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Objective:To establish an isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) method for the determination of L-tryptophan and its metabolites in serum.Methods:The methodology was established and evaluated using serum samples collected from 166 healthy subjects undergoing physical examinations at West China Hospital from November 2022 to January 2023 were collected. Isotope-labeled markers of L-tryptophan (Trp), L-kynurenine (Kyn), and kynurenic acid (KA) were used as internal standards. After protein precipitation treatment of serum samples, LC-MS/MS was used to determine Trp, Kyn, and KA simultaneously. The selectivity, specificity, linearity, detection limit (LOD), quantification limit (LOQ), carry-over, precision, recovery rate, matrix effect, and dilution integrity of the method were evaluated.Results:The linearity of Trp, Kyn, and KA was demonstrated to be 0.999. The LODs were 0.10 μmol/L, 0.01 μmol/L and 1.00 nmol/L, respectively. The LOQs were 0.20 μmol/L, 0.04 μmol/L and 2.00 nmol/L, respectively. The intra-batch precision and inter-batch precision were below<10%. The average recovery rate and the relative matrix effect were all about 100%. The samples over the upper limit of quantitation can be diluted up to 16 times. The Trp concentration, Kyn concentration, KA concentration, Kyn/Trp ratio, and KA/Kyn ratio in serum of healthy subjects were 59.55±10.92 μmol/L, 1.85±0.43 μmol/L, 39.89±17.93 nmol/L, (31.64±8.19)×10 -3 and 21.51±6.72, respectively. Conclusion:An ID-LC-MS/MS method was successfully established for the quantitative determination of Trp, Kyn, and KA in serum. The method proved to be simple, rapid, sensitive, accurate, and reliable, providing robust support for clinical research related to these analytes.
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Objective:To correlate bone mineral density with serum bone metabolism indexes in patients with hyperthyroidism.Methods:Thirty patients with hyperthyroidism who received treatment in the General Hospital of Taiyuan Iron and Steel (Group) Co., Ltd. from January 2018 to August 2019 were included in the hyperthyroidism group. Additional 30 healthy subjects who concurrently received routine physical examination were included in the control group. Bone mineral density in all subjects was measured by dual energy X-ray absorptiometry. Bone metabolism indexes in all subjects were measured using a Roche chemiluminescence instrument: 25-hydroxyvitamin D level [25(OH)D], aminoterminal propeptide of type I procollagen (PINP) and beta-cardiotoxin (β-CTX). Correlation between bone mineral density and serum bone metabolism indexes was analyzed using Spearman method.Results:Bone mineral density in lumbar vertebrae 1-4 [(0.86 ± 0.14) g/cm 3], left femoral neck [(0.79 ± 0.07) g/cm 3] and left hip joint [(0.72 ± 0.10) g/cm 3] in the hyperthyroidism group was significantly lower than that in the control group [(1.28 ± 0.21) g/cm 3, (1.03 ± 0.18) g/cm 3, (0.86 ± 0.13) g/cm 3, t = 9.115, 6.806, 4.675, all P < 0.001]. There were 6 cases of osteoporosis, 12 cases of osteopenia and 12 cases of normal bone in the hyperthyroidism group. There was 1 case of osteoporosis, 6 cases of osteopenia and 23 cases of normal bone in the control group. There was significant difference in the number of cases developing osteoporosis between hyperthyroidism and control groups ( Z = 2.968, P < 0.05). Serum level of 25(OH)D in the hyperthyroidism group was significantly lower than that in the control group [(16.89 ± 4.31) μg/L vs. (24.13 ± 5.48) μg/L, t = 5.688, P < 0.001]. Serum levels of PINP and β-CTX in the hyperthyroidism group were significantly lower than those in the control group [PINP: (49.37 ± 10.23) μg/L vs. (47.68 ± 6.49) μg/L; β-CTX: (774.56 ± 159.67) ng/L vs. (534.32 ± 167.48) ng/L, t = 45.974 and 5.687, both P < 0.001]. In the hyperthyroidism group, bone mineral density at lumbar vertebrae 1-4, left femoral neck and left hip joint was positively correlated with serum level of 25(OH)D ( r = 0.417, 0.396, 0.401, all P < 0.05), and it was negatively correlated with serum levels of PINP and β-CTX ( r = -0.414, -0.399, -0.432, -0.404, -0.387, -0.412, all P < 0.05). Conclusion:Hyperthyroidism patients generally have low bone mineral density and accelerated bone metabolism. It is of great significance to regularly monitor bone mineral density and serum bone metabolism indexes in hyperthyroidism patients to prevent osteoporosis.
