxidant and anti-inflammatory effects without causing cytotoxic or embryotoxic effects, even at higher concentrations. This indicates their potential for safe clinical application in treating oxidative and inflammatory diseases.

A method for the quantitative analysis of Lycium barbarum polysaccharides (LBPs) using Fourier-transform infrared spectroscopy (FTIR): From theoretical computation to experimental application

Lycium barbarum polysaccharides (LBPs) is one of the most important active substances in Lycium barbarum (LB). It is a challenge to quantitatively determine the content due to their complex structures and lack of suitable reference standard in practice. In this study, a quantitative analysis method of LBPs in LB was established based on Fourier-transform infrared spectroscopy (FTIR). The stretching vibration of CO on the pyranose ring of saccharide at 921 cm−1 was selected as the characteristic absorption band by theoretical calculation, which can’t be impacted by the preparation methods and interfered by the component monosaccharides. The molecular weight CRM of dextran (Mw 63.3 kDa) served as the reference standard. The introducing internal standard (KSCN) can obtain a good precision (RSD = 1.10 %) and effectively compensate for the analysis errors caused by the environment, quality loss and uneven distribution during the tablet pressing processes. The methodological verification suggested that the method had good accuracy according to the recovery rate (96.61 %–105.45 %) and the blank recovery (92.39 %–99.37 %), respectively. The LOD and LOQ of CRMD were 0.10 mg and 0.32 mg, respectively. The polysaccharide content of LB from 24 different regions (0.50–2.54 %) and 10 batches of LB extracts (7.09–10.56 %) determined by the developed method less than the ones using phenol–sulfuric acid assay (1.95 %–4.83 % for LB and 9.83–15.53 % for extracts, respectively). The established method based on FTIR could be served as a supplement to phenol–sulfuric acid assay and a rapid quantitative assay for polysaccharides products. In additional, this study provided a new idea for the quantitative analysis of plant polysaccharides.

Phytochemical profiling and bioactivity analysis of Citrus japonica leaves volatile oil from Palestine

BackgroundExploring the therapeutic potential of unutilized plant parts from agricultural crops represents a promising strategy for discovering novel medications with high positive economic value. This study aimed to characterize the composition, antidiabetic, anti-obesity, antioxidant, and cytotoxic effects of volatile oil (VO) extracted from the leaves of Citrus japonica trees. This is the first research to assess the C. japonica VO’s anti-obesity, anti-lipase, and cytotoxic properties. Using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assays and gas chromatography–mass spectrometry (GC–MS) analysis, the components of VO and its capacity to suppress the growth of cancer and other abnormal cells were ascertained, respectively. Stable 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals, p-nitrophenyl butyrate (PNPB), and dinitrosalicylic acid (DNSA) assays were employed to determine antioxidant, anti-obesity, and antidiabetic activities, respectively.ResultsThe C. japonica leaf showed significant diversity in type and percentage of VO molecules. Overall, 45 compounds were identified in the VO, constituting 99.69% of the total oil composition. γ-Muurolene (28.12%), β-eudesmol (10.93%), γ-eudesmol (8.44%), germacrene B (7.39%), and elemol (7.27%) are the major characterized molecules. According to the inhibition percentage results of DPPH free radicals, porcine pancreatic lipase, and α-amylase, the VO exhibits strong antioxidant properties and weak inhibitory effects on lipase and α-amylase enzymes. The C. japonica VO showed a moderate cytotoxic effect against Hep3B and considerable activity on B16F1, CaCo-2, HeLa, MCF-7, and HepG2, with IC50 doses in the range of 69.7–171.96 µg/mL. The VO cytotoxic effect IC50 against the normal cell line LX-2 was 224.95 µg/mL.ConclusionThe current study collectively presented the chemical constituents of C. japonica leaf VO from Palestine for the first time and demonstrated its inhibitory effects against DPPH free radicals, porcine pancreatic lipase, and α-amylase. The results suggest that C. japonica leaf VO has the potential to be used as a natural supplement to prevent or treat cancer, as well as in the food industry as a natural antioxidant.Graphical

Radical-Scavenging and Anti-inflammatory Activities of Fermented Eucheuma cottonii from Lombok

