Emergency diagnosis made easy: matrix removal and analyte enrichment from raw saliva using paper-arrow mass spectrometry.

Paracetamol overdose is a leading cause of acute liver failure that can prove fatal. Establishing paracetamol concentration accurately and quickly is critical. Current detection methods are invasive, time-consuming and/or expensive. Non-invasive, rapid and cost-effective techniques are urgently required. To address this challenge, a novel approach, called Paper-Arrow Mass Spectrometry (PA-MS) has been developed. This technique combines sample collection, extraction, enrichment, separation and ionisation onto a single paper strip, and the entire analysis process, from sample to result, can be carried out in less than 10 min requiring only 2 µL of raw human saliva. PA-MS achieved a LOQ of 185 ng mL-1, mean recovery of 107 ± 7%, mean accuracy of 11 ± 8% and precision ≤5% using four concentrations, and had excellent linearity (r2 = 0.9988) in the range of 0.2-200 µg mL-1 covering the treatment concentration range, surpassing the best-in-class methods currently available for paracetamol analysis. Furthermore, from a panel of human saliva samples, inter-individual variability was found to be <10% using this approach. This technique represents a promising tool for rapid and accurate emergency diagnosis.

Rapid differentiation of cystic fibrosis-related bacteria via reagentless atmospheric pressure photoionisation mass spectrometry.

Breath analysis is an area of significant interest in medical research as it allows for non-invasive sampling with exceptional potential for disease monitoring and diagnosis. Volatile organic compounds (VOCs) found in breath can offer critical insight into a person's lifestyle and/or disease/health state. To this end, the development of a rapid, sensitive, cost-effective and potentially portable method for the detection of key compounds in breath would mark a significant advancement. Herein, we have designed, built and tested a novel reagent-less atmospheric pressure photoionisation (APPI) source, coupled with mass spectrometry (MS), utilising a bespoke bias electrode within a custom 3D printed sampling chamber for direct analysis of VOCs. Optimal APPI-MS conditions were identified, including bias voltage, cone voltage and vaporisation temperature. Calibration curves were produced for ethanol, acetone, 2-butanone, ethyl acetate and eucalyptol, yielding R2 > 0.99 and limits of detection < 10 pg. As a pre-clinical proof of concept, this method was applied to bacterial headspace samples of Escherichia coli (EC), Pseudomonas aeruginosa (PSA) and Staphylococcus aureus (SA) collected in 1 L Tedlar bags. In particular, PSA and SA are commonly associated with lung infection in cystic fibrosis patients. The headspace samples were classified using principal component analysis with 86.9% of the total variance across the first three components and yielding 100% classification in a blind-sample study. All experiments conducted with the novel APPI arrangement were carried out directly in real-time with low-resolution MS, which opens up exciting possibilities in the future for on-site (e.g., in the clinic) analysis with a portable system.

Insights into serum metabolic biomarkers for early detection of incident diabetic kidney disease in Chinese patients with type 2 diabetes by random forest.

Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease (ESRD) worldwide. Early detection is critical for the risk stratification and early intervention of progressive DKD. Serum creatinine (sCr) and urine output are used to assess kidney function, but these markers are limited by their delayed changes following kidney pathology, and lacking of both sensitivity and accuracy. Hence, it is essential to illustrate potential diagnostic indicators to enhance the precise prediction of early DKD. A total of 194 Chinese individuals include 30 healthy participants (Stage 0) and 164 incidents with type 2 diabetes (T2D) spanning from DKD's Stage 1a to 4 were recruited and their serums were subjected for untargeted metabolomic analysis. Random forest (RF), a machine learning approach, together with univariate linear regression (ULR) and multivariate linear regression (MvLR) analysis were applied to characterize the features of untargeted metabolites of DKD patients and to identify candidate DKD biomarkers. Our results indicate that 2-(α-D-mannopyranosyl)-L-tryptophan (ADT), succinyladenosine (SAdo), pseudouridine and N,N,N-trimethyl-L-alanyl-L-proline betaine (L-L-TMAP) were associated with the development of DKD, in particular, the latter three that were significantly elevated in Stage 2-4 T2D incidents. Each of the four metabolites in combination with sCr achieves better performance than sCr alone with area under the receiver operating characteristic curve (AUC) of 0.81-0.91 in predicting DKD stages. An average of 3.9 years follow-up study of another cohort including 106 Stage 2-3 patients suggested that "urinary albumin-to-creatinine ratio (UACR) + ADT + SAdo" can be utilized for better prognosis evaluation of early DKD (average AUC = 0.9502) than UACR without sexual difference.

