PHENOLIC ANTIOXIDANTS DETERMINATION IN FOOD ITEMS USING REVERSED-PHASE HPLC

The determination of synthetic phenolic antioxidants （SPAs） including propyl gallate （PG）, tertiary butyl hydroquinone （TI3HQ）, butylated hydroxyanisole （BHA） and butylated hydroxytoluene （BHT） in food items is reported using high performance liquid chromatography （HPLC）. A Cls column is used as the stationary phase, acetonltrile and water：Acetic acid （1%） is used as the mobile phase of gradient elution and the UV detec- tor is set at 280 nm. Under the above conditions, four antioxidents is completely separated within 8 rain. The limit of detection, linear range, and reproducibility of HPLC are evaluated. Isolation parameters of SPAs from different types of food items （cooking oil, margarine and butter, and cheese） are optimized. SPAs are extracted from food items through extraction with methanol/acetonitrile （1 ： 1, in volume）, vortex, ultrasonic treatment and precipitation in a freezer （2 h）. For cooking oil margarine, butter and cheese at 50 and 200 rag/L, recoveries of SPAs are 93.3%0--108.3% （PG）, 85.3~^--108.3~~ （TBHQ）, 96.7~^--101.2~/6 （BHA）, and 73.9^-- 94.6% （BHT）. The method is applied to the determination of SPAs in 38 food items （16 cooking oils, 8 mar- garine, 6 butter and 6 cheese samples）. The levels of SPAs in positive samples are all below the legal limits of China.

Application of HPLC and ESI-MS techniques in the analysis of phenolic acids and flavonoids from green leafy vegetables (GLVs)

Diets containing high proportions of fruits and vegetables reduce the risk of onset of chronic diseases. The role of herbal medicines in improving human health is gaining popularity over the years, which also increases the need for safety and efficiency of these products. Green leafy vegetables （GLVs） are the richest source of phenolic compounds with excellent antioxidant properties. Increased consumption of diets containing phenolic compounds may give positive and better results to human health and significantly improves the immune system. Highly selective, susceptible and versatile analytical techniques are necessary for extraction, identifica- tion, and quantification of phenolic compounds from plant extracts, which helps to utilize their important biological properties. Recent advances in the pre-treatment procedures, separation techniques and spectro- metry methods are used for qualitative and quantitative analysis of phenolic compounds. The online coupling of liquid chromatography with mass spectrometry （LC-MS） has become a useful tool in the metabolic profiling of plant samples. In this review, the separation and identification of phenolic acids and flavonoids from GLVs by LC-MS have been discussed along with the general extraction procedures and other sources of mass spectrometer used. The review is devoted to the understanding of the structural configuration, nature and accumulation pattern of phenolic acids and flavonoids in plants and to highlighting the recent developments in the chemical investigation of these compounds by chromatographic and spectroscopic techniques. It concludes with the advantages of the combination of these two methods and prospects.

Temperature and Acidified Solvent Effect on Total Anthocyanins and RP-HPLC Phenolic Acids Determination in Selected Spices

Total anthocyanins of spices (Syzygium aromaticum L., Coriadrum sativum L., Cuminum cyminum L., Zingiber officinale Rosc., Elettaria cardamomum, Curcuma longa, Rhus coriaria L., Cinnamomum zeylanicum Blume, Foeniculum vulgare Mill and Laurus nobilis L.) were determined using acidified (1% HCl) solvents (methanol, ethanol and acetone) at three temperatures (20℃, 40℃ and 60℃). Also phenolic acids were separated and identified by RP-HPLC. Results showed that sumac and cinnamon had the highest levels of anthocyanins, while for the acetone the cinnamon indicated the highest amount of anthocyanins when methanol and ethanol were used as extracting solvents at 20℃. At 40℃ using ethanol, sumac showed the highest level of anthocyanins whereas acetone solvent yielded the highest anthocyanin contents for cinnamon. At 60℃, cinnamon showed the highest level of anthocyanins when methanol and acetone were the solvents, while sumac had the highest anthocyanins level using ethanol as solvent. HPLC results showed ten phenolic acids found in those spices and varied in their concentrations. Gallic acid had the highest level (1642.3 mg/100g) (cloves). Gentisic acid had the lowest level (1.2 mg/100g) in ginger. Also sumac showed the highest level of chlorogenic acid (1528.7 mg/100g). Some acids were not found in some spices, for instance, benzoic acid was not found in coriander, cumin, ginger, green cardamom, cinnamon and sweet laurel.