Eucheuma cottonii, a widely cultivated seaweed from the Island of Lombok, Indonesia, contains various bioactive compounds. Its utilization, however, is mainly limited based on its high carbohydrate content. Nevertheless, plant fermentation using lactic acid bacteria is renowned for increasing bioactive compounds and enhancing bioactivities. This study unveiled the potential of Lactobacillus plantarum-fermented E. cottonii as a functional food. E. cottonii was fermented using L. plantarum for 24 h. The folin-Ciocalteau method was used to determine the total phenolic content. The antioxidant capacity was measured using a 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay. Anti-inflammatory activity was carried out using cyclooxygenase (COX) inhibition assay against cervical cancer (HeLa) and colon cancer (WiDr) cells. Overall, fermentation successfully enhanced the bioactivities of E. cottonii. Fermented products exhibited higher antioxidant capacity than unfermented ones. Interestingly, the bioactivities only showed a moderate correlation with total phenolic content. Regarding anti-inflammatory activity, fermented extracts exhibited higher cyclooxygenase inhibition against HeLa cells, whereas no significant differences were observed between the fermented and unfermented products in WiDr cells. These findings indicate that L. plantarum-fermented E. cottonii holds promise to be a profitable functional food and has the potential to be utilized as an additional food therapy for cancer treatment. Keywords: Eucheuma cottonii; fermentation; Lactobacillus plantarum; antioxidant, anti-inflammation.

Screening of potential targets and small-molecule drugs related to lipid metabolism in ovarian cancer based on bioinformatics

Backgroundabout 70 % of ovarian cancer (OC) patients with postoperative chemotherapy relapse within 2–3 years due to drug resistance and metastasis, and the 5-year survival rate is only about 30 %. Lipid metabolism plays an important role in OC. We try to explore the potential targets and drugs related to lipid metabolism to provide clues for the treatment of OC. Methodsthe gene expression profiles of OC and normal ovarian tissue samples were obtained from the cancer genome atlas (TCGA) and genotype-tissue expression databases (GTEx). The differentially expressed genes (DEGs) were analyzed. Lipid metabolism related genes (LMRGs) were downloaded from MSigDB database. The DEGs related to lipid metabolism in OC was obtained by intersection. And gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) analyses were performed. The protein-protein interaction (PPI) network of lipid metabolism related DEGs was constructed, and seven algorithms were used to screen core potential target genes. Its expression in OC and prognostic ability were analyzed by Univariate Cox. Cmap database mining OC lipid metabolism related potential small-molecular drugs and docking. CCK8, scratch assay, transwell test and free fatty acid (FFA) assay, fluorescence detection of cellular fatty acid uptake, and the reactivity assay of CPT1A were used to detect the biological effects of drugs on OC cell.Rreverse transcription PCR(RT-qPCR) and WesternBlot were performed to measure the expression of core targets. Results437 DEGs related to lipid metabolism of OC were screened. GO and KEGG analysis showed that these DEGs were lipid metabolism, fatty acid metabolism, sphingolipid metabolism, PPAR signal pathway and so on. The PPI network based on lipid metabolism DEGs consists of 301 nodes and 1107 interaction pairs, and 6 core target genes were screened. ROC curve analysis showed that all of the 6 genes could predict the prognosis of OC. Three small molecular drugs Cephaeline, AZD8055 and GSK-1059615 were found by cmap and molecular docking showed that they all had good binding ability to target gene. Cephaeline has the strongest inhibitory effect on SKOV3 cells of OC, and could significantly inhibit cell migration and invasion regulate the mRNA and protein expression of some targets, and inhibit lipid metabolism process in ovarian cancer cells. Conclusionsix OC potential genes related to lipid metabolism were identified and verified, which can be used as potential biomarkers and therapeutic targets to evaluate the prognostic risk of OC patients. In addition, three small-molecular drugs that may be effective in the treatment of OC were unearthed, among which Cephaeline has the most potential. We speculate that Cephaeline may target six genes to inhibit progression of OC by affecting lipid metabolism.

