不同液相色谱-质谱联用法分析贝类样品中神经毒素β-N-甲氨基-L-丙氨酸的比较

基于液相色谱-串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS),比较了两种亲水交互作用(hydrophilic liquid interaction chromatography column,HILIC)色谱柱和C18色谱柱分析扁玉螺(Nev...     

天津市售主要经济贝类中麻痹性贝类毒素污染现状及特征分析

使用液相色谱串联质谱法(LC-MS/MS)检测9种天津市售主要经济贝类中13种麻痹性贝类毒素(paralytic shellfish poisoning,PSP)的含量,分析PSP的污染现状和成分特征。结果显示,共计6个品种(紫贻贝、虾夷扇贝、牡蛎、毛蚶、脉红螺、菲律宾蛤仔)检出PSP。其中虾夷扇贝、紫贻贝、毛蚶3个品种的检出率较高且含量较高,检出率分别为66.7%、58.3%、58.3%。PSP中检出率较高的毒素包括N-磺酰胺甲酰基类毒素C1、C2和膝沟藻毒素GTX1、GTX2、GTX3,毒力占比主要以膝沟藻毒素GTX1-4为主,毛蚶蓄积STX毒素的能力强于其他贝类。全年的PSP含量季节变化为夏季最高,冬季最低。PSP主要存在于贝类的消化腺内,全年不同月份的PSP含量均在安全食用范围内。     

Quantitation Overcoming Matrix Effects of Lipophilic Toxins in Mytilus galloprovincialis by Liquid Chromatography-Full Scan High Resolution Mass Spectrometry Analysis (LC-HR-MS)

The analysis of marine lipophilic toxins in shellfish products still represents a challenging task due to the complexity and diversity of the sample matrix. Liquid chromatography coupled with mass spectrometry (LC-MS) is the technique of choice for accurate quantitative measurements in complex samples. By combining unambiguous identification with the high selectivity of tandem MS, it provides the required high sensitivity and specificity. However, LC-MS is prone to matrix effects (ME) that need to be evaluated during the development and validation of methods. Furthermore, the large sample-to-sample variability, even between samples of the same species and geographic origin, needs a procedure to evaluate and control ME continuously. Here, we analyzed the toxins okadaic acid (OA), dinophysistoxins (DTX-1 and DTX-2), pectenotoxin (PTX-2), yessotoxin (YTX) and azaspiracid-1 (AZA-1). Samples were mussels (Mytilus galloprovincialis), both fresh and processed, and a toxin-free mussel reference material. We developed an accurate mass-extracted ion chromatogram (AM-XIC) based quantitation method using an Orbitrap instrument, evaluated the ME for different types and extracts of mussel samples, characterized the main compounds co-eluting with the targeted molecules and quantified toxins in samples by following a standard addition method (SAM). An AM-XIC based quantitation of lipophilic toxins in mussel samples using high resolution and accuracy full scan profiles (LC-HR-MS) is a good alternative to multi reaction monitoring (MRM) for instruments with HR capabilities. ME depend on the starting sample matrix and the sample preparation. ME are particularly strong for OA and related toxins, showing values below 50% for fresh mussel samples. Results for other toxins (AZA-1, YTX and PTX-2) are between 75% and 110%. ME in unknown matrices can be evaluated by comparing their full scan LC-HR-MS profiles with those of known samples with known ME. ME can be corrected by following SAM with AM-XIC quantitation if necessary.     