Identification and characterization of phenolics and terpenoids from ethanolic extracts of Phyllanthus species by HPLC-ESI-QTOF-MS/MS

Phyllanthus species plants are a rich source of phenolics and widely used due to their medicinal properties. A liquid chromatography–tandem mass spectrometry(LC–MS/MS) method was developed using high-pressure liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry(HPLC-ESI-QTOFMS/MS) for the identification and characterization of quercetin, kaempferol, ellagic acid and their derivatives in ethanolic extracts of Phyllanthus species. The chromatographic separation was carried out on Thermo Betasil C_8 column(250 mm×4.5 mm, 5 μm) using 0.1% formic acid in water and 0.1% formic acid in methanol as the mobile phase. The identification of diagnostic fragment ions and optimization of collision energies were carried out using 21 reference standards. Totally 51 compounds were identified which include 21 compounds identified and characterized unambiguously by comparison with their authentic standards and the remaining 30 were tentatively identified and characterized in ethanolic extracts of P. emblica, P. fraternus, P. amarus and P.niruri.

基于HPLC指纹图谱的高酚酸耐盐金银花品种筛选

为了筛选出高酚酸含量且耐盐的金银花品种,在河北、山东、河南不同地点采集金银花品种17个,统一移植到河北省农林科学院滨海农业研究所实验基地的盐渍土(NaCl含量0.1%)上栽培繁殖,盐胁迫14 d后取枝条进行酚酸物质含量检测。采用高效液相色谱法(HPLC)与指纹图谱综合质量评价相结合的方法,以酚酸含量高为目标,对金银花品种进行筛选。结果表明:不同的金银花品种具有共同的特征色谱峰12个,特征峰中以绿原酸、异绿原酸A、异绿原酸C、木犀草苷这4种酚酸物质含量表现较高;不同品种的酚酸物质含量差异较大,其中变异株的绿原酸(15.7 mg/g)以及总酚酸物质含量(20.35 mg/g)均为最高,其次是古郡红(绿叶)(绿原酸含量为11.87 mg/g,总酚酸物质含量为17.05 mg/g),二者指标值均显著>其他品种。变异株和古郡红(绿叶)为本次试验筛选出的适合在盐碱地推广种植的品种。

HPLC法同时检测陕西平利绞股蓝中13种酚酸含量

为建立同时检测陕西平利绞股蓝(Gynostemma pentaphyllum)中13种酚酸含量的测定方法,采用高效液相色谱法(HPLC),利用甲醇水溶液(80%)常温涡旋,超声波提取,色谱柱为Dimensions C18(4.6 mm×150 mm,5μm,P/N:227-30011-07,C/N:18C03033),流动相为甲醇(100%)和磷酸水溶液(0.5%),洗脱方法为梯度洗脱,检测波长为280 nm,柱温为30℃,流速为1.0 mL/min,进样体积为5μL。结果表明,13种酚酸在浓度为5~100 mg/L时线性关系良好,相关系数(R2)≥0.998;在样本加标回收试验中,平均加标回收率为61.8%~131.0%。该测定方法精密度、重复性、稳定性好,适用于陕西平利绞股蓝中酚酸含量的测定。

HPLC同时检测葡萄酒中10种单体酚的方法

本研究建立了用HPLC同时测定葡萄酒中没食子酸、(+)-儿茶素等10种单体酚的方法。色谱条件为:Hibar RT Lichrospher反相C_18柱,流动相A为水:乙酸(98:2),流动相B为乙腈,梯度洗脱,流速为1.0ml/min,检测波长为280nm。在该色谱条件下各个单体酚在30min内得到了良好的分离,各物质的含量与峰面积呈良好的线性关系,且回收率较高,重复性好,且具有定量准确、快速等特点。结果表明,三种葡萄酒均被检测到了这10种单体酚,且它们在红葡萄酒中的含量显著高于白葡萄酒。

HPLC法测定不同葡萄品种成熟过程中单体酚的变化

建立了用HPLC法同时测定黑比诺、梅尔诺、霞多丽葡萄成熟过程中6种酚(没食子酸、安息香酸、儿茶素、丁香酸、芦丁、阿魏酸)的方法,测定了其含量,并比较了其变化差异。结果表明,采用Hibar RT Lichrospher C18柱(250 mm×4．0 mm×5μm),16％乙腈水溶液(含1％乙酸),1．0 mL／min等压洗脱,可以较好地分离测定葡萄中的主要单体酚。在葡萄成熟过程中,红葡萄(梅尔诺和黑比诺)中单体酚含量均高于白葡萄(霞多丽)。儿茶素、芦丁和丁香酸含量随着葡萄成熟逐渐减少,其中儿茶素含量减少幅度最大;安息香酸含量在葡萄成熟前期迅速减少,其后下降缓慢,采收前1周迅速增加;在梅尔诺中没食子酸含量先增加后减少,而在黑比诺和霞多丽中呈先增加后减少再增加的趋势。在供试葡萄品种中,儿茶素含量最高,阿魏酸含量最低,依次为儿茶素>安息香酸>没食子酸>芦丁>丁香酸>阿魏酸。