Development of chitosan nanoparticle loaded with Tricholoma fracticum extract and evaluation of in vitro antioxidant activity

SummaryThe objective of this study was to develop Tricholoma fracticum extract‐loaded chitosan nanoparticles (TFNPs) by ionic gelation method and to evaluate their in vitro antioxidant activity. Phenolic and flavonoid contents in the T. fracticum extract were measured spectrophotometrically and chromatographically. Characterisation of NPs was evaluated by field emission scanning electron microscopy (FE‐SEM), transmission electron microscopy (TEM), ZETA analysis, Fourier‐transform infrared spectroscopy (FT‐IR), UV–visible spectroscopy (UV–Vis) and thermogravimetric analysis (TGA). In vitro antioxidant capacity was determined using 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) and 2,2′‐azino‐bis(3‐ethylbenzothiazoline‐6‐sulfonic acid) (ABTS) assays. The phenolic and flavonoid contents in the T. fracticum extract were measured as 7.1 ± 0.3 mg Gallic Acid Equivalent/g extract and 5.5 ± 0.6 mg Quercetin Equivalent/g extract, respectively. The particle size, polydispersity index (PDI) and zeta potential of Blank NP1 and TFNP were 265.5 ± 15.8 nm, 0.4, 38.7 ± 4.0 mV and 333.2 ± 16.3 nm, 0.4, 37.0 ± 4.1 mV, respectively. The highest antioxidant activity was observed in TFNP, followed by T. fracticum extract, chitosan and blank NP, respectively. The preserved or enhanced antioxidant activity observed in the encapsulated T. fracticum extract indicates the potential for loading similar mushroom extracts onto chitosan and thus preserving their bioactive properties.

Degraded sweet corn cob polysaccharides modulate T2DM-induced abnormalities in hepatic lipid metabolism via the bile acid-related FXR-SHP and FXR-FGF15-FGFR4 pathways

Type 2 diabetes mellitus (T2DM) is a metabolic disorder characterized by dysregulation of glucose and lipid metabolism. This study aimed to elucidate the mechanism through which a degraded sweet corn cob polysaccharide (UE-DSCCP-A) mitigates T2DM by modulating hepatic lipid metabolism. Biochemical indices pertinent to lipid metabolism were assessed, and liver pathology was examined in T2DM mice following UE-DSCCP-A treatment. Additionally, metabolomics, PCR, and Western blot analyses were employed to investigate the underlying mechanisms involved. These findings indicated that UE-DSCCP-A ameliorated hepatic lipid metabolism disorders, decreased lipid accumulation, and mitigated hepatic fibrosis. Untargeted metabolomics analysis revealed that UE-DSCCP-A modulated pathways associated with steroid biosynthesis and bile acid synthesis and metabolism in T2DM mice. The bile acid assay results demonstrated that UE-DSCCP-A treatment reduced bile acid levels in both the serum and liver but increased fecal bile acid levels in T2DM mice. Furthermore, alterations in bile acid distribution within the liver were observed. UE-DSCCP-A has the capacity to activate the hepatic FXR-SHP pathway as well as the gut-liver axis involving FXR-FGF15-FGFR4 signaling. Consequently, UE-DSCCP-A is capable of modulating critical target genes and proteins associated with bile acid synthesis and metabolism, regulating steroid biosynthesis, and influencing bile acid synthesis and transport within the liver. Additionally, it has beneficial effects on lipid metabolism disorders in individuals with T2DM. Thus, UE-DSCCP-A represents a promising candidate for functional foods with inherent hypoglycemic properties.

Antibacterial and Antioxidant Activities of Hydroalcoholic and Phenolic Extracts from Ternstroemia dentisepala and T. lineata Leaves.

Traditional Mexican medicine commonly uses infusions of Ternstroemia spp. to treat insomnia, injuries, and infections. The antibacterial activities of Ternstroemia dentisepala and Ternstroemia lineata were evaluated for the first time against a panel of Gram-positive and Gram-negative bacteria that have implications for human health, including Enterococcus faecalis, Streptococcus agalactiae, Streptococcus pyogenes, Salmonella typhi, Pseudomonas aeruginosa, and Vibrio parahaemolyticus. Furthermore, the scavenging potential of the hydroalcoholic (HAEs) and total phenolic extracts (TPEs) from the leaves of both plants by a 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) assay (ABTS•+) was determined. Also, the total phenolic contents of the HAEs using the Folin-Ciocalteu reagent were assayed. T. dentisepala HAE and TPE were active against all bacterial strains tested, with a minimum inhibitory concentration between 1.0 and 6.0 mg/mL, with the last one being the most active. However, the T. lineata extracts only demonstrated effectiveness against S. typhi and P. aeruginosa. The TPEs from T. dentisepala and T. lineata improved the activity by approximately 30% in all bacteria tested in comparison with the HAEs. The T. dentisepala HAE had a higher total phenolic content than the T. lineata extract, which was consistent with its ABTS•+-scavenging activity. The two HAEs had different chemical profiles, mostly because of the types and amounts of phenolic compounds they contained. These profiles were obtained using thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), and proton nuclear magnetic resonance (1H NMR) experiments.

