Bioactive ZnO Nanoparticles: Biosynthesis, Characterization and Potential Antimicrobial Applications.

In recent years, biosynthesized zinc oxide nanoparticles (ZnONPs) have gained tremendous attention because of their safe and non-toxic nature and distinctive biomedical applications. A diverse range of microbes (bacteria, fungi and yeast) and various parts (leaf, root, fruit, flower, peel, stem, etc.) of plants have been exploited for the facile, rapid, cost-effective and non-toxic synthesis of ZnONPs. Plant extracts, microbial biomass or culture supernatant contain various biomolecules including enzymes, amino acids, proteins, vitamins, alkaloids, flavonoids, etc., which serve as reducing, capping and stabilizing agents during the biosynthesis of ZnONPs. The biosynthesized ZnONPs are generally characterized using UV-VIS spectroscopy, TEM, SEM, EDX, XRD, FTIR, etc. Antibiotic resistance is a serious problem for global public health. Due to mutation, shifting environmental circumstances and excessive drug use, the number of multidrug-resistant pathogenic microbes is continuously rising. To solve this issue, novel, safe and effective antimicrobial agents are needed urgently. Biosynthesized ZnONPs could be novel and effective antimicrobial agents because of their safe and non-toxic nature and powerful antimicrobial characteristics. It is proven that biosynthesized ZnONPs have strong antimicrobial activity against various pathogenic microorganisms including multidrug-resistant bacteria. The possible antimicrobial mechanisms of ZnONPs are the generation of reactive oxygen species, physical interactions, disruption of the cell walls and cell membranes, damage to DNA, enzyme inactivation, protein denaturation, ribosomal destabilization and mitochondrial dysfunction. In this review, the biosynthesis of ZnONPs using microbes and plants and their characterization have been reviewed comprehensively. Also, the antimicrobial applications and mechanisms of biosynthesized ZnONPs against various pathogenic microorganisms have been highlighted.

Are Fermented Foods Effective against Inflammatory Diseases?

Fermented foods have been used over the centuries in various parts of the world. These foods are rich in nutrients and are produced naturally using various biological tools like bacteria and fungi. Fermentation of edible foods has been rooted in ancient cultures to keep food for preservation and storage for a long period of time with desired or enhanced nutritional values. Inflammatory diseases like rheumatoid arthritis, osteoarthritis, and chronic inflammatory pain are chronic disorders that are difficult to treat, and current treatments for these disorders fail due to various adverse effects of prescribed medications over a long period of time. Fermented foods containing probiotic bacteria and fungi can enhance the immune system, improve gastrointestinal health, and lower the risk of developing various inflammatory diseases. Foods prepared from vegetables by fermentation, like kimchi, sauerkraut, soy-based foods, or turmeric, lack proper clinical and translational experimental studies. The current review has focused on the effectiveness of various fermented foods or drinks used over centuries against inflammation, arthritis, and oxidative stress. We also described potential limitations on the efficacies or usages of these fermented products to provide an overarching picture of the research field.

CRISPR-Based Programmable Nucleic Acid-Binding Protein Technology Can Specifically Detect Fatal Tropical Disease-Causing Pathogens.