液相色谱串联质谱检测动物源性食品中莫西丁克残留

[目的]建立动物源性食品中莫西丁克残留的检测方法。[方法]首先对样品净化方法和试验参数进行了优化。利用乙腈对样品中莫西丁克进行提取,利用分散型固相萃取净化莫西丁克,利用液相色谱串联质谱测定动物源性食品中莫西丁克的含量。[结果]最佳质谱条件为：离子源温度300℃、喷雾针压力15 Psi、干燥气流速6 L/min。在此条件下,子离子峰的响应值最高。该方法的检测限为0.005 mg/kg,当在添加浓度0.005～0.02 mg/kg时,回收率为81.08%～113.72%,RSD为4.21%～9.69%。[结论]该方法步骤简单、可操作性强、有机溶剂用量少、成本低、杂质干扰少,有良好的精密度及较低的方法检测低限,适于动物源性食品中莫西丁克的快速测定。     

中国3个主栽烟草品种的差异蛋白质组学研究

【目的】从全蛋白质组学水平研究烟草,筛选具有重要特性的蛋白质,阐明其相关代谢通路,使得烟草的基础科学研究和品种培育工作有所突破。【方法】利用苯酚法提取中国3个主栽烟草品种红花大金元、K326和云烟87叶片的蛋白质,酶解后采用串联质量标签技术(tandem mass tag,TMT)标记,结合二维液相色谱串联质谱技术(2D LC-MS/MS)对3个烟草品种叶片的蛋白质组成进行分析,运用Proteome Discoverer软件提取谱图后在MASCOT搜索引擎上鉴定蛋白质和肽段,并对鉴定的蛋白质进行系统的生物信息学分析,包括数据相关性分析、分层聚类分析和主成分分析,同时按照2倍上下调的原则采用火山图筛选不同品种烟草中表达差异显著的蛋白质,最后利用KEGG通路分析烟草中重要蛋白质的分布及功能。【结果】鉴定红花大金元、K326和云烟87烟草样品的蛋白质Group总数为3 079,蛋白质ID数为10 343。从分层聚类分析和主成分分析中蛋白质的整体表达情况可见,K326和云烟87蛋白质表达情况较相似,而红花大金元与前两者相差较大。后续的差异蛋白质筛选也验证了该结果,表现为云烟87和K326之间仅存在29个差异表达的蛋白质,其中13个蛋白质覆盖8条代谢通路,包括类黄酮生物合成,参与类黄酮合成路径的查尔酮合成酶在K326中的相对含量显著高于云烟87。红花大金元和云烟87之间有160个蛋白质差异表达,其中103个蛋白质覆盖42条代谢通路,包括谷胱甘肽代谢,相对定量结果显示谷胱甘肽代谢途径相关的3种酶,谷胱甘肽过氧化物酶、磷脂过氧化氢物谷胱甘肽过氧化物酶和谷胱甘肽S-转移酶在红花大金元中的含量均显著低于云烟87。红花大金元和K326之间存在119个差异蛋白质,其中89个蛋白质覆盖41条代谢通路,包括外源物质细胞色素P450代谢。筛选获得的重要差异蛋白质的相对定量结果显示一些与抗性相关的蛋白质,如蛋白酶抑制剂,在云烟87中的表达量远低于K326和红花大金元,同时结果表明红花大金元中的超氧化物歧化酶含量处于较低水平。【结论】TMT技术结合2D LC-MS/MS可对不同品种烟草的叶片蛋白质进行有效地分离和鉴定,鉴定出的蛋白质大部分是参与光合作用、物质代谢或者与抗逆性相关。     

液相色谱-质谱法检测油菜子、植株和土壤中三唑醇的残留

建立液相色谱-质谱法检测油菜子、植株和土壤中三唑醇残留量的方法。油菜子和植株经乙腈提取,经Carbon/NH2柱净化,实现样品的制备;土壤则经乙腈提取浓缩后直接检测。在岛津VP-ODS色谱柱上,以乙腈+0.1%甲酸水为流动相,梯度洗脱,采用电喷雾正离子(ESI+)sim扫描离子模式,以m/z 296为定量检测离子。结果表明,三唑醇保留时间在11.9 min左右。在0.02~2.00 mg/L的浓度范围内呈良好的线性关系,其相关系数为0.999 7。在油菜子、植株、土壤中添加0.02、0.20、2.0 mg/kg 3个浓度的三唑醇标样,其回收率为71%~110%,相对标准偏差(n=5)均小于10%,最低检出量(S/N=10)为1.7×10-9 g。该方法准确度和精密度均能满足检测要求。     

