超声辅助酶法提取-液相色谱-原子荧光光谱法测定富硒黑木耳中硒形态

建立了富硒黑木耳中硒代胱氨酸、硒代半胱氨酸、亚硒酸、硒蛋氨酸、硒酸5种硒形态的液相色谱-原子荧光光谱分析方法.通过链酶蛋白酶E酶解,结合超声提取后,选取Hamilton PRP-X100离子交换色谱柱(250 mm×4.1 mm,10μm),40 mmol/L的磷酸氢二铵为流动相,在16 min内,5种硒形态完全达到基...     

液相色谱-氢化物发生原子荧光光谱法测定富硒酵母中硒的形态

建立了富硒酵母中硒酸Se(Ⅵ)、亚硒酸Se(Ⅵ)、硒代蛋氨酸(SeMet)和硒甲基硒代半胱氨酸(SeMeCys)4种硒形态的高效液相色谱-氢化物发生原子荧光光谱(HPLC-HG-AFS)分析方法。样品采用蛋白酶和胰蛋白酶酶解提取,20 mmol/L(NH4)2HPO4为流动相,经PRP-X100阴离子交换色谱柱分离后HG-AFS测定。结果显示,4种硒形态标准曲线的线性关系良好(R2≥0.9995),方法检出限为0.5~5.0μg/kg,平均回收率为82.5%~101.2%,日内相对标准偏差≤8.6%,日间相对标准偏差≤14.5%。本方法前处理简单、灵敏度高、设备便宜、运行费用低廉,适用于富硒饲料产品中硒的形态分析。     

高效液相色谱-原子荧光光谱联用技术测定富硒鸡蛋中的硒形态

为监测市场上富硒鸡蛋中蛋清及蛋黄中硒形态的种类及分布规律,优化建立了一种高效液相色谱-氢化物发生-原子荧光光谱联用技术测定富硒鸡蛋蛋清及蛋黄粉中硒形态的方法。随机选取富硒鸡蛋样品,进行蛋清蛋黄分离、冷冻干燥及蛋黄脱脂处理,处理后样品经Tris-HCl缓冲液（5 mmol/L, pH=7.5）55 oC恒温振荡、酶解提取,高速离心机（10000 r/min）离心10 min,上清液过0.22 μm有机滤膜后采用高效液相色谱-氢化物发生-原子荧光光谱联用技术进行测定,外标法定量。结果表明,该方法能在11 分钟内实现5种硒形态的基线分离,且各种硒形态的线性相关系数(r)均大于0.9994,其检出限在0.5-3.0 μg/L之间,回收率在81.6%~110.4%之间,可以满足检测需求,按照该实验方法测定市场上富硒鸡蛋中的硒形态,方法灵敏度高、准确性好,且测得的富硒鸡蛋蛋清样品中硒形态主要成分为硒代蛋氨酸,而蛋黄样品主要成分为硒代半胱氨酸,还有少量的硒代蛋氨酸,同时,一些鸡蛋还含有极少量的亚硒酸根,但蛋清及蛋黄中硒代氨基酸的含量总和占总硒含量的83.3-98.4%,适用于富硒鸡蛋中的硒形态提取。由此可见,市场上富硒鸡蛋中的硒主要以硒代氨基酸的形式存在,比较适合日常补硒,且该方法可以对市场上富硒食品起到一定监测作用。     

高效液相色谱-电感耦合等离子体质谱法测定富硒碎米荠中的硒形态

建立了富硒碎米荠中Se(Ⅵ)、Se(Ⅳ)、硒代胱氨酸(SeCys_2)、甲基硒代半胱氨酸(MeSeCys)、硒代蛋氨酸(SeMet) 5种硒形态的高效液相色谱-电感耦合等离子体质谱(HPLC-ICP-MS)分析方法。通过风味蛋白酶和蛋白酶E两种酶先后添加的顺序提取不同形态硒化合物,经两次酶解提取后,稀释到合适浓度。选取Thermo Scientific Hypersil GOLD C8色谱柱(250 mm×4.6 mm×5μm),以含0.05%七氟丁酸与3%甲醇的20 mmol/L KH_2PO_4作为流动相进行等度洗脱,可在6 min内将5种硒形态完全分离。5种硒形态在线性范围内相关系数(r~2)均大于0.999;加标回收率均在85.8%～106.3%之间;变异数小于5%;方法的检出限为0.08～0.25μgSe/L;定量限为0.25～0.54μgSe/L。应用该方法测定实际样品,发现不同碎米荠中有机硒形态占总硒的71.2%～87.9%,其中以SeCys_2和SeMet两种硒形态为主,同时还含有少量无机硒和未知形态硒。     