CHAMP: A Centrifugal Microfluidics-Based CRISPR/Cas12b-Combined Real-Time LAMP One-Pot Method for Mycoplasma pneumoniae Infection Diagnosis.

The Mycoplasma pneumoniae outbreak poses health risks to community residents. However, it still has limitations for current clinical diagnostic methods (qPCR nucleic acid assay or IgM immunoassay), including specialized handling, expensive equipment, prolonged turnaround time, and false positives and negatives, highlighting the need to improve clinical diagnostic methods. Herein, we present a novel centrifugal microfluidics-based method for rapidly diagnosing M. pneumoniae infections (CHAMP system). This user-friendly method combines CRISPR/Cas12b and real-time loop-mediated isothermal amplification (LAMP) in a one-pot reaction, offering high sensitivity, specificity, and simplicity for methodology. By adding fully automated nucleic acid magnetic bead-extracted samples to a prepackaged centrifugal microfluidics chip, 48 samples can be automated tested simultaneously within 15 to 60 min at 60 °C. 427 clinical nasopharyngeal swab specimens were used for validation, demonstrating good positive and negative predictive values and good diagnostic sensitivity, specificity, and significant time savings. This method is particularly suitable for detecting low nucleic acid copies of M. pneumoniae samples.

Effects of high temperature short time (HTST) pasteurization on milk and whey during commercial whey protein concentrate production

Two pasteurization steps are often used in the preparation of whey protein concentrate (WPC) before evaporation into a dry product. The Pasteurized Milk Ordinance (PMO) in the United States requires that raw bovine milk be pasteurized using a process that meets minimum heat treatment requirements to achieve reductions in pertinent microorganisms. In addition, WPC produced from USDA-approved plants must comply with CFR Subpart B §58.809, which dictates that all fluid whey used in the manufacture of dry whey products shall be pasteurized before being condensed. These heat treatments are effective at inactivating the most thermally resistant bacterium, such as Coxiella burnetii; however, they can also alter milk proteins-inducing denaturation, aggregation and reduced bioactivity. Though the impact of thermal treatments on whey proteins has been examined, the specific influence of 2 high-temperature-short-time (HTST) pasteurization steps on the retention of proteins in WPC remains unknown. This study aimed to investigate the effect of commercial-scale HTST pasteurization of both raw milk and the resulting sweet whey on the products' overall protein profile. Three distinct batches of raw milk (RM) and corresponding pasteurized milk (PM), the resulting whey (RW) and pasteurized whey (PW) produced at commercial scale were analyzed. Assessments of denaturation were conducted through solubility testing at pH 4.6 and hydrophobicity evaluation via anilinonaphthalene-1-sulfonic acid assay (ANS). Additionally, enzyme-linked immunosorbent assay (ELISA), PAGE (PAGE) and liquid chromatography tandem mass spectroscopy (LC-MS/MS) were employed to compare the retention of key bioactive proteins before and after each HTST pasteurization step. The percentage of soluble whey protein decreased from RM to PM and from RW to PW, but no significant differences were observed via hydrophobicity assay. ELISA revealed a significant reduction in key bioactive proteins, such as lactoferrin, immunoglobulin A and immunoglobulin M, but not immunoglobulin G, after HTST pasteurization of RM and RW. PAGE and LC-MS/MS revealed a significant decrease in the retention of lactoferrin and key milk fat globular membrane proteins, such as xanthine dehydrogenase oxidase/xanthine oxidase, lactadherin and fatty acid binding protein. Additionally, xanthine oxidase activity was significantly reduced after HTST pasteurization of milk and whey. This research helps to identify the limitations of the current processing techniques used in the dairy industry and could lead to innovation in improving the retention of bioactive proteins.