Attaching protein-adsorbing silica particles to the surface of cotton substrates for bioaerosol capture including SARS-CoV-2.

The novel coronavirus pandemic (COVID-19) has necessitated a global increase in the use of face masks to limit the airborne spread of the virus. The global demand for personal protective equipment has at times led to shortages of face masks for the public, therefore makeshift masks have become commonplace. The severe acute respiratory syndrome caused by coronavirus-2 (SARS-CoV-2) has a spherical particle size of ~97 nm. However, the airborne transmission of this virus requires the expulsion of droplets, typically ~0.6-500 µm in diameter (by coughing, sneezing, breathing, and talking). In this paper, we propose a face covering that has been designed to effectively capture SARS-CoV-2 whilst providing uncompromised comfort and breathability for the wearer. Herein, we describe a material approach that uses amorphous silica microspheres attached to cotton fibres to capture bioaerosols, including SARS CoV-2. This has been demonstrated for the capture of aerosolised proteins (cytochrome c, myoglobin, ubiquitin, bovine serum albumin) and aerosolised inactivated SARS CoV-2, showing average filtration efficiencies of ~93% with minimal impact on breathability.

Assessment of creatinine concentration in whole blood spheroids using paper spray ionization-tandem mass spectrometry.

Accurate quantification of blood creatinine is important to estimate the glomerular filtration rate. Existing techniques using liquid chromatography tandem mass spectrometry (LC-MS/MS) have a high accuracy and eliminate most interferences encountered in routine enzymatic and Jaffé methods. However, they require laborious and time-consuming sample treatment and data acquisition. The aim of this study is to develop a fast and simple method to enable a direct analysis of whole blood creatinine with performance measures that are comparable to conventional LC-MS/MS. 5µL whole blood is formed as a three-dimensional spheroid on hydrophobic silanized paper substrates which then undergoes paper-spray ionization-tandem mass spectrometry (PSI-MS/MS). The method is validated using real human samples and compared with LC-MS/MS. PSI-MS/MS whole blood analysis exhibited a lower limit of quantification of 2.5 µg/mL, precision ≤ 6.3%, recovery in the range of 88-94% and excellent linearity (R2 > 0.99; 2.5-20 µg/mL) covering the normal range for creatinine levels. Creatinine levels were comparable to those measured by LC-MS/MS with small deviations of less than 0.3 µg/mL. This simple, fast and accurate microsampling technique for direct analysis of creatinine from whole blood shows promise for routine clinical screening and monitoring. This approach can be readily extended for other analytes of interest and, due to inherent advantages relating to cost, storability, speed, and simplicity, it can be especially advantageous for use in resource-limited settings.

Lactobacillus casei Strain Shirota Ameliorates Dextran Sulfate Sodium-Induced Colitis in Mice by Increasing Taurine-Conjugated Bile Acids and Inhibiting NF-κB Signaling via Stabilization of IκBα.