高效液相色谱-电感耦合等离子体质谱（HPLC-ICP-MS）联用测定富硒小麦中硒代氨基酸

为科学补硒和促进富硒小麦的种植推广,建立了高效液相色谱-电感耦合等离子体质谱联用技术(HPLC-ICP-MS)检测富硒小麦中硒代氨基酸的方法。用蛋白酶XIV辅助微波振荡提取富硒小麦中硒代氨基酸,采用C18 分离柱分离,以30.0mmol/L磷酸氢二铵+1.0%甲醇+2.0mmol/L四丁基溴化铵溶液(pH=6.5)为流动相,能在10min内实现5种硒代氨基酸的分离。在高能氦气模式（HEHe）下,用78Se的色谱峰积分面积作为定量依据,5种硒代氨基酸在1.0～200.0μg/L范围内线性相关性良好,检出限在 0.11~0.29μg/L之间。以富硒小麦为基体进行加标回收试验,除硒代胱氨酸（SeCys2）可能不稳定,易分解造成回收率偏低外,其他4种硒代氨基酸的加标回收率在92.34～102.46%之间,相对标准偏差为 1.6 %~4.2 %（n=7）。用该方法测定了农业科技工作者种植推广的富硒小麦,结果发现小麦中的硒赋存形态多为硒代蛋氨酸（SeMet）,此外,小麦中还含有少量硒代胱氨酸（SeCys2）、硒代半胱氨酸（SeCys）、甲基硒代半胱氨酸（MeSeCys）和硒代乙硫氨酸（SeEt)。该方法具有良好的精密度和准确度,适用于富硒小麦中硒代氨基酸的形态分析。     

高效液相色谱-电感耦合等离子体质谱法测定人尿中硒形态

采用高效液相色谱-电感耦合等离子体质谱法(HPLC-ICP/MS)测定人尿中硒代胱氨酸(SeCys_2)、甲基硒代半胱氨酸(MeSeCys)、亚硒酸盐[Se(Ⅳ)]、硒代蛋氨酸(SeMet)、硒酸盐[Se(Ⅵ)]5种硒形态。样品经超纯水稀释后,采用Hamilton PRP-X100色谱柱(250 mm×4 mm,10μm)分离,以40 mmol/L磷酸氢二铵(含1%甲醇,pH 5)为流动相进行等度洗脱,13 min内可将5种硒形态分离。5种硒形态的线性范围为0～300.0μg/L,相关系数(r)均大于0.999,检出限为0.2～0.5μg/L。除SeCys_2的加标回收率为37.7%～70.4%外,MeSeCys、Se(Ⅳ)、SeMet、Se(Ⅵ)的加标回收率为80.0%～123%;5种硒形态的相对标准偏差(RSD)均不大于7.8%。应用该方法测定实际样品,结果显示人尿中硒形态主要以SeCys_2为主,同时含有少量MeSeCys、SeMet、无机硒及未知含硒化合物。     

高效液相色谱-电感耦合等离子体质谱分析几种富硒产品的硒总量及其形态

本文采用高效液相色谱-电感耦合等离子体质谱(HPLC-ICP-MS)分析了几种富硒产品中硒的总量及其形态。结果表明,四种富硒产品的硒总量均有不同程度增加,富硒大米和富硒茶叶中的硒总量较高,分别为1.300μg/g和0.459μg/g,富硒大米和富硒大葱的富硒倍率相对较高,分别为未富硒产品的24.1和17.8倍。以0.1mol/LHCl辅助超声预处理样品进行富硒螺旋藻和富硒大葱中硒的形态分析,发现富硒大葱中的硒主要为有机态的甲基硒代半胱氨酸(MeSeCys)以及部分硒代蛋氨酸(SeMet),而富硒螺旋藻中的硒大部分为无机硒。说明经根部吸收等富集方式可有效将无机硒转化为有机硒,而某些方式未能完全转化。     

高效液相色谱与电感耦合等离子体质谱联用技术用于胁迫富硒海带硒形态研究

通过对海带胁迫富硒培养, 研究了海带对硒的富集总量及硒化学形态转变. 建立了反相离子对缓冲盐等3个色谱系统的高效液相色谱与电感耦合等离子体质谱联用(RP-HPLC-ICP-MS)技术测定亚硒酸钠、硒甲基半胱氨酸(MeSeCys)、硒代蛋氨酸(SeMet)三种硒形态. 采用3种提取溶剂, 超声提取缩短提取时间, 分离检测富硒海带的主要硒形态为亚硒酸钠、硒甲基半胱氨酸和硒代蛋氨酸.     

