Comparison of two Analytical Methods (ELISA and LC-MS/MS) for Determination of Aflatoxin B1 in Corn and Aflatoxin M1 in Milk

The aim of this paper is to assess the closeness of agreement between results of ELISA and LC-MS/MS methods for determination of aflatoxin B₁ in corn and aflatoxin M₁ in milk. Samples of corn (n=100) and milk (n=250) were simultaneously analyzed using ELISA and LC-MS/MS methods, after the severe drought that affected Serbia in summer 2012 resulting in occurrence of aflatoxin B1 in corn and aflatoxin M₁ in milk. Regression analysis showed higher level of agreement between aflatoxin B₁ samples (R²=0.994), compared to aflatoxin M₁ samples (R²=0.920). However, both techniques were satisfactory in meeting the requirements for official control purposes.     

Characterization of the TAG of peanut oil by electrospray LC-MS-MS

A study of processed peanut oil was undertaken to assess the utility of HPLC combined with tandem MS to obtain data easily regarding the number of TAG of fats and oils and their FA composition. Mass chromatograms and spectra corresponding to only TAG of a single M.W. were obtained for the full range of TAG in the sample. Analysis of the mass spectra allowed the identification of more than 160 TAG in the sample by their FA composition. In addition, it was possible to estimate relative abundances of the TAG and suggest the position of the FA on glycerol for a limited number of cases. This technique greatly simplifies the task of assigning FA to coeluting TAG and facilitates identification of TAG present in trace quantities in mixtures, with possible application in circumstances where such trace TAG could be significant markers. Results are quickly obtained without extensive sample preparation or prefractionation of the sample.     

Rapid and Sensitive Liquid Chromatography-Tandem Mass Spectrometry for Quantification of Glycyrrhetic Acid in Human Plasma

A simple, rapid and sensitive liquid chromatography-tandem mass spectrometry(LC-MS/MS) for the determination of glycyrrhetic acid in human plasma with ginsenoside Rh2 as internal standard was developed and validated. The plasma samples were prepared via liquid-liquid extraction with ethyl acetate. Chromatographic separation was accomplished on a Venusil MP-C18(50 mm×2.1 mm, 5μm i.d.) column at 25℃. The mobile phase consisted of acetonitrile/5 mmol·L-1 ammonium acetate(10:90, volume ratio) at a flow rate of 0.4 mL/min. Negative electrospray ionization was utilized as the ionization source. Glycyrrhetic acid and internal standard were determined via the mutiple reaction monitoring of precursor→production ion transitions at m/z 469→425, 409 and m/z 621→161,respectively. Each sample was chromatographed within 2.5 min. The lower limit of uantification was 0.50 ng/mL for 200 μL of plasma sample and the linear range was from 0.50 ng/mL to 800 ng/mL. The intra-and inter-day precisions were less than 8.76% in terms of relative standard deviation(RSD), and the accuracy was within a range of -3.25%-1.32% in terms of relative error(RE). The method was successfully applied to the pharmacokinetic studies of glycyrrhetic acid in healthy male Chinese volunteers after a single oral administration of 75 mg of glycyrrhizin.     

A Simple and Efficient Method for the Substrate Identification of Amino Acid Decarboxylases

Amino acid decarboxylases convert amino acids into different biogenic amines which regulate diverse biological processes. Therefore, identifying the substrates of amino acid decarboxylases is critical for investigating the function of the decarboxylases, especially for the new genes predicted to be amino acid decarboxylases. In the present work, we have established a simple and efficient method to identify the substrates and enzymatic activity of amino acid decarboxylases based on LC-MS methods. We chose GAD65 and AADC as models to validate our method. GAD65 and AADC were expressed in HEK 293T cells and purified through immunoprecipitation. The purified amino acid decarboxylases were subjected to enzymatic reaction with different substrate mixtures in vitro. LC-MS analysis of the reaction mixture identified depleted or accumulated metabolites, which corresponded to candidate enzyme substrates and products, respectively. Our method successfully identified the substrates and products of known amino acid decarboxylases. In summary, our method can efficiently identify the substrates and products of amino acid decarboxylases, which will facilitate future amino acid decarboxylase studies.     

Cytoskeleton Elements Contribute to Prion Peptide-Induced Endothelial Barrier Breakdown in a Blood–Brain Barrier In Vitro System

The mechanisms involved in the interaction of PrP 106-126, a peptide corresponding to the prion protein amyloidogenic region, with the blood–brain barrier (BBB) were studied. PrP 106-126 treatment that was previously shown to impair BBB function, reduced cAMP levels in cultured brain endothelial cells, increased nitric oxide (NO) levels, and changed the activation mode of the small GTPases Rac1 (inactivation) and RhoA (activation). The latter are well established regulators of endothelial barrier properties that act via cytoskeletal elements. Indeed, liquid chromatography-mass spectrometry (LC-MS)-based proteomic profiling study revealed extensive changes in expression of cytoskeleton-related proteins. These results shed light on the nature of the interaction between the prion peptide PrP 106-126 and the BBB and emphasize the importance of the cytoskeleton in endothelium response to prion- induced stress.     