Inflammatory bowel disease (IBD) is a chronic progressive intestinal inflammatory disease, characterized by an altered gut microbiota composition and accompanying alterations in circulatory bile acids. Increasing evidence supports the beneficial effect of probiotics intake on health. Introduction of probiotics to the intestines can modulate gut microbiota composition and in turn regulate the host immune system and modify the inflammatory response. Probiotics can also improve intestinal barrier function and exhibit a positive impact on host physiological and pathological conditions via gut microbiota-derived metabolites. Previous studies have demonstrated that Lactobacillus casei strain Shirota (LcS) treatment could inhibit clinical manifestation of colitis in dextran sulfate sodium (DSS)-induced mice, however, the underlying mechanisms remain unknown. In this study, we employed the DSS-induced acute colitis mouse model to investigate the anti-inflammatory effects of LcS and related mechanisms. Administration of LcS ameliorated the severity of DSS-induced colitis and enhanced intestinal integrity via induction of mucin-2 and occludin expression in colons. Fecal microbiota analysis showed that LcS increased the relative abundance of beneficial bacterial species in colitic mice, whereas the relative abundance of pathobionts was reduced. Additionally, LcS treatment modulated circulating bile acid profiles in colitic mice. In mice treated with LcS, we identified increased levels of primary taurine-conjugated bile acids, including taurocholic acid (TCA) and taurochenodeoxycholic acid (TCDCA). LcS treatment also increased the levels of secondary taurine-conjugated bile acids, including taurodeoxycholic acid (TDCA) and tauroursodeoxycholic acid (TUDCA). Moreover, LcS treatment exhibited a suppressive effect on the hydroxylated primary bile acids α-muricholic acid (α-MCA) and ß-muricholic acid (ß-MCA). We further demonstrated that LcS treatment suppressed the expression of pro-inflammatory mediators interferon-gamma (IFN-γ) and nitric oxide (NO), and increased the expression of the anti-inflammatory mediator interleukin-10 (IL-10) in colon tissues, potentially as a result of altered bile acid profiles. Mechanistically, we showed that LcS treatment suppressed the activation of nuclear factor-kappa B (NF-κB) signaling via stabilization of inhibitor of NF-κB alpha (IκBα). Altogether, we have demonstrated the therapeutic effects of LcS in DSS-induced colitis, providing new insights into its effect on bile acid metabolism and the related anti-inflammatory mechanisms. Our findings provide support for the application of LcS in the treatment of IBD.

HBx-K130M/V131I Promotes Liver Cancer in Transgenic Mice via AKT/FOXO1 Signaling Pathway and Arachidonic Acid Metabolism.

Chronic hepatitis B viral (HBV) infection remains a high underlying cause for hepatocellular carcinoma (HCC) worldwide, while the genetic mechanisms behind this remain unclear. This study elucidated the mechanisms contributing to tumor development induced by the HBV X (HBx) gene of predominantly Asian genotype B HBV and its common HBx variants. To compare the potential tumorigenic effects of K130M/V131I (Mut) and wild-type (WT) HBx on HCC, the Sleeping Beauty (SB) transposon system was used to deliver HBx Mut and WT into the livers of fumarylacetoacetate hydrolase (Fah)-deficient mice and in the context of transformation related protein 53 (Trp53) deficiency. From our results, HBx Mut had a stronger tumorigenic effect than its WT variant. Also, inflammation, necrosis, and fibrosis were evident in HBx experimental animals. Reduction of forkhead box O1 (FOXO1) with increased phosphorylation of upstream serine/threonine kinase (AKT) was detected under HBx Mut overexpression. Thus, it is proposed that HBx Mut enhances disease progression by reducing FOXO1 via phosphorylation of AKT. At the metabolomic level, HBx altered the expression of genes that participated in arachidonic acid (AA) metabolism, as a result of inflammation via accumulation of proinflammatory factors such as prostaglandins and leukotriene in liver. Taken together, the increased rate of HCC observed in chronic hepatitis B patients with K130M/V131I-mutated X protein, may be due to changes in AA metabolism and AKT/FOXO1 signaling. IMPLICATIONS: Our findings suggested that HBx-K130M/V131I-mutant variant promoted HCC progression by activating AKT/FOXO1 pathway and inducing stronger inflammation in liver via AA metabolism.

A Metabolomics Study on the Bone Protective Effects of a Lignan-Rich Fraction From Sambucus Williamsii Ramulus in Aged Rats.