Potential antimicrobial and fruit juice clarification activity of amylase enzyme from Bacillus strains

The hydrolytic enzyme, amylase possesses wide industrial applications and its production from bacterial sources by submerged fermentation is much simplified and economical. The research aimed to characterize amylase-producing bacteria and evaluate their potential for amylase activity regarding antimicrobial and fruit juice clarification. In current study, Bacillus licheniformis, Bacillus amyloliquifaciens, Bacillus cereus, Bacillus subtilis and Bacillus paramycoides was identified by 16S rRNA sequencing. After submerged fermentation, amylase activity of bacteria was measured by 3, 5-dinitro salicylic acid (DNS) assay. A substantial amount of amylase (423.47 mg/ml) in crude extract was measured by Bradford protein assay. Later, ammonium sulfate (80 %) precipitated partially purified amylase showed 1.6 times enhanced amylase activity (1484.94 U/ml) compared to crude amylase (973.23 U/ml). For highest amylase production, 72 h of optimum fermentation period was recorded at pH 7 with 2 % starch as substrate. Potent thermophilic amylase activity was observed at 65 °C. In apple juice clarification activity of amylase, turbidity of juice was reduced to 54.18 %. Potential antimicrobial property of amylase was detected with largest zone of inhibition against Escherichia coli ATCC 25922 (22.36 mm) and Mucor sp. ATCC 48559 (22.45 mm). Considering promising amylase properties, amylase-producing Bacillus strains from rice mill soil can be fermented for large scale amylase production providing application for industrial purposes including fruit juice clarification and antimicrobial activities. It will also overthrow the requirement of employing expensive and harmful chemicals in fruit juice clarification and combating pathogens.

Green Ultrasound-Assisted Extraction of Bioactive Compounds from Cumari-Do-Pará Peppers (Capsicum chinense Jacq.) Employing Vegetable Oils as Solvents.

Capsaicin, carotenoids, and phenolic compounds from cumari-do-Pará peppers (Capsicum chinense Jacq.) harvested from two different locations in Pará, Brazil, and at different ripening stages were extracted by employing green methodologies as an alternative to organic solvents. Edible vegetable oils from soybeans (Glycine max), Brazilian nuts (Bertholettia excelsa H.B.), and palm olein were used in combination with ultrasonic-assisted extraction (UAE). The proximate composition of the pepper extracts and vitamin C were determined through AOAC methods, total phenolics and carotenoids were assessed by UV/Vis spectrophotometry, and capsaicin by high-performance liquid chromatography. Antioxidant cumari-do-Pará extract activities were evaluated by the ABTS radical scavenging and β-carotene/linoleic acid assays. The vegetable oils were suitable for extracting and preserving bioactive pepper compounds, especially mature ones harvested from Igarapé-Açu. Bioactive compound content and antioxidant activity varied with harvesting location and ripening stage. Soybean oil was the most effective in extracting bioactive pepper compounds, particularly carotenoids, with 69% recovery. Soybean oil extracts enriched in capsaicin, carotenoids, and phenolics obtained from cumari-do-Pará can be used as spices in foodstuffs and/or as additives in pharmaceutical and nutraceutical formulations. Edible vegetable oils combined with UAE are promising for bioactive compound extraction, representing an environmentally friendly, safe, low-cost, versatile, and fast alternative.

3,3’4-trimethoxy-4’-rutinosylellagic acid and its acetylated derivative: Antioxidant activity and antiproliferative effects on breast cancer cells and molecular docking study