HPLC-DRC-ICP-MS测定富硒蔬菜中的硒形态

建立了高效液相色谱-动态反应池-电感耦合等离子体质谱(HPLC-DRC-ICP-MS)联用测定硒代胱氨酸(SeCys2)、甲基硒代半光氨酸(MeSeCys)、亚硒酸盐(SeIV)、硒代蛋氨酸(SeMet)和硒酸盐(SeVI)的方法。样品通过胃蛋白酶提取,采用Hamilton PRP X-100色谱柱(250 mm×4.6 mm,10μm),使用5 mmol/L的柠檬酸溶液(pH 4.7)作为流动相,在10 min内可以完全分离5种硒形态。在80Se质量数下,采用甲烷作为DRC反应气,可有效消除40Ar40Ar+对80Se的干扰,提高检测灵敏度。电感耦合等离子体质谱(ICP-MS)检测,各硒形态的线性相关系数均大于0.9990,Se(IV)、Se(VI)、SeMet、SeCys2、MeSeCys的定量限分别为5,10,10,5,5μg/kg,5种硒形态的加标回收率在80.1%～99.2%之间,可满足蔬菜中硒形态定量分析。     

HPLC-ICP-MS或HPLC-FAAS法分离测定硒化合物（英文）

提出了一种用高效液相色谱（HPLC）分离和用电感偶合等离子体质谱仪（ICP－MS）或火焰原子吸收光谱仪（FAAS）作元素专一检测器在线测定硒的化学形态的方法。在优化的HPLC条件下,用ESAⅢ阴离子色谱柱（250mm×4．6mm）,以柠檬酸铵为流动相（5．5mmol／L,pH5．5,流速1．5mL／min）,进样量100μL,分离和测定三甲基硒离子、硒代蛋氨酸、亚硒酸和硒酸盐只需8min。HPLC－FAAS在线分析4种硒化合物的检测限为p（Se）=1mg／L。用超声雾化器作ICP－MS的接口,HPLC－ICP－MS在线分析4种硒化合物的检测限分别为P（Se）=0．34μg／L（亚硒酸）,0．18μg／L（硒代蛋氨酸）,0．08μg／L（三甲基硒离子）和0．07μg／L（硒酸盐）。与气动雾化器接口相比,信号强度增加7至31倍。     

HPLC-ICP-MS或HPLC-FAAS法分离测定硒化合物（英文）

 提出了一种用高效液相色谱（HPLC）分离和用电感偶合等离子体质谱仪（ICP－MS）或火焰原子吸收光谱仪（FAAS）作元素专一检测器在线测定硒的化学形态的方法。在优化的HPLC条件下,用ESAⅢ阴离子色谱柱（250mm×4．6mm）,以柠檬酸铵为流动相（5．5mmol／L,pH5．5,流速1．5mL／min）,进样量100μL,分离和测定三甲基硒离子、硒代蛋氨酸、亚硒酸和硒酸盐只需8min。HPLC－FAAS在线分析4种硒化合物的检测限为p（Se）=1mg／L。     

Determination of trimethylselenonium iodide, selenomethionine, selenious acid, and selenic acid using high-performance liquid chromatography with on-line detection by inductively coupled plasma mass spectrometry or flame atomic absorption spectrometry

