CN111521719A - Hilic-CAD联合检测脱氢抗坏血酸的方法 - Google Patents
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- SBJKKFFYIZUCET-UHFFFAOYSA-N Dehydroascorbic acid Natural products OCC(O)C1OC(=O)C(=O)C1=O SBJKKFFYIZUCET-UHFFFAOYSA-N 0.000 title claims abstract description 39
- 235000020960 dehydroascorbic acid Nutrition 0.000 title claims abstract description 39
- 239000011615 dehydroascorbic acid Substances 0.000 title claims abstract description 39
- 238000000034 method Methods 0.000 title claims abstract description 28
- 239000007788 liquid Substances 0.000 claims abstract description 8
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 claims description 30
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 24
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 claims description 15
- 235000019253 formic acid Nutrition 0.000 claims description 15
- 239000000243 solution Substances 0.000 claims description 14
- 239000007864 aqueous solution Substances 0.000 claims description 9
- 239000012159 carrier gas Substances 0.000 claims description 5
- 239000012046 mixed solvent Substances 0.000 claims description 3
- 238000005303 weighing Methods 0.000 claims description 3
- HQVFCQRVQFYGRJ-UHFFFAOYSA-N formic acid;hydrate Chemical compound O.OC=O HQVFCQRVQFYGRJ-UHFFFAOYSA-N 0.000 claims description 2
- 238000002347 injection Methods 0.000 claims description 2
- 239000007924 injection Substances 0.000 claims description 2
- 238000001514 detection method Methods 0.000 abstract description 5
- 239000003153 chemical reaction reagent Substances 0.000 abstract description 4
- 230000014759 maintenance of location Effects 0.000 abstract description 4
- 238000000105 evaporative light scattering detection Methods 0.000 abstract description 3
- 238000001819 mass spectrum Methods 0.000 abstract description 3
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 abstract description 3
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 12
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 4
- 229930003268 Vitamin C Natural products 0.000 description 4
- 235000019154 vitamin C Nutrition 0.000 description 4
- 239000011718 vitamin C Substances 0.000 description 4
- 150000002500 ions Chemical class 0.000 description 3
- 235000010323 ascorbic acid Nutrition 0.000 description 2
- 239000011668 ascorbic acid Substances 0.000 description 2
- 229960005070 ascorbic acid Drugs 0.000 description 2
- 150000000996 L-ascorbic acids Chemical class 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000002013 hydrophilic interaction chromatography Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000004043 responsiveness Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
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- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/62—Detectors specially adapted therefor
- G01N30/72—Mass spectrometers
- G01N30/7233—Mass spectrometers interfaced to liquid or supercritical fluid chromatograph
- G01N30/724—Nebulising, aerosol formation or ionisation
- G01N30/7266—Nebulising, aerosol formation or ionisation by electric field, e.g. electrospray