Vitamin D as a Possible COVID-19 Prevention Strategy

Vitamin D is no longer considered an agent only affecting calcium phosphate metabolism. A number of studies over the past few years have demonstrated its role in immunomodulation and its influence on the development and functioning of the brain and nervous system. In the current epidemiological crisis caused by coronavirus disease 2019 (COVID-19), the immunoprotective role of vitamin D has been discussed by some authors regarding whether it contributes to protection against this serious disease or whether its use does not play a role. Non-standard approaches taken by laboratories in examining the serum levels of the vitamin D metabolite calcidiol have contributed to inconsistent results. We examined the serum of 60 volunteers in the spring and autumn of 2021 who declared whether they were taking vitamin D at the time of sampling. Furthermore, the tested participants noted whether they had experienced COVID-19. A newly developed liquid chromatography–tandem mass spectrometry (LC-MS/MS) method was used to measure calcidiol levels. The analysis of variance (ANOVA) model of Statgraphics Centurion 18 statistical software from Statgraphics Technologies was used for calculations. The results of this study showed that those who took vitamin D suffered significantly less often from COVID-19 than those who did not take vitamin D.     

油菜蜂花粉黄酮体外降糖活性研究

以总黄酮含量为考察指标,利用溶剂萃取和大孔树脂对油菜蜂花粉乙醇提取物进行分离纯化,富集黄酮,然后对不同极性组分进行抑制α-葡萄糖苷酶实验,并利用红外光谱（IR）和液-质联用（LC-MS）对体外降糖活性最高的组分进行化学成分分析。结果表明,黄酮类物质在抑制α-葡萄糖苷酶活性中起主要作用;AB-8大孔树脂纯化得到的PEFS-3组分总黄酮含量为68.77%,IC50为72.16μg/m L,远小于阿卡波糖的IC50（1124.86μg/m L）,表明其抑制效果强于阿卡波糖;PEFS-3组分中共鉴定出5种主要物质,其中4种为槲皮素-3-O-β-D-吡喃葡萄糖基-（1→2）-O-β-D-吡喃葡萄糖苷、山奈酚-3,4’-双-O-β-D-吡喃葡萄糖苷、异鼠李素-3-O-β-D-吡喃葡萄糖基-（1→2）-β-D-吡喃葡萄糖苷、山奈酚-3-O-β-D-吡喃葡萄糖基-（1→2）-β-D-吡喃葡萄糖苷,第5种推断为亥茅酚苷或黄烷醇,具体结果还需进一步研究。      

蜂花粉中黄酮类化合物的鉴定和含量测定

以24种不同蜂花粉为材料,对其中微量特征成分黄酮类化合物进行鉴定和归纳。建立一种高压液相色谱-串联三重四级杆质谱法快速测定蜂花粉中8种黄酮类化合物的方法。破壁的试样经乙醇溶液提取后加酸水解,经反相色谱分离,质谱定性并测定8种黄酮类化合物含量,外标法定量。该方法线性良好,R2均大于0.999,检出限为0.15³.0 mg/kg,精密度为3.2%⁶.6%,回收率在91.2%¹00.2%,此方法操作简便,抗基质干扰、结果可靠。分析结果证实,蜂花粉中黄酮类化合物通常有槲皮素、山奈酚、芦丁等。可为识别蜂花粉的植物来源,鉴别真伪和建立相应的指纹图谱提供研究方法和实验依据。      

LC-MS/MS法测定小鼠脑组织中色氨酸及其3种代谢物

目的:建立检测小鼠脑组织中色氨酸、5-羟基色氨酸、5-羟色胺和5-羟基吲哚乙酸含量的LC-MS/MS分析方法。方法:采用TSK Gel amide 80（2.0 mm×15 cm,3μm）色谱柱,柱温35℃,乙腈:甲酸铵水溶液（15 mmol·L-1,p H5.5）（40∶60,v/v）为流动相,流速0.4 m L·min-1,采用正离子多反应监测（MRM）模式进行扫描。结果:4种待测物均在5 min内完成测定,色氨酸、5-羟基色氨酸、5-羟色胺和5-羟基吲哚乙酸最低定量限分别为0.2、0.1、0.2、0.2 ng·m L-1,最低检测限分别为0.05、0.02、0.05、0.05 ng·m L-1,在1⁵00 ng·m L-1浓度范围内线性关系均良好,方法的精密度和加样回收率均符合生物样品的分析要求。将该方法应用于口服萱草花总黄酮的小鼠脑提取物检测后发现:脑内四种待测物浓度明显发生改变,且变化规律与阳性药物帕罗西汀相同。结论:本方法简便、准确、灵敏度高,专属性强,重复性好,适用于脑组织中色氨酸-五羟色胺代谢过程中的4种成分的测定,可应用于神经精神类药物的药理作用机制研究。      

食品中丙烯酰胺分析方法研究进展

自从食品中发现一种潜在的致癌物质———丙烯酰胺后,世界各国的科学家进行了广泛深入的研究,在食品中丙烯酰胺的分析方法、形成机理、减少措施、暴露研究及风险评估等方面均有所进展。文中主要集中在食品中丙烯酰胺分析方法研究方面,主要包括水源水中丙烯酰胺的分析、聚丙烯酰胺中残留丙烯酰胺的分析以及采用GCMS、LCMS和LCMS/MS连用的方法精确分析食品中的丙烯酰胺,并提出了对食品中丙烯酰胺研究的进一步展望。