Diagnostic approaches capable of ultrasensitive pathogen detection from low-volume clinical samples, running without any sophisticated instrument and laboratory setup, are easily field-deployable, inexpensive, and rapid, and are considered ideal for monitoring disease progression and surveillance. However, standard pathogen detection methods, including culture and microscopic observation, antibody-based serologic tests, and primarily polymerase chain reaction (PCR)-oriented nucleic acid screening techniques, have shortcomings that limit their widespread use in responding to outbreaks and regular diagnosis, especially in remote resource-limited settings (RLSs). Recently, clustered regularly interspaced short palindromic repeats (CRISPR)-based programmable technology has emerged to challenge the unmet criteria of conventional methods. It consists of CRISPR-associated proteins (Cas) capable of targeting virtually any specific RNA or DNA genome based on the guide RNA (gRNA) sequence. Furthermore, the discovery of programmable trans-cleavage Cas proteins like Cas12a and Cas13 that can collaterally damage reporter-containing single-stranded DNA or RNA upon formation of target Cas-gRNA complex has strengthened this technology with enhanced sensitivity. Current advances, including automated multiplexing, ultrasensitive single nucleotide polymorphism (SNP)-based screening, inexpensive paper-based lateral flow readouts, and ease of use in remote global settings, have attracted the scientific community to introduce this technology in nucleic acid-based precise detection of bacterial and viral pathogens at the point of care (POC). This review highlights CRISPR-Cas-based molecular technologies in diagnosing several tropical diseases, namely malaria, zika, chikungunya, human immunodeficiency virus and acquired immunodeficiency syndrome (HIV-AIDS), tuberculosis (TB), and rabies.

Integrated Multilayer Omics Reveals the Genomic, Proteomic, and Metabolic Influences of Histidyl Dipeptides on the Heart.

Background Histidyl dipeptides such as carnosine are present in a micromolar to millimolar range in mammalian hearts. These dipeptides facilitate glycolysis by proton buffering. They form conjugates with reactive aldehydes, such as acrolein, and attenuate myocardial ischemia-reperfusion injury. Although these dipeptides exhibit multifunctional properties, a composite understanding of their role in the myocardium is lacking. Methods and Results To identify histidyl dipeptide-mediated responses in the heart, we used an integrated triomics approach, which involved genome-wide RNA sequencing, global proteomics, and unbiased metabolomics to identify the effects of cardiospecific transgenic overexpression of the carnosine synthesizing enzyme, carnosine synthase (Carns), in mice. Our result showed that higher myocardial levels of histidyl dipeptides were associated with extensive changes in the levels of several microRNAs, which target the expression of contractile proteins, ß-fatty acid oxidation, and citric acid cycle (TCA) enzymes. Global proteomic analysis showed enrichment in the expression of contractile proteins, enzymes of ß-fatty acid oxidation, and the TCA in the Carns transgenic heart. Under aerobic conditions, the Carns transgenic hearts had lower levels of short- and long-chain fatty acids as well as the TCA intermediate-succinic acid; whereas, under ischemic conditions, the accumulation of fatty acids and TCA intermediates was significantly attenuated. Integration of multiple data sets suggested that ß-fatty acid oxidation and TCA pathways exhibit correlative changes in the Carns transgenic hearts at all 3 levels. Conclusions Taken together, these findings reveal a central role of histidyl dipeptides in coordinated regulation of myocardial structure, function, and energetics.

The bifidogenic effects and dental plaque deformation of non-digestible isomaltooligosaccharides synthesized by dextransucrase and alternansucrase.

Non-digestible isomaltooligosaccharides (NDIMOS) are functional food and beverage ingredients that contributed to human health benefits through metabolism of gastrointestinal microorganism. In this study, NDIMOS were synthesized by combine dextransucrase from Leuconostoc mesenteroides B512F/KM and alternansucrase from L. mesenteroides NRRL 1355CF10/KM using sucrose as substrate and maltose as acceptor. Their digestibility was confirmed by using digestive enzymes including α-amylase and amyloglucosidase. NDIMOS inhibited insoluble glucan formation through mutansucrase from Streptococcus mutans. The bifidogenic effect of NDIMOS was investigated by growth of four strains of Bifidobacterium in MRS broth containing NDIMOS, compared with MRS broth contain glucose and negative control. Additionally, Bifidobacterium bifidum or Bifidobacterium adolescentis inhibited the growth of Salmonella enterica serovar typhimurium when they were co-cultivation in MRS broth containing NDIMOS. These results suggested that NDIMOS is potential functional ingredient for food, beverage, and pharmaceutical application.