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
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Abstract
本发明涉及一种Hilic‑CAD联合检测脱氢抗坏血酸的方法，它包括以下方法：流动相配制步骤、脱氢抗坏血酸待测溶液配制步骤、高效液相色谱仪参数设置步骤与检测并输出对应的色谱图步骤。本发明与在普通氨基柱上检测相比，脱氢抗坏血酸的保留明显增强，方法重现性好，无需使用昂贵且不易清除的离子对试剂，更经济方便。本发明与用DAD及ELSD检测器相比，脱氢抗坏血酸的响应度明显提高，与质谱检测器相比，操作更简便。
Description
技术领域
本发明涉及一种脱氢抗坏血酸的方法，具体地说是一种Hilic-CAD联合检测脱氢抗坏血酸的方法。
背景技术
脱氢抗坏血酸是抗坏血酸（维生素C）的一种氧化降解杂质。通过监测脱氢抗坏血酸的含量，可以优化含有维生素C的食品及药品的生产工艺，控制维生素C的降解，优化含有维生素C产品的贮存条件。脱氢抗坏血酸的定量检测方法有报道，多采用普通氨基柱分离，离子对作为流动相添加剂，检测器通常为DAD、ELSD或者质谱检测器。这些方法普遍存在脱氢抗坏血酸在色谱柱上保留时间不足、响应度不足或者采用离子对试剂的问题。目前，尚未见Hilic-CAD联用且不需使用离子对试剂的技术检测脱氢抗坏血酸的报道。
发明内容
本发明的目的是克服现有技术中存在的不足，提供一种重现性好且更加经济方便的Hilic-CAD联合检测脱氢抗坏血酸的方法。
按照本发明提供的技术方案，所述Hilic-CAD联合检测脱氢抗坏血酸的方法包括以下方法：
步骤一、将体积浓度为0.01%~1%的甲酸水溶液与乙腈按照体积比（1~5）:（95~99）混合形成流动相；
步骤二、称取脱氢抗坏血酸，加甲酸水溶液与乙腈配成的体积比为（1~5）:（95~99）混合溶剂溶解稀释后得到10~100μg/ml的脱氢抗坏血酸待测溶液；
步骤三、高效液相色谱仪采用Hilic色谱柱，色谱柱产品参数为50~300mm×2.1~10mm、1.7~10μm，柱温设置为25℃~40℃，流速为0.5~1.5ml/min，CAD温度为30℃~50℃，载气压力为：50~70psi，进样量为：5~50μl；
步骤四、采用高效液相色谱仪对脱氢抗坏血酸待测溶液进行检测并输出对应的色谱图。
作为优选，步骤一和步骤二中，甲酸水溶液中甲酸的体积浓度为0.01%~1%。
作为优选，步骤一和步骤二中，甲酸水溶液中甲酸的体积浓度为0.1%。
作为优选，步骤一和步骤二中，甲酸水溶液与乙腈的体积比为3: 97。
作为优选，步骤二中，脱氢抗坏血酸待测溶液的质量浓度为100μg/ml。
作为优选，步骤三中，柱温设置为25℃。
作为优选，步骤三中，流速设置为1ml/min。
作为优选，步骤三中，CAD温度设置为35℃。
作为优选，步骤三中，载气压力设置为50psi。
作为优选，步骤三中，进样量设置为5μl。
本发明与在普通氨基柱上检测相比，脱氢抗坏血酸的保留明显增强，方法重现性好，无需使用昂贵且不易清除的离子对试剂，更经济方便。本发明与用DAD及ELSD检测器相比，脱氢抗坏血酸的响应度明显提高，与质谱检测器相比，操作更简便。
附图说明
图1是实施例1中脱氢抗坏血酸溶液的色谱图。
具体实施方式
下面结合具体实施例对本发明作进一步说明。
实施例1
一种采用亲水作用色谱法（Hilic）与电喷雾检测器（CAD）联用检测脱氢抗坏血酸的方法包括以下步骤：
步骤一、将体积浓度为0.1%的甲酸水溶液与乙腈按照体积比3: 97混合形成流动相；
步骤二、称取一定量的脱氢抗坏血酸，加甲酸水溶液与乙腈配成的体积比为3: 97混合溶剂溶解稀释后得到100μg/ml的脱氢抗坏血酸待测溶液，该步骤中甲酸水溶液体积浓度为0.1%；
步骤三、高效液相色谱仪采用Hilic色谱柱，色谱柱产品参数为100mm×4.6mm、2.7μm，将高效液相色谱仪柱温设置为25℃，流速为1.0ml/min，CAD温度为35℃，载气压力为：50psi，进样量为：5μl；
步骤四、采用高效液相色谱仪对脱氢抗坏血酸待测溶液进行检测并输出对应的色谱图。
检测结果：脱氢抗坏血酸的保留时间为7.476分钟，响应度为22PA。
以上所述仅为本发明的较佳实施例，并不用以限制本发明，凡在本发明的精神和原则之内，所作的任何修改、等他替换、改进等，均应包含在本发明的保护范围之内。
Claims (10)
1.一种Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是该方法包括以下方法：
步骤一、将体积浓度为0.01%~1%的甲酸水溶液与乙腈按照体积比（1~5）:（95~99）混合形成流动相；
步骤二、称取脱氢抗坏血酸，加甲酸水溶液与乙腈配成的体积比为（1~5）:（95~99）混合溶剂溶解稀释后得到10~100μg/ml的脱氢抗坏血酸待测溶液；
步骤三、高效液相色谱仪采用Hilic色谱柱，色谱柱产品参数为50~300mm×2.1~10mm、1.7~10μm，柱温设置为25℃~40℃，流速为0.5~1.5ml/min，CAD温度为30℃~50℃，载气压力为：50~70psi，进样量为：5~50μl；
步骤四、采用高效液相色谱仪对脱氢抗坏血酸待测溶液进行检测并输出对应的色谱图。
2.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤一和步骤二中，甲酸水溶液中甲酸的体积浓度为0.01%~1%。
3.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤一和步骤二中，甲酸水溶液中甲酸的体积浓度为0.1%。
4.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤一和步骤二中，甲酸水溶液与乙腈的体积比为3: 97。
5.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤二中，脱氢抗坏血酸待测溶液的质量浓度为100μg/ml。
6.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤三中，柱温设置为25℃。
7.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤三中，流速设置为1ml/min。
8.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤三中，CAD温度设置为35℃。
9.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤三中，载气压力设置为50psi。
10.根据权利要求1所述的Hilic-CAD联合检测脱氢抗坏血酸的方法，其特征是：步骤三中，进样量设置为5μl。
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|---|---|---|---|---|
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