同位素内标-液质联用法测定鸡可食性组织中替米考星残留量

建立了鸡四种可食性组织中替米考星的液相色谱串联质谱内标法。样品经磷酸二氢钾缓冲液和乙腈提取后,C18固相萃取柱净化,以0.1%甲酸乙腈溶液和0.1%甲酸水为流动相,液相色谱串联质谱法测定,内标法定量。试验结果表明：鸡四种可食性组织中替米考星在1 μg/kg~100 μg/kg的浓度范围内呈线性关系,相关系数r大于0.99;替米考星在鸡四种可食性组织中的检测限为0.5 μg/kg,定量限为1 μg/kg;四种组织中替米考星添加浓度为1 μg/kg~4800 μg/kg时,批间平均回收率为94.6%~100.9%,批内、批间变异系数在1.11%-5.06%范围之间。本方法适用于鸡四种可食性组织中替米考星含量的测定。     

阿诺宁两活性成分在大鼠肝微粒体中的酶动力学研究

[目的]建立番荔枝内酯类化合物squamocin和bullatacin的液相色谱串联质谱(LC-MS/MS)检测方法,并测定其在大鼠肝微粒体中的酶动力学参数。[方法]建立微粒体孵育体系,启动反应后测定不同时间孵育体系内剩余的squamocin和bullatacin的浓度,用底物消除法计算其酶动力学参数。[结果]在大鼠肝微粒体中,squamocin和bullatacin的表观米氏常数KM值分别为(1.6±0.5)和(2.0±0.4)μmol/L。[结论]该方法适用于squamocin和bullatacin的酶动力学常数的测定。     

Level and clinical significance of RNA oxidative damage in human gastric cancer and para-carcinoma tissues

AIM: To investigate the RNA oxidative damage in human gastric cancer tissue and para-carcinoma tissue for exploring the role of RNA oxidation in the occurrence of gastric cancer. METHODS: Immunohistochemical observation and LC-MS/MS analysis were performed in 61 cases of gastric carcinoma. The position and concentration of 8-oxoguanosine (8-oxoGsn) were detected, respectively. RESULTS: The results of immunohistochemical observation showed that 8-oxoGsn was obviously up-regulated in the gastric cancer. The positive staining mainly accumulated in the cytoplasm of the tumor cells. The results of mass spectrometry showed that the level of 8-oxoGsn in the gastric cancer tissues was higher than that in the para-carcinoma tissues (P<0.05). CONCLUSION: 8-oxoGsn is up-regulated in gastric cancer. RNA oxidative damage may play important roles in the occurrence of gastric cancer.     

固相萃取-GC/LC-MS/MS测定茶叶中79种农药残留

建立了一套以一次固相萃取前处理方法,运用气相色谱-串联质谱（GC-MS/MS）和液相色谱-串联质谱（LC-MS/MS）两种检测技术,测定茶叶中79种农药残留的可靠、快速、高通量检测方法。该方法用20βmL乙腈一次均质提取,分取提取液10βmL,用乙腈+甲苯（3∶1）20βmL洗脱过Carbon/NH₂小柱净化,用5βmL乙腈定容混匀,于GC-MS/MS和LC-MS/MS同时检测。结果表明,79种农药在0.01~0.40βmg·L^-1范围内线性良好,相关系数（R²）在0.995以上;高、低、中3种水平加标回收率在67.3%~130.8%内;相对标准偏差（RSD）在15%内;98.7%的农药定量限（LOQ）≤0.01βmg·kg^-1;使用多种类茶叶实际检测,结果均合格。因此,该方法能同时检测含有机磷、有机氯、拟除虫菊酯类、氨基甲酸酯类和有机杂环类5大类农药,更有利于大批量茶叶样品的多农残检测。