Pinibacter aurantiacus gen. nov., sp. nov., isolated from rhizospheric soil of a pine tree.

A Gram-stain-negative, aerobic and rod-shaped novel bacterial strain, designated MAH-26T, was isolated from rhizospheric soil of a pine tree. The colonies were orange coloured, smooth, spherical and 0.7-1.8 mm in diameter when grown on Reasoner's 2A (R2A) agar for 2 days. Strain MAH-26T was able to grow at 10-40 °C, at pH 6.0-9.0 and with 0-1.0 % NaCl. Cell growth occurred on nutrient agar, R2A agar, tryptone soya agar and Luria-Bertani agar. The strain gave positive results in oxidase and catalase tests. Strain MAH-26T was closely related to Flavihumibacter sediminis CJ663T and Parasegetibacter terrae SGM2-10T with a low 16S rRNA gene sequence similarity (92.8 and 92.9 %, respectively) and phylogenetic analysis indicated that the strain formed a distinct phylogenetic lineage from the members of the closely related genera of the family Chitinophagaceae. Strain MAH-26T has a draft genome size of 6 857 405 bp, annotated with 5173 protein-coding genes, 50 tRNA and two rRNA genes. The genomic DNA G+C content was 41.5 mol%. The predominant isoprenoid quinone was menaquinone 7. The major fatty acids were identified as iso-C15:0, iso-C15:1 G and iso-C17:0 3OH. On the basis of phylogenetic inference and phenotypic, chemotaxonomic and molecular properties, strain MAH-26T represents a novel species of a novel genus of the family Chitinophagaceae, for which the name Pinibacter aurantiacus gen. nov., sp. nov. is proposed. The type strain of Pinibacter aurantiacus is MAH-26T (=KACC 19749T=CGMCC 1.13701T).

Prospects for Protective Potential of Moringa oleifera against Kidney Diseases.

Kidney diseases are regarded as one of the major public health issues in the world. The objectives of this study were: (i) to investigate the causative factors involved in kidney disease and the therapeutic aspects of Moringa oleifera, as well as (ii) the effectiveness of M. oleifera in the anti-inflammation and antioxidant processes of the kidney while minimizing all potential side effects. In addition, we proposed a hypothesis to improve M. oleifera based drug development. This study was updated by searching the key words M. oleifera on kidney diseases and M. oleifera on oxidative stress, inflammation, and fibrosis in online research databases such as PubMed and Google Scholar. The following validation checking and scrutiny analysis of the recently published articles were used to explore this study. The recent existing research has found that M. oleifera has a plethora of health benefits. Individual medicinal properties of M. oleifera leaf extract, seed powder, stem extract, and the whole extract (ethanol/methanol) can up-increase the activity of antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH), while decreasing the activity of inflammatory cytokines such as TNF-α, IL-1ß, IL-6, and COX-2. In our study, we have investigated the properties of this plant against kidney diseases based on existing knowledge with an updated review of literature. Considering the effectiveness of M. oleifera, this study would be useful for further research into the pharmacological potential and therapeutic insights of M. oleifera, as well as prospects of Moringa-based effective medicine development for human benefits.

Wastewater based surveillance system to detect SARS-CoV-2 genetic material for countries with on-site sanitation facilities: an experience from Bangladesh