An analytical method has been developed for the determination of selenious acid, selenic acid, trimethylselenonium ion, and selenomethionine. The four selenium compounds were separated by HPLC on a column (25 cm×4 mm I.D.) of the anion-exchanger ESA Anion III with a mobile phase (1.5 ml/min) of 0.0055 M ammonium citrate (pH 5.5). Detection was carried out using an on-line inductively coupled plasma mass spectrometer (ICP-MS) or a flame atomic absorption spectrometer (FAAS) as the selenium-specific detector. The chromatographic parameters and the chemical factors affecting the separation of the selenium species were optimized. The four selenium compounds could be separated within 8 minutes. The detection limits of the coupled HPLC–FAAS system were approximately 1 mg Se/l for each compound (100 μl injection), estimated as three times the base-line noise of the chromatograms. More powerful selenium detection was achieved with an ICP-MS. Selenium was measured at m/z 78. To increase the nebulization efficiency, the Meinhard concentric glass nebulizer was replaced by an ultrasonic nebulizer. The ICP-MS signal intensity was increased with the ultrasonic nebulization by a factor of 7 times for selenious acid and 24 to 31 times for trimethylselenonium ion, selenomethionine, and selenic acid compared to that with the Meinhard nebulization. The detection limits achieved by the HPLC–ICP-MS with the ultrasonic nebulization were 0.08 μg Se/l for trimethylselenonium ion, 0.34 μg Se/l for selenious acid, 0.18 μg Se/l for selenomethionine, and 0.07 μg Se/l for selenic acid, respectively.     

Speciation of selenoamino acids by on-line HPLC ETAA spectrometry

An analytical method was developed to determine selenoamino acids in the presence of other compounds. Separation has been achieved by High Performance Liquid Chromatography (HPLC) using electrothermal atomic absorption (ETAA) spectrometry as a very sensitive and element-specific detector. On-line HPLC ETAAS speciation of selenocystine and selenomethionine has been studied, using a laboratory made interface. Analytical characterization of the method has been realized with standard solutions. Using a 100μl sample loop, the detection limits were calculated as 8 μgl^?1 for selenomethionine and 10 μgl^?1 for selenocystine with repeatability and reproducibility of 4% and 7% respectively. The method has been applied to the determination of selenoamino acids in an extract of white clover (CRM402) certified for total selenium.     

柱前衍生化-HPLC法测定恩施3种富硒蔬菜中硒氨基酸

采用HPLC与ICP-MS间隙联用的方法,以柱前衍生化-HPLC法进行定性定量分析,使用ICP-MS鉴定,建立一种富硒蔬菜中硒氨基酸的分离检测方法。结果表明:所测定的硒代氨基酸在其线性范围内呈现良好的线性关系(R2>0.999)。该方法中硒代蛋氨酸和硒代胱氨酸的检出限分别为0.204 mg/L和0.680 mg/L,加标回收率分别为97.4%和94.0%,RSD分别为2.1%和0.69%。检测恩施富硒蔬菜样品,白菜、萝卜叶和苋菜含有硒代蛋氨酸和硒代胱氨酸。此方法可用于富硒蔬菜中硒代氨基酸的含量检测。     

高效液相色谱-电感耦合等离子体质谱联用检测食品中的五种硒形态

建立了高效液相色谱-电感耦合等离子体质谱(HPLC-ICP-MS)联用检测硒酸盐(SeVI)、亚硒酸盐(SeIV)、硒代蛋氨酸(SeMet)、硒代胱氨酸(SeCys2)和硒代乙硫氨酸(SeEt)的方法。采用Hamilton PRP X-100色谱柱（250 mm×4.6 mm, 5 μm）,使用5 mmol/L的柠檬酸溶液(pH 4.5)作为流动相,电感耦合等离子体质谱(ICP-MS)检测,在21 min内可以完全分离5种硒形态。各形态硒的线性相关系数均大于0.9995, SeVI、SeIV、SeMet、SeCys2、SeEt的检出限分别为0.4、0.4、5.6、0.9、1.2 μg/L。探讨了不同提取方法的提取效果,鲜蘑菇和猪肉样品加标回收实验表明,对水溶性良好的无机硒和硒代蛋氨酸而言,采用柠檬酸溶液提取的效果非常好,SeIV和SeVI的回收率均在100%左右,SeMet的回收率为85.0%～95.3%;用蛋白酶水解提取,SeCys2和SeEt的回收率为79.9%～91.5%。该方法可完全满足食品中这5种硒形态的准确定量分析。     

Retention study of inorganic and organic selenium compounds on a silica-based reversed phase column with mixed ion-pairing reagents