The lignan-rich fraction (SWR) of Sambucus Williamsii Ramulus, a folk herbal medicine in China for treatment of bone diseases, has previously reported to exert protective effects on bone without exerting uterotrophic effects in ovariectomized (OVX) mice. The aim of the present study was to identify the potential metabolites and the associated metabolic pathways that contribute to the beneficial effects of SWR on bone in vivo. Aged female Sprague Dawley rats (9 months old) were either sham-operated or ovariectomized for 12 weeks, before receiving treatment for another 12 weeks with the following treatment groups (n = 12 each): vehicle (Sham), vehicle (OVX), Premarin (130 µg/kg) or low (57 mg/kg), medium (114 mg/kg), and high (228 mg/kg) doses of SWR. The results showed that SWRH significantly suppressed bone loss, improved bone micro-architecture and increased bone strength on tibia without stimulating uterus weight gain in OVX rats. Premarin exerted similar bone protective effects as SWRH but elicited uterotrophic effects in OVX rats. The metabolic profiles of serum samples were analyzed by using ultra-performance liquid chromatography quadrupole time-of flight mass spectrometry and gas chromatography time-of flight mass spectrometry, and the metabolites that were significantly altered were identified by multivariate statistical analysis. Our study indicated that SWRH effectively restored the changes of 26 metabolites induced by estrogen-deficiency in OVX rats, which related to lipids, amino acids, tryptophan metabolisms, and anti-oxidative system. A subsequent validation showed that the serum level of superoxide dismutase and catalase were indeed up-regulated, while the serotonin level in a tryptophan hydroxylase 1 (TPH1) high expressing cells (rats RBL-2H3 cells) was down regulated after treatment with SWR. The results also suggested that the gut-microbiota may play an important role on the bone protective effects of SWR. The current study provides insight for understanding the unique mechanism of actions of SWR that might be involved in achieving bone protective effects in vivo.

Clinical and histopathological features of chronic hepatitis B virus infected patients with high HBV-DNA viral load and normal alanine aminotransferase level: A multicentre-based study in China.

BACKGROUND: The aim of this study is to reveal the clinical and histopathological features of HBsAg-positive and HBeAg-positive chronic hepatitis B infected patients with high level of HBV DNA, from 17 hospitals and medical centres in China, with alanine aminotransferase levels within the lower region of normal range versus those with levels within the upper region of normal range and to investigate the clinical risk factors for the requirement of treatment through the examination of liver biopsy. METHODS: Liver biopsy was performed on high level of HBV DNA of 455 patients with HBsAg-positive and HBeAg-positive chronic hepatitis B infection and persistently normal alanine aminotransferase level. Liver necroinflammation and fibrosis were graded per the Knodell histological activity index and Ishak's fibrosis score, respectively. Univariate analysis of the clinical parameters versus necroinflammation and fibrosis was carried out. RESULTS: Of the subjects in this multicentre-based study, 5.49% and 10.11% had significant necroinflammation with Knodell histological activity index ≥ 9 and hepatic fibrosis stages with Ishak scores ≥ 3, respectively. The subjects were stratified into three age groups (30-39, 40-49 and ≥ 50 years), and our data clearly suggested that age, particularly in the age group over 50, was an independent predictor of liver necroinflammation and fibrosis. Lower HBV-DNA viral levels were found in patients with Knodell histological activity index ≥ 9 or advanced fibrosis (Ishak scores ≥ 3). CONCLUSION: Our results showed that histological changes in liver tissues were observed in a significant proportion of patients with persistently normal alanine aminotransferase level. According to the data evaluation results, liver biopsy is advisable for HBeAg-positive chronic hepatitis B infected patients aged older than 40 and high HBV-DNA viral load in China.

Chemical Analysis of Astragali Complanati Semen and Its Hypocholesterolemic Effect Using Serum Metabolomics Based on Gas Chromatography-Mass Spectrometry.

The hypocholesterolemic protective effect of the dried seed of Astragalus complanatus (ACS) was investigated in rats fed with normal diet, high cholesterol diet (HCD), and HCD plus 70% ethanol extract of ACS (600 mg/kg/day) by oral gavage for four weeks. ACS extract was tested to be rich in antioxidants, which may be contributed to its high content of phenolic compounds. Consumption of ACS remarkably suppressed the elevated total cholesterol (p < 0.01) and LDL-C (p < 0.001) induced by HCD. Chemical constituents of ACS extract were analyzed by ultra-performance liquid chromatography coupled with electrospray ionization orbitrap mass spectrometry and the results showed that the ACS extract mainly consisted of phenolic compounds including flavonoids and flavonoid glycosides. In addition, based on the serum fatty acid profiles, elucidated using gas chromatography-mass spectrometry, free and esterified fatty acids including docosapentaenoic acid, adrenic acid, dihomo-γ-linolenic acid and arachidonic acid were regulated in ACS treatment group. Western blot results further indicated the protein expression of peroxisome proliferator-activated receptor alpha (PPARα) (p < 0.05) in liver was upregulated in ACS treatment group. To conclude, our results clearly demonstrated that ACS provides beneficial effect on lowering HCD associated detrimental change.