The presence of SARS-CoV-2 genetic materials in wastewater has become a matter of grave for many countries of the world. Wastewater based epidemiology, in this context, emerged as an important tool in developed countries where proper sewage system is available. Due to the recent shift in the spread of the infection from urban to rural areas, it is now equally important to develop a similar mechanism for rural areas as well. Considering the urgency of the issue a study was conducted in 14 districts of Bangladesh and a total of 238 sewage samples were collected in two different periods from December 2020 to January 2021. We are the first to propose a surveillance system for both urban and rural areas where a proper sewage system is absent. Based on RT-PCR analysis of the water samples, in more than 92% of cases, we found the presence of the SARS-COV-2 gene (ORF1ab, N, and Internal Control-IC). The trend of Ct value varies for different study locations. The spread of genetic material for on-site ({Delta}m = 0.0749) sanitation system was found more prominent than that of off-site sewage system ({Delta}m = 0.0219); which indicated the shift of genetic material from urban to rural areas. Wastewater samples were also measured for physicochemical parameters, including pH (6.30 - 12.50) and temperature (22.10 - 32.60) {o}C. The highest viral titer of 1975 copy/mL in sewage sample was observed in a sample collected from the isolation ward of the SARS-COV-2 hospital. Additionally, a correlation was found between bacterial load and SARS-CoV-2 genetic materials. The results indicated the association of increased Ct values with decreasing number of patients and vice versa. The findings reported in this paper contributed to the field of wastewater-based epidemiology dealing with SARS-COV-2 surveillance for developing countries where proper sewage system is absent and highlighting some of the challenges associated with this approach in such settings. HighlightsO_LIDevelopment of wastewater-based surveillance system based on on-site sanitation system for developing countries. C_LIO_LIAssociation of different environmental parameters with the presence of SARS CoV-2 genetic material in wastewater. C_LIO_LIPrediction of the viral concentration of sewage system using viral load and copy number parameter. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=122 SRC="FIGDIR/small/21261347v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1f7847dorg.highwire.dtl.DTLVardef@11b2c93org.highwire.dtl.DTLVardef@10b9fe1org.highwire.dtl.DTLVardef@2d9e8d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Sex-specific epidemiological and clinical characteristics of Covid-19 patients in the southeast region of Bangladesh

BackgroundCOVID-19 has become a global pandemic with a high growth rate of confirmed cases. In Bangladesh, both mortality and affected rates are increasing at an alarming rate. Therefore, more comprehensive studies of the epidemiological and clinical characteristics of COVID-19 are required to control this pandemic. PurposeThe present study aimed to compare and analyze the sex-specific epidemiological, clinical characteristics, comorbidities, and other information of confirmed COVID-19 patients from the southeast region in Bangladesh for the first time. Methods385 lab-confirmed cases were studied out of 2,471 tested samples between 5 June and 10 September 2020. RT-PCR was used for COVID-19 identification and SPSS (version 25) for statistical data analysis. ResultsWe found that male patients were roughly affected compared to females patients (male 74.30% vs. female 25.7%) with an average age of 34.86 {+/-} 15.442 years, and B (+ve) blood group has been identified as a high-risk factor for COVID-19 infection. Workplace, local market, and bank were signified as sex-specific risk zone (p < 0.001). Pre-existing medical conditions such as diabetes, hypertension, cardiovascular and respiratory diseases were identified among the patients. Less than half of the confirmed COVID-19 cases in the southeast region were asymptomatic (37.73%) and more prevalent among females than males (male vs. female: 36.84% vs. 40.51%, p = 0.001). ConclusionsThe findings may help health authorities and the government take necessary steps for identification and isolation, treatment, prevention, and control of this global pandemic.

CRISPR is a useful biological tool for detecting nucleic acid of SARS-CoV-2 in human clinical samples.

The recent pandemic of novel coronavirus disease (COVID-19) has spread globally and infected millions of people. The quick and specific detection of the nucleic acid of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) remains a challenge within healthcare providers. Currently, quantitative reverse transcription-polymerase chain reaction (RT-qPCR) is the widely used method to detect the SARS-CoV-2 from the human clinical samples. RT-qPCR is expensive equipment and needs skilled personnel as well as lengthy detection time. RT-qPCR limitation needed an alternative healthcare technique to overcome with a fast and cheaper detection method. By applying the principles of CRISPR technology, several promising detection methods giving hope to the healthcare community. CRISPR-based detection methods include SHERLOCK-Covid, STOP-Covid, AIOD-CRISPR, and DETECTR platform. These methods have comparative advantages and drawbacks. Among these methods, AIOD-CRISPR and DETECTR are reasonably better diagnostic methods than the others if we compare the time taken for the test, the cost associated with each test, and their capability of detecting SARS-CoV-2 in the clinical samples. It may expect that the promising CRISPR-based methods would facilitate point-of-care (POC) applications in the CRISPR-built next-generation novel coronavirus diagnostics.