Summary Separation of seven inorganic and organic selenium compounds, namely selenic acid [Se(VI)], selenous acid [Se(IV)], trimethylselenonium iodide (TMSe⁺), selenocystine (SeCys), selenomethionine (SeMet), selenoethionine (Seet), and selenocystamine (SeCM), has been performed on a LiChrosorb C 18 column by using mixed ion-pair reagents; 1-butanesulfonic acid and tetramethylammonium hydroxide. Flame atomic absorption spectrometry (FAAS) was used as an element-specific detector. The retention behaviors of selenium compounds in terms of several chromatographic parameters, such as pH of the mobile phase, the concentrations of ion-pair reagents, and the content of organic modifier (methanol) were investigated. It was found that the separation of both inorganic and organic selenium compounds can be achieved within 12 min with a mobile phase of 10 mM 1-butanesulfonic acid −4 mM tetramethylammonium hydroxide −4 mM malonic acid −0.05% methanol adjusted to pH 4.5 at a flow rate of 1.0 mL min⁻¹. The results obtained in this study showed that the use of mixed ion-pair reagents is very useful to improve the separation of selenium compounds. The applicability of this technique for the speciation of selenium compounds in real samples was demonstrated by the determination of selenium compounds in a selenium nutritional supplement. The results were found to be in good agreement with those obtained by ion-exchange HPLC-ICP-MS.     

Selenoamino acid speciation using HPLC–ETAAS following an enzymic hydrolysis of selenoprotein

A high-pressure liquid chromatography–electrothermal atomic absorption spectroscopy (HPLCETAAS) hyphenated technique was used for the determination of seleno compounds present in a selenium-enriched yeast. Conditions were optimized for the separation and quantification of the selenoamino acids, selenocystine and selenomethionine, in the presence of other compounds. The separation was achieved by ion-pairing chromatography using sodium heptanesulphonate as the anionic counterion. On-line detection was carried out using electrothermal atomic absorption with palladium(II) as a matrix modifier. Different extraction procedures were tested on a seleniumenriched yeast. A 92% recovery of the total selenium present in the material was obtained. Attempts to evaluate selenium speciation were carried out; selenomethionine and selenocystine were identified as the major components (42% and 35% respectively).     

Determination of selenoamino acids using two-dimensional ion-pair reversed phase chromatography with on-line detection by inductively coupled plasma mass spectrometry

Analytical methods for the speciation of nine selenium species (selenite, selenate, selenourea, trimethylselenonium ion, selenocystamine, selenocystine, selenocysteine, selenomethionine and selenoethionine) that are commonly encountered in biological and environmental samples were developed. Good separation was achieved by either a mixed ion-pair reversed phase chromatography (LiChrosorb RP 18, 2.5 mM 1-butanesulfonate-8 mM tetramethylammonium hydroxide-4 mM malonic acid-0.05% methanol, pH 4.5) or a conventional ion-pair reversed phase chromatography (Inertsil ODS, 10 mM tetraethylammonium hydroxide-4.5 mM malonic acid, pH 6.8) with on-line ICP-MS detection. Using a 20-μl sample loop, low detection limits around 1 ng ml⁻¹ expressed as Se were achieved for the examined selenium species. The methods were used for the determination of selenoamino acids in a selenium nutritional supplement. The developed methods were found to be rather robust. No alteration of the separation was observed when the protease enzymatic extracts were analyzed without dilution. Both water extracts and enzymatic extracts were chromatographed first with the mixed ion-pair reversed phase chromatographic system, then the major chromatographic peaks were collected and analyzed by the second ion-pair reversed phase chromatographic system for a further verification of their identity. Selenomethionine was found to be the major selenium species in the supplement. A major unknown species, probably Se-adenosylhomocysteine, could be determined in the extracts. A biological reference material, Dolt-2, was also examined for the selenoamino acids. Selenocystine and selenomethionine could be detected in its enzymatic extract, suggesting that Dolt-2 may be used as a reference material for the identification of selenoamino acids in biological and environmental samples. As selenoethionine does not occur naturally in the investigated samples, it is added as an internal standard in this study.     

用离子抑制色谱法分析二（2,2,6,6—四甲基—4—哌啶基）马来酸酯合？…

建立了用离子抑制色谱法分析二（２,２,６,６－四甲基－４－哌啶基）马来酸酯合成反应液的方法。平均回收率为９８．８％,相对标准偏差为０．５６％,测量的平均相对偏差不大于５．０％,方法简单,快速,可用于工艺条件的选择和质量检测。     

硒酵母中有机硒及硒代氨基酸含量的测定方法

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