Cancers account for many deaths worldwide and natural compounds and their derivatives are interesting chemotherapeutic agents for cancer drug development. In this study, a natural compound 3,3’4-trimethoxy-4’-rutinosylellagic acid (TR2) and its acetylated derivative 3,3’4-trimethoxy-4’-hexaacetylrutinosylellagic acid (TR22) were evaluated for their antioxidant and anticancer effects against estrogen sensitive (MCF-7) and estrogen non-sensitive (MDA-MB 231) breast adenocarcinoma. In the β-Carotene-linoleic acid assay, DPPH• radical scavenging and CUPRAC assay, the compound TR2 had better activity than the standard α-Tocopherol, while in the ABTS•+ assay, it was more active than both standards α- α-Tocopherol and BHA. Both compounds had good antioxidant effects with TR2 being more active than TR22. Both compounds inhibited growth of breast carcinoma cells when compared to the untreated controls after 72 h. Compound TR22 significantly (p < 0.001) inhibited proliferation of both MCF-7 and MDA-MB 231 breast carcinoma cell lines suggesting that acetylation reaction improves inhibition of breast cancer cells growth. On the contrary, TR2 exhibited better inhibitory effect of clone formation than TR22 suggesting that acetylation reduces the activity in this assay. Both compounds inhibited migration of the cancer cells when compared to the untreated control cells and compound TR2 exhibited greater cellular anti-migration effect than TR22 at the same concentration and after the same period of incubation. Molecular docking studies supplemented the results and revealed that TR2 and TR22 had appreciable interactions with tyrosine kinase with negative binding energies suggesting that they are potent receptor tyrosine kinase inhibitors which can impede on cancer progression.

Metabolic signatures of metabolites of the purine degradation pathway in human plasma using HILIC UHPLC–HRMS

The metabolic disorders in the purine degradation pathway have proven to be closely associated with several human diseases. However, the etiology is not yet fully understood. Profile assay of purine intermediates and uric acid involved in the metabolic pathway can provide additional insight into the nature and severity of related diseases. Purine metabolites are endogenous chemicals with high hydrophilicity, polarity, and similar structures, thus there is a great need for a specific method to quantify them directly in biological fluids with a short running time. Herein, eight purine degradation pathway metabolites, including xanthine, hypoxanthine, guanine, xanthosine, inosine, guanosine, adenosine and uric acid, in human plasma were quantitatively measured using hydrophilic interaction chromatography-tandem high-resolution mass spectrometry (HILIC-HRMS) in a short running time of 10 min. The method was systematically validated for specificity, linearity of the calibration curve, the limit of detection, the limit of quantification, the lower limit of quantification, precision, accuracy, extraction recovery, matrix effect, and stability. The results showed that the method was linear (R2 > 0.99), accurate (the intra- and inter-day recoveries of all analytes ranged from 90.0 % to 110.0 %), and precise (the intra- and inter-day precisions were less than 6.7 % and 8.9 %, respectively) with the lower limits of quantification ranging from 3 to 10,000 ng/mL. The extraction recoveries and matrix effects were repeatable and stable. All the analytes were stable in the autosampler and could be subject to three freeze-thaw cycles. The developed method was ultimately applied to 100 plasma specimens from healthy individuals. The results showed that the concentrations of different purine metabolites varied dramatically in plasma specimens. Diet and body mass index (BMI) were the most significant factors determining purine levels, followed by drinking and sex. Age, smoking and bedtime showed a very weak correlation with purine metabolism. The findings of the present work reveal the characteristics of purine metabolism in human plasma under non-pathological conditions. The results also highlight the factors that can cause changes in purine metabolism, which are useful in developing effective treatment strategies for metabolic disorders of purines, particularly for those caused by lifestyle factors.

Investigating the synergistic antibacterial effects of chlorogenic and p-coumaric acids on Shigella dysenteriae

Chlorogenic acid (CGA) is a well-known plant secondary metabolite exhibiting multiple physiological functions. The present study focused on screening for synergistic antibacterial combinations containing CGA. The combination of CGA and p-coumaric acid (pCA) exhibited remarkably enhanced antibacterial activity compared to that when administering the treatment only. Scanning electron microscopy revealed that a low-dose combination treatment could disrupt the Shigella dysenteriae cell membrane. A comprehensive analysis using nucleic acid and protein leakage assay, conductivity measurements, and biofilm formation inhibition experiments revealed that co-treatment increased the cell permeability and inhibited the biofilm formation substantially. Further, the polyacrylamide protein- and agarose gel-electrophoresis indicated that the proteins and DNA genome of Shigella dysenteriae severely degraded. Finally, the synergistic bactericidal effect was established for fresh-cut tomato preservation. This study demonstrates the remarkable potential of strategically selecting antibacterial agents with maximum synergistic effect and minimum dosage exhibiting excellent antibacterial activity in food preservation.