HPLC法测定薄荷苯酚软膏中苯酚的含量

目的建立HPLC 法测定薄荷苯酚软膏中苯酚的含量.方法：采用超声提取方法处理薄荷苯酚软膏,用HPLC 法进行含量测定.色谱柱：Diamonsil C18 柱（250 mm × 4.6 mm,5 μm）,流动相：甲醇- 水（60∶40）,流速：1.0 mL·min-1,检测波长：271 nm,柱温：45 ℃.结果：苯酚在9 - 360 μg·mL-1 范围内线性关系良好（r = 0.999 9）;样品平均回收率为100.2%,RSD为 0.68%（ n = 6） .结论：本方法简便、准确、重复性好,可用于薄荷苯酚软膏中苯酚的含量测定.

HPLC测定不同采收期球花石斛中酚类成分的含量

目的:比较不同采收期球花石斛叶酚类成分(联苄、菲和芴酮类成分)的含量变化,为球花石斛的合理采收及质量评价提供科学依据。方法:采用高效液相色潜法对不同采收期球化石斛中联苄类化合物石斛酚(gigantol)和杓唇石斛素(moscatil-in),菲类化合物 moscatin 及芴酮类化合物 dendroflorin、dengibsin 和 denchrysan A 的含量进行测定。色谱条件:色谱柱为 Beck-man Coulter ODS(250 mm×4.6 mm,5μm);流动相为乙腈和0,1%二氟醋酸水溶液,梯度洗脱[32%A(0～15 min),32%A→65%A(15～45 min),65%A→32%A(45～55 min)],流速:1.0 mL·min^(-1);检测波长280 nm;柱温25℃。结果:石斛酚、杓唇石斛素、moscatin、dendroflorin、dengibsin 和 denchrysan A 在测定的范围内均表现出良好的线性(r>0.9992);方法的回收率在95.8%～102.4%之间(RSD<2.8%)。结论:通过对同一年12个月不同采收期的球花石斛中联苄、菲和芴酮类成分的含量测定,证明方法灵敏,准确可靠,重复性好。总酚类成分的含量在4,5月份和10,11月份相对高于其他月份。

苹果树皮中12种酚类化合物的HPLC测定方法

【目的】为了建立高效液相色谱法(HPLC)同时测定苹果树皮中没食子酸、儿茶酚、儿茶素、绿原酸、咖啡酸、表儿茶素、香豆酸、阿魏酸、芦丁、根皮苷、槲皮素、根皮素12种酚类化合物的方法,【方法】选用Eclipse XDB C18色谱柱(150 mm×4.6 mm i.d,5μm),以甲醇和1%乙酸溶液作为流动相进行梯度洗脱,利用VWD检测器,在各种酚类物质的最优吸收波长下进行检测,【结果】得到12种酚类化合物的线性回归方程,相关系数均在0.999以上,检出限(3RSN)分别为0.02、0.02、0.03、0.01、0.02、0.03、0.01、0.01、0.03、0.03、0.05和0.01 mg.L-1。色谱测定方法和包含样品处理方法的加标回收率分别在56.54%～98.83%和90.95%～100.90%。【结论】整个提取和分析测定流程简便、快速、准确,适合苹果树皮中酚类化合物的分析测定。

骨肽注射液中苯酚含量的RP-HPLC法测定

目的:建立高效液相色谱法测定骨肽注射液中苯酚的含量。方法:采用 ZORBAX C_(18)柱(4.6mm×250mm,5μm);流动相为甲醇-水(25:75);流速1.0 mL·mm^(-1);检测波长:270nm;进样量:10μL。结果:方法的线性范围为0.001-1.0mg·mL^(-1)(r=0.9998);检测限为0.2μg·mL^(-1);精密度 RSD 为0.23%(n=7);高、中、低浓度回收率(n=3)分别为98.8%(RSD为0.5%)、99.2%(RSD 为0.2%,)、99.0%(RSD 为0.3%)。结论:本法具有专属、灵敏、快速、准确的特点,可用于骨肽注射液中的苯酚测定。

HPLC法测定苹果浓缩汁中的多酚类物质

利用高效液相色谱法(HPLC)分析了儿茶素、绿原酸、咖啡酸、表儿茶素、香豆酸、阿魏酸等六种酚类物质在鲁加1号和鲁加5号苹果浓缩汁中的的含量。色谱条件:色谱柱为EclipseXDB-C18色谱柱(150mm×4.6mmid,5μm),以甲醇-1%乙酸水溶液作为流动相进行梯度洗脱。柱温30℃,流速为1.0ml/min,检测波长为280nm。在此色谱条件下,各组分均得到很好的分离。经测定鲁加1号苹果浓缩汁中含有儿茶素、绿原酸、咖啡酸、表儿茶素、香豆酸、阿魏酸等六种酚类物质;鲁加5号苹果浓缩汁中含有较少的绿原酸、咖啡酸、表儿茶素、香豆酸、阿魏酸等五种酚类物质。