A review on the traditional Chinese medicinal herbs and formulae with hypolipidemic effect.

Hyperlipidemia, characterized by the abnormal blood lipid profiles, is one of the dominant factors of many chronic diseases such as diabetes, obesity, and cardiovascular diseases (CVD). For the low cost, effectiveness, and fewer side effects, the popularity of using traditional Chinese medicine (TCM) to handle hyperlipidemia is increasing and its role in health care has been recognized by the public at large. Despite the importance of TCM herbs and formulations, there is no comprehensive review summarizing their scientific findings on handling hyperlipidemia. This review summarizes the recent experimental and clinical results of nine representative single Chinese herbs and seven classic TCM formulae that could improve lipid profiles so as to help understand and compare their underlying mechanisms. Most of single herbs and formulae demonstrated the improvement of hyperlipidemic conditions with multiple and diverse mechanisms of actions similar to conventional Western drugs in spite of their mild side effects. Due to increasing popularity of TCM, more extensive, well-designed preclinical and clinical trials on the potential synergistic and adverse side effects of herb-drug interactions as well as their mechanisms are warranted. Hyperlipidemic patients should be warned about the potential risks of herb-drug interactions, particularly those taking anticoagulants and antiplatelet drugs.

Semen Astragali Complanati: an ethnopharmacological, phytochemical and pharmacological review.

ETHNOPHARMACOLOGICAL RELEVANCE: Semen Astragali Complanati (SAC), the dried ripe seed of Flatstem Milkvetch (Astragalus complanatus Bunge) (Leguminosae), is commonly used in traditional Chinese medicine (TCM) for treating muscle, liver, kidney, blood, skin and reproductive system diseases. MATERIALS AND METHODS: Relevant information about SAC was gathered via "Google Scholar", "ISI Web of Knowledge", "PubMed", "ScienceDirect", "Medline Plus", "ACS", "CNKI" and "Wiley Online Library" and from books in local libraries. RESULTS: The major contents of SAC include fatty acids, amino acids, polysaccharides, flavonoids, triterpene glycosides and trace elements. Previous scientific studies have reported that SAC exhibits a number of therapeutic effects on chronic diseases such as cardiovascular diseases, diabetes mellitus and cancers. It has been found that flavonoids are the main bioactive component in SAC. However most of the previous studies have shown the effects brought by the total flavonoid fraction extracted from SAC only; further studies are warranted for the biological effects produced by individual components. There are only a few studies on the toxicity of SAC and the overall results show that its toxicity is quite low or even non-existent. CONCLUSIONS: SAC is a valuable TCM herb with multiple pharmacological effects for treating some chronic diseases. More studies on SAC will help us to have a better understanding of its pharmacological mechanisms so as to provide more scientific evidence to explain its traditional uses, identify its therapeutic potential on other diseases and understand its possible harmful effects. Based on previous studies, it is easy to identify that antioxidant effect of SAC might play an important role on its pharmacological effects. Studying the effects of SAC on handling intracellular reactive oxygen species may be a potential direction to help understanding the molecular mechanisms of SAC on preventing and/or treating chronic diseases.

A review of the phytochemistry and pharmacological activities of raphani semen.

The dried ripe seed of Raphanus sativus L., commonly known as radish seed (or Raphani Semen), is used as traditional Chinese medicine (TCM) to treat constipation, chronic tracheitis, and hypertension. The major active compounds in Raphani Semen are alkaloids, glucosinolates, brassinosteroids, and flavonoids. Fatty acids are its main nutritional contents. Raphani Semen has been demonstrated to have beneficial effects on hypertension, obesity, diabetes mellitus, constipation, and cough. So far, there is no report about the adverse/toxic effects of this herb on humans. However, Raphani Semen processed by roasting was reported to exhibit some adverse effects on mice. Additionally, erucic acid, the main fatty acid in Raphani Semen, was shown to enhance the toxicity of doxorubicin. Thus, Raphani Semen has a potential risk of causing toxicity and drug interaction. In summary, Raphani Semen is a valuable TCM herb with multiple pharmacological effects. More studies on Raphani Semen could help better understand its pharmacological mechanisms so as to provide clear scientific evidence to explain its traditional uses, to identify its therapeutic potential on other diseases, and to understand its possible harmful effects.