Comprehensive Two-Dimensional Gas Chromatography Mass Spectrometry-Based Metabolomics.

Compared to one-dimensional gas chromatography with mass spectrometry (GC-MS), GC × GC-MS provides significantly increased peak capacity, resolution, and sensitivity for analysis of complex biological samples. In the last decade, GC × GC-MS has been increasingly applied to the discovery of metabolite biomarkers and elucidation of metabolic mechanisms in human diseases. The recent development of coupling GC × GC with a high-resolution mass spectrometer further accelerates these metabolomic applications. In this chapter, we will briefly review the instrumentation, sample preparation, data analysis, and applications of GC × GC-MS-based metabolomic analysis.

Helicobacter pylori infection and micronutrient deficiency in pregnant women: a systematic review and meta-analysis.

BACKGROUND: Over the last few years, epidemiological studies have shown that infection with Helicobacter pylori has a major effect on micronutrient deficiency as well as on adverse pregnancy outcomes. Importantly, there are gaps in understanding the linkage of H. pylori infection with micronutrients deficiency in pregnant women. OBJECTIVE: We conducted a systematic review and meta-analysis to estimate the association between H. pylori infection and micronutrient deficiencies in pregnant women. METHODS: A systematic literature search was conducted for relevant articles using PubMed, Web of Science, and Scopus database from inception to March 2020. The OR with 95% CIs was determined by meta-analysis of data extracted from the selected studies. RESULTS: From 2384 primary articles, 6 studies were selected for systematic reviews and 4 studies distinctively (with 1274 participants: 553 cases and 721 controls) were selected for meta-analysis. The meta-analysed fixed effect model estimated the odds of having H. pylori infection was not significantly higher among pregnant women with micronutrient deficiencies than those without deficiencies (OR=1.12, 95% CI 0.88 to 1.42, p=0.37). In the subgroup analysis, no correlation was found between H. pylori infection and vitamin B12 (OR=0.74, 95% CI 0.45 to 1.21, p=0.22), folate (OR=1.07, 95% CI 0.73 to 1.58, p=0.73), and ferritin (OR=0.81, 95% CI 0.51 to 1.31, p=0.4). However, a positive correlation was found between iron-deficiency anaemia (IDA) and H. pylori infection (OR=16.23, 95% CI 4.19 to 62.93, p<0.0001) during pregnancy. CONCLUSION: H. pylori infection is associated with increased risk of IDA but not with deficiency of other micronutrients in pregnancy. PROSPERO REGISTRATION NUMBER: CRD42019135683.

First detection of SARS-CoV-2 genetic material in the vicinity of COVID-19 isolation centre through wastewater surveillance in Bangladesh