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BACKGROUNDThe outbreak of COVID-19 caused by a novel Coronavirus (termed SARS-CoV-2) has spread to over 140 countries around the world. Currently, reverse transcription quantitative qPCR (RT-qPCR) is used as the gold standard for diagnostics of SARS-CoV-2. However, the positive rate of RT-qPCR assay of pharyngeal swab samples are reported to vary from 30[~]60%. More accurate and sensitive methods are urgently needed to support the quality assurance of the RT-qPCR or as an alternative diagnostic approach. METHODSWe established a reverse transcription digital PCR (RT-dPCR) protocol to detect SARS-CoV-2 on 194 clinical pharyngeal swab samples, including 103 suspected patients, 75 close contacts and 16 supposed convalescents. RESULTSThe limit of blanks (LoBs) of the RT-dPCR assays were [~]1.6, [~]1.6 and [~]0.8 copies/reaction for ORF 1ab, N and E genes, respectively. The limit of detection (LoD) was 2 copies/reaction. For the 103 fever suspected patients, the sensitivity of SARS-CoV-2 detection was significantly improved from 28.2% by RT-qPCR to 87.4% by RT-dPCR. For close contacts, the suspect rate was greatly decreased from 21% down to 1%. The overall sensitivity, specificity and diagnostic accuracy of RT-dPCR were 90%, 100% and 93 %, respectively. In addition, quantification of the viral load for convalescents by RT-dPCR showed that a longer observation period was needed in the hospital for elderly patients. CONCLUSIONRT-dPCR could be a confirmatory method for suspected patients diagnosed by RT-qPCR. Furthermore, RT-dPCR was more sensitive and suitable for low viral load specimens from the both patients under isolation and those under observation who may not be exhibiting clinical symptoms.
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The age of 18 is an important criterion in judicial trial, immigrant and competitive sports. Consequentially, the estimation of age 18 is a key issue in forensic practice and research. The extremitas sternalis claviculae, iliac crest, third molar, and the proximal limb of the limb bone were usually used as indictors of age 18. The results of the previous studies demonstrated that those indictors could be beneficial to the estimation of age 18. The X-ray, CT, ultrasound and MRI of different indictors were widely utilized for the estimation of age 18, particularly the thin-layer CT. But due to the non-radiation, MRI will be a trend for forensic age estimation in the future. Whilst in the previous studies, the descriptive analysis was applied for the estimation of age 18, but due to the low statistic efficiency, it is unsuitable for forensic age estimation, and the future studies should pay attention to the high efficiency statistical methods, for instance, the ROC curve or the data mining.
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The low mass cutoff ( LMCO) is the main weakness of ion trap when it performs tandem mass analysis by collision induced dissociation (CID). LMCO means that some daughter ions of m/ z are less than about 1 / 3 of the m/ z of parent ion could not be detected during the tandem mass spectrometry processing. A new method which can significantly improve the effect of low mass cutoff was proposed and investigated. By simply changing the scan method of digital potential frequency, some low mass ions can be effectively observed during the tandem mass spectrometric experiment. In the experiment, the frequency of the digital ion trapping power and ion activation power were scanned from lower value to higher value, and some lower mass product ions could be detected during CID process. For example, some lower mass ions were observed during the CID of reserpine precursor ion when the frequency of its digital trapping power was scanned from 500 kHz to 560 kHz. The tandem mass spectra of Reserpine ion showed that the experimental results both from this work and the triple quadrupole mass spectrometer were exactly the same.
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A new method has been developed for the determination of selected polybrominated diphenylethers (PBDEs) and polybrominated biphenyl (PBB) in four polymers: high-density polyethylene (HDPE), polystyrene (PS), acrylonitrile-butadiene-styrene copolymer (ABS), and polypropylene (PP). PBDEs and PBB in the polymers were extracted with toluene, using ultrasonic-assisted extraction (UAE). The extracts were then determined by high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS), using external calibration (single-point). Extraction parameters of UAE and several ICP-MS parameters were optimized. Extraction efficiencies almost reached 100%. The relative standard deviations (RSDs) were in the range of 0.7%-5.4%. The results demonstrate that the method possesses advantages of good precision, as well as high extraction efficiency and accuracy. The method especially overcomes the problem of the thermal degradation of highly brominated PBDEs, such as PBDE-209.