金钗石斛中黄酮类和酚类成分HPLC指纹图谱研究

目的:建立金钗石斛药材中黄酮类和酚类成分的HPLC指纹图谱。方法:采用Phenomenex prodigy ODS(3)C18色谱柱(250mm×4.6mm,5μm),乙腈-0.1%乙酸溶液梯度洗脱,柱温25℃,流速1.0mL/min,检测波长254nm。结果:该方法可使金钗石斛中黄酮类和酚类成分得到较好的分离,并根据检测结果选定了10个共有指纹峰。结论:该法操作简便、重现性好,可作为金钗石斛药材质量控制的方法。

用HPLC测定苹果酒中的酚类物质

确立了用HPLC测定苹果酒中酚类物质的方法 ,各酚的回收率均在 85 %之上 。

用HPLC法测定干红葡萄酒中的酚类物质

建立了用HPLC测定干红葡萄酒中 11种酚类物质的方法。该法具有定量准确 ,快速的特点。在实验条件下 ,各物质的含量与峰面枳呈良好的线性关系 。

“紫娟”晒青绿茶色素的HPLC-ESI-MS分离鉴定及其稳定性研究

研究"紫娟"晒青绿茶主要呈色成分及其稳定性。采用酸性甲醇-水和水分别提取"紫娟"晒青绿茶色素,利用HPLC-ESI-MS/MS方法对其主要色素进行了分离鉴定。结果显示,"紫娟"晒青绿茶含有14种花色素苷、24种黄酮、8种酚类以及3种生物碱。主要的花色苷类物质为天竺葵素-3-葡萄糖苷或天竺葵素-3-半乳糖苷、天竺葵素-3-(6-香豆酰基)-葡萄糖苷、飞燕草素-3-葡萄糖苷、飞燕草素-3-芸香糖苷、飞燕草素-3-O-(6-香豆酰基)-葡萄糖苷、Prodelphinidin B-2(or 4)3′-O-gallate、Procyanidin B-2(or 4)3′-O-gallate、Petunidin-3-glc-4-vinylphenol和Malvidin-3-glc-4-vinylepicatechin等。酸性甲醇-水提取的"紫娟"晒青绿茶红色素水溶性好,颜色随pH值变化而变化。酸性条件下该色素对光、低温有很好的稳定性,耐氧化性与耐还原性较强。Cu2+、Fe3+使色素稳定性变差。明确了"紫娟"晒青绿茶呈现紫色的主要物质基础。

HPLC法同时测定苍耳类药材中酚酸及蒽醌类成分的含量

建立HPLC同时测定苍耳子、苍耳草药材中绿原酸、新绿原酸、原儿茶醛、原儿茶酸、隐绿原酸、咖啡酸、1,3-二咖啡酰奎宁酸、阿魏酸、3,4-二咖啡酰奎宁酸、3,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸、芦荟大黄素、大黄素及大黄酚含量的方法。采用Agilent ZORBAX SB-C18色谱柱（250×4.6 mm,5μm）,以甲醇（A）-0.1%甲酸水溶液（B）为流动相,梯度洗脱,流速1.0 m L/min,柱温35℃,检测波长254 nm,进样量10μL。14种成分的浓度与峰面积的线性关系良好（r〉0.9931）;加样回收率为97.18%~101.09%。苍耳类药材中酚酸和蒽醌类14种成分的含量存在差异,其中绿元素、原儿茶酸含量较高,芦荟大黄素、大黄素、大黄酚含量较低。该方法简单、准确,重现性较好,可用于苍耳类药材内在质量的评价和控制。

HPLC-DAD法同时测定金银花中7种多酚类物质

建立了高效液相色谱-二极管阵列法（HPLC—DAD）同时测定金银花中绿原酸、咖啡酸、阿魏酸、香豆酸、肉桂酸、芦丁和木犀草苷的方法。采用岛津Shim—packVP—ODSC镕（250mm×4．6mm，5μm）色谱柱，2％甲酸-2％甲酸甲醇溶液为流动相梯度洗脱，柱温35℃，流速0．7mL/min。结果表明：绿原酸、咖啡酸、阿魏酸、香豆酸、肉桂酸、芦丁和木犀单苷分别在1．04-520、1．02-510、1．59～530、1．26～630、1．89～560、1．62～540、0．29～58mg／L范围内线性关系良好（r值范围为0．9964-0．9999），检出限（RSN=3）为0．001～0．078mg／L，精密度RSD小于3．27％，平均回收率为93．42％～102．53％。本方法操作简单，灵敏度高、重现性好，可用于金银花中7种多酚类物质的定量分析和质量监控。