In the course of a COVID-19 pandemic, 0.33 million people got infected in Bangladesh, we made the first and successful attempt to detect SARS-CoV-2 viruses genetic material in the vicinity wastewaters of an isolation centre i.e. Shaheed Bhulu Stadium, situated at Noakhali. The idea was to understand the genetic loading variation, both temporal and distance-wise in the nearby wastewater drains when the number of infected COVID-19 patients is not varying much. Owing to the fact that isolation center, in general, always contained a constant number of 200 COVID-19 patients, the prime objective of the study was to check if several drains carrying RNA of coronavirus are actually getting diluted or accumulated along with the sewage network. Our finding suggested that while the temporal variation of the genetic load decreased in small drains over the span of 50 days, the main sewer exhibited accumulation of SARS-CoV-2 RNA. Other interesting finding displays that probably distance of sampling location in meters is not likely to have a significant impact on gene detection concentration, although the quantity of the RNA extracted in the downstream of the drain was higher. These findings are of immense value from the perspective of wastewater surveillance of COVID-19, as they largely imply that we do not need to monitor every wastewater system, and probably major drains monitoring may illustrate the city health. Perhaps, we are reporting the accumulation of SARS-CoV-2 genetic material along the sewer network i.e. from primary to tertiary drains. The study sought further data collection in this line to simulate conditions prevailed in the most of south Asian country and to shed further light on the temporal variation and decay/accumulation processes of the genetic load of the SARS-COV-2. HIGHLIGHTSO_LIFirst detection report of SARS-CoV-2 RNA in the wastewaters of Bangladesh. C_LIO_LIWe traced the genetic load in the vicinity of the isolation center with 200 COVID-19 patients. C_LIO_LIAbout 75% of positive samples were found during the monitoring period. C_LIO_LIMain sewer and canal exhibit temporal accumulation of genetic load of SARS-CoV-2 C_LIO_LIIn general, shredding shows a variable trend during the sampling tenure. C_LI

Cardiospecific Overexpression of ATPGD1 (Carnosine Synthase) Increases Histidine Dipeptide Levels and Prevents Myocardial Ischemia Reperfusion Injury.

BACKGROUND Myocardial ischemia reperfusion (I/R) injury is associated with complex pathophysiological changes characterized by pH imbalance, the accumulation of lipid peroxidation products acrolein and 4-hydroxy trans-2-nonenal, and the depletion of ATP levels. Cardioprotective interventions, designed to address individual mediators of I/R injury, have shown limited efficacy. The recently identified enzyme ATPGD1 (Carnosine Synthase), which synthesizes histidyl dipeptides such as carnosine, has the potential to counteract multiple effectors of I/R injury by buffering intracellular pH and quenching lipid peroxidation products and may protect against I/R injury. METHODS AND RESULTS We report here that ß-alanine and carnosine feeding enhanced myocardial carnosine levels and protected the heart against I/R injury. Cardiospecific overexpression of ATPGD1 increased myocardial histidyl dipeptides levels and protected the heart from I/R injury. Isolated cardiac myocytes from ATPGD1-transgenic hearts were protected against hypoxia reoxygenation injury. The overexpression of ATPGD1 prevented the accumulation of acrolein and 4-hydroxy trans-2-nonenal-protein adducts in ischemic hearts and delayed acrolein or 4-hydroxy trans-2-nonenal-induced hypercontracture in isolated cardiac myocytes. Changes in the levels of ATP, high-energy phosphates, intracellular pH, and glycolysis during low-flow ischemia in the wild-type mice hearts were attenuated in the ATPGD1-transgenic hearts. Two natural dipeptide analogs (anserine and balenine) that can either quench aldehydes or buffer intracellular pH, but not both, failed to protect against I/R injury. CONCLUSIONS Either exogenous administration or enhanced endogenous formation of histidyl dipeptides prevents I/R injury by attenuating changes in intracellular pH and preventing the accumulation of lipid peroxidation derived aldehydes.

Decreased ω-6:ω-3 PUFA ratio attenuates ethanol-induced alterations in intestinal homeostasis, microbiota, and liver injury.

Ethanol (EtOH)-induced alterations in intestinal homeostasis lead to multi-system pathologies, including liver injury. ω-6 PUFAs exert pro-inflammatory activity, while ω-3 PUFAs promote anti-inflammatory activity that is mediated, in part, through specialized pro-resolving mediators [e.g., resolvin D1 (RvD1)]. We tested the hypothesis that a decrease in the ω-6:ω-3 PUFA ratio would attenuate EtOH-mediated alterations in the gut-liver axis. ω-3 FA desaturase-1 (fat-1) mice, which endogenously increase ω-3 PUFA levels, were protected against EtOH-mediated downregulation of intestinal tight junction proteins in organoid cultures and in vivo. EtOH- and lipopolysaccharide-induced expression of INF-γ, Il-6, and Cxcl1 was attenuated in fat-1 and WT RvD1-treated mice. RNA-seq of ileum tissue revealed upregulation of several genes involved in cell proliferation, stem cell renewal, and antimicrobial defense (including Alpi and Leap2) in fat-1 versus WT mice fed EtOH. fat-1 mice were also resistant to EtOH-mediated downregulation of genes important for xenobiotic/bile acid detoxification. Further, gut microbiome and plasma metabolomics revealed several changes in fat-1 versus WT mice that may contribute to a reduced inflammatory response. Finally, these data correlated with a significant reduction in liver injury. Our study suggests that ω-3 PUFA enrichment or treatment with resolvins can attenuate the disruption in intestinal homeostasis caused by EtOH consumption and systemic inflammation with a concomitant reduction in liver injury.

Integrating comprehensive two-dimensional gas chromatography mass spectrometry and parallel two-dimensional liquid chromatography mass spectrometry for untargeted metabolomics.

The diverse characteristics and large number of entities make metabolite separation challenging in metabolomics. To date, there is not a singular instrument capable of analyzing all types of metabolites. In order to achieve a better separation for higher peak capacity and accurate metabolite identification and quantification, we integrated GC × GC-MS and parallel 2DLC-MS for analysis of polar metabolites. To test the performance of the developed system, 13 rats were fed different diets to form two animal groups. Polar metabolites extracted from rat livers were analyzed by GC × GC-MS, parallel 2DLC-MS (-) and parallel 2DLC-MS (+), respectively. By integrating all data together, 58 metabolites were detected with significant change in their abundance levels between groups (p≤ 0.05). Of the 58 metabolites, three metabolites were detected in two platforms and two in all three platforms. Manual examination showed that discrepancy of metabolite regulation measured by different platforms was mainly caused by the poor shape of chromatographic peaks resulting from low instrument response. Pathway analysis demonstrated that integrating the results from multiple platforms increased the confidence of metabolic pathway assignment.

Global Plasma Profiling for Colorectal Cancer-Associated Volatile Organic Compounds: a Proof-of-Principle Study.

Volatile organic compounds (VOCs) could reflect changes resulting from ongoing pathophysiological processes and altered body metabolisms, and thus have been studied for various types of cancers. We aimed to test an advanced global metabolomic technique to characterize circulating VOCs in patients diagnosed with colorectal cancer (CRC). We employed solid-phase microextraction (SPME) and comprehensive two-dimensional gas chromatography mass-spectrometry (GC × GC-MS). We analyzed 30 random plasma samples from incident cases of CRC. The 30 samples were from population controls enrolled in a large population-based case-control study. The number of metabolite peaks detected in the cases was significantly lower than that detected in the controls (median 1530 vs. 1694, P = 0.02). Partial least squares-discriminant analysis showed clear VOC profile differences between the CRC and the controls. After adjustment for multiple comparisons at the 5% false discovery rate level, five VOCs were differentially expressed between the cases and the controls. Among these five VOCs, 2,3,4-trimethyl-hexane (decreased) and 2,4-dimethylhept-1-ene (increased) were both lipid peroxidation products but not previously reported for CRC. In summary, this study pointed to an intriguing observation that the richness of volatile metabolites may be reduced in CRC cases and demonstrated the utility of SPME GC × GC-MS in discovery of candidate markers for further validation.

Ethanol and unsaturated dietary fat induce unique patterns of hepatic ω-6 and ω-3 PUFA oxylipins in a mouse model of alcoholic liver disease.

Alcoholic liver disease (ALD), a significant health problem, progresses through the course of several pathologies including steatosis, steatohepatitis, fibrosis, and cirrhosis. There are no effective FDA-approved medications to prevent or treat any stages of ALD, and the mechanisms involved in ALD pathogenesis are not well understood. Bioactive lipid metabolites play a crucial role in numerous pathological conditions, as well as in the induction and resolution of inflammation. Herein, a hepatic lipidomic analysis was performed on a mouse model of ALD with the objective of identifying novel metabolic pathways and lipid mediators associated with alcoholic steatohepatitis, which might be potential novel biomarkers and therapeutic targets for the disease. We found that ethanol and dietary unsaturated, but not saturated, fat caused elevated plasma ALT levels, hepatic steatosis and inflammation. These pathologies were associated with increased levels of bioactive lipid metabolites generally involved in pro-inflammatory responses, including 13-hydroxy-octadecadienoic acid, 9,10- and 12,13-dihydroxy-octadecenoic acids, 5-, 8-, 9-, 11-, 15-hydroxy-eicosatetraenoic acids, and 8,9- and 11,12-dihydroxy-eicosatrienoic acids, in parallel with an increase in pro-resolving mediators, such as lipoxin A4, 18-hydroxy-eicosapentaenoic acid, and 10S,17S-dihydroxy-docosahexaenoic acid. Elucidation of alterations in these lipid metabolites may shed new light into the molecular mechanisms underlying ALD development/progression, and be potential novel therapeutic targets.

Simultaneous quantification of straight-chain and branched-chain short chain fatty acids by gas chromatography mass spectrometry.

Biomedical research in areas such as metabolic disorders, neuromodulatory, and immunomodulatory conditions involves lipid metabolism and demands a reliable and inexpensive method for quantification of short chain fatty acids (SCFAs). We report a GC-MS method for analysis of all straight-chain and branched-chain SCFAs using pentafluorobenzyl bromide (PFBBr) as derivatization reagent. We optimized the derivatization and GC-MS conditions using a mixture containing all eight SCFA standards, i.e., five straight-chain and three branched-chain SCFAs. The optimal derivatization conditions were derivatization time 90 min, temperature 60 °C, pH 7, and (CH3)2CO:H2O ratio 2:1 (v:v). Comparing the performance of different GC column configurations, a 30 m DB-225ms hyphenated with a 30 m DB-5ms column in tandem showed the best separation of SCFAs. Using the optimized experiment conditions, we simultaneously detected all SCFAs with much improved detection limit, 0.244-0.977 µM. We further applied the developed method to measure the SCFAs in mouse feces and all SCFAs were successfully quantified. The recovery rates of the eight SCFAs ranged from 55.7% to 97.9%.

Surface fitting for calculating the second dimension retention index in comprehensive two-dimensional gas chromatography mass spectrometry.

Comprehensive two-dimensional gas chromatography mass spectrometry (GC × GC-MS) has been widely used for analysis of volatile compounds. However, the second dimension retention index (I) of each compound is not widely used to aid compound identification owing to the limited accuracy of I calculation. We report a surface fitting approach to the calculation of I using n-alkanes (C7-C30) as references, where the second dimension retention time (2tR) and the second dimension column temperature (2Te) formed the X-Y plane and the I was the Z-axis to form the I surface. Compared to the conventional approach for calculating I using isovolatility curves, the surface fitting approach eliminated the construction of isovolatility curves for the reference compounds and gives better reproducibility. The goodness of the proposed surface fitting achieved R2 = 0.9999 and RMSE = 6.1 retention index units (iu). Ten-fold cross validation demonstrated the surface fitting approach had a good predictability with average R2 = 0.9999 and RMSE = 6.6 iu. The developed method was also applied to calculate the second dimension retention indices of compound standards in two commercial mixtures MegaMix A and MegaMix B. The mean standard deviation of the calculated I was only 1.6 iu for compounds in MegaMix A and 3.4 iu for compounds in MegaMix B. Compared with the literature results, the small value of standard deviation in the calculated retention index using surface fitting method shows that the surface fitting method has less measurement variability than the conventional isovolatility curve approach.