超高效液相色谱-飞行时间-串联质谱法对3种蓝靛果忍冬果实中花青苷的比较分析

利用超高效液相色谱-飞行时间-串联质谱技术,对伊利亚特、别列里和格尔达3种蓝靛果忍冬果实中的花青苷进行分析。结果表明,在3种蓝靛果忍冬果实中共鉴定出24种花青苷,包含14种矢车菊素类花青苷、6种芍药素类花青苷、2种飞燕草素类花青苷和2种天竺葵素类花青苷,其中9种花青苷为首次在蓝靛果忍冬中被发现。3种蓝靛果忍冬果实中主要的花青苷均为矢车菊素-3-葡萄糖苷、芍药素-3-葡萄糖苷、矢车菊素-3-芸香糖苷和矢车菊素-3,5-双葡萄糖苷。比较分析表明,3种蓝靛果忍冬果实中花青苷的组成不同,分别在伊利亚特、别列里和格尔达果实中检测到24、23种和12种花青苷;此外,伊利亚特和别列里果实中花青苷的相对含量明显高于格尔达。本实验为进一步研究花青苷在蓝靛果忍冬中合成与积累的分子遗传机制以及选育富含花青苷的蓝靛果忍冬新品种奠定基础。     

黑果腺肋花楸花色苷提取工艺优化及其抗氧化活性和组成鉴定

目的：优化超声波辅助提取黑果腺肋花楸花色苷的条件,测定花色苷的抗氧化能力,鉴定黑果腺肋花楸花色苷提取物的组成成分。方法：采用响应面法优化花色苷提取条件,通过1,1-二苯基-2-三硝基苯肼自由基清除能力、2,2’-联氨-双（3-乙基苯并噻唑啉-6-磺酸）二铵盐自由基清除能力及Fe3＋还原能力方法评价其抗氧化能力,高效液相色谱-质谱联用法进行成分分析。结果表明：提取温度60?℃、超声功率100?W、料液比1∶30（g/mL）、超声时间31?min、乙醇体积分数64%和pH?2.0时为最优提取条件,此时黑果腺肋花楸花色苷含量可达（3.61±0.01）mg/g。体外抗氧化实验表明,在相同质量浓度条件下,黑果腺肋花楸花色苷提取物的抗氧化活性明显高于VC。组分鉴定表明黑果腺肋花楸花色苷提取物中共有6?种花色苷,其中矢车菊-己糖苷二聚体和矢车菊-3,5-二己糖苷为新检测出的2?种花色苷。     

黑果腺肋花楸挥发油化学成分及总黄酮含量分析研究

目的研究黑果腺肋花楸挥发性化学成分的组成及质量分数,分析其总黄酮含量。方法采用超声波提取法提取黑果腺肋花楸果实中的挥发油和总黄酮,挥发油的化学成分通过气相色谱-质谱法(gas chromatography-mass spectrometer, GC-MS)进行分析鉴定,各挥发性化学成分的质量分数采用色谱峰面积归一化法计算。以芦丁为标准品,用紫外可见分光光度仪进行总黄酮含量的分析测定。结果黑果腺肋花楸挥发油经GC-MS分析,共鉴定出36个挥发性化学成分,占挥发油色谱总峰面积的84.14%。芦丁标准品呈现良好线性关系(r~2=0.9997),总黄酮类化合物的平均含量为0.233%,标准偏差(standard deviation,SD)为0.01291,相对标准偏差(relative standard deviation, RSD)为5.53%。结论黑果腺肋花楸中的挥发性成分以烷烃类、酯类、羧酸类、醇类几类化合物为主,其果实中总黄酮类化合物的含量较低。     

植物乳杆菌发酵黑果腺肋花楸果汁的工艺优化

本文以黑果腺肋花楸为原料,利用植物乳杆菌C8-1对其发酵,研制具有特殊果香风味和营养价值的黑果腺肋花楸发酵饮料。采用单因素试验和正交试验对植物乳杆菌发酵黑果腺肋花楸果汁进行条件优化,以菌种接种量、发酵温度、发酵时间和料液比为影响因素,总酸含量作为指标,进行正交试验分析。结果表明,黑果腺肋花楸发酵饮料最佳发酵条件为:植物乳杆菌接种量9 lg CFU/mL,发酵温度35 ℃,发酵时间22 h,黑果腺肋花楸料液比为1:3(v:v),在此条件下,总酸含量达到6.60 mg/mL;发酵后的黑果腺肋花楸果汁中的总酚含量增加10.51%,总酸含量增加74.60%,还原糖含量降低4.41%,可溶性固形物含量降低6.15%;对色泽研究可知,发酵对黑果腺肋花楸果汁的亮度、红色色调和黄色色调均有增强作用;抗氧化实验表明,经发酵处理的黑果腺肋花楸果汁对DPPH自由基清除能力、OH自由基清除能力和总还原能力均极显著高于鲜榨的黑果腺肋花楸果汁(p<0.01)。感官评分较鲜榨果汁有明显提高。结果表明,植物乳杆菌C8-1可以在黑果腺肋花楸果汁中进行正常的生长代谢活动,并能够提高黑果腺肋花楸果汁的品质和抗氧化能力。     

五种寒地果树果实的多酚含量、抗氧化活性及抗α-淀粉酶活性分析

为了探究寒地果树资源果实中的营养价值及体外活性,本试验测定了软枣猕猴桃、黑果腺肋花楸、草原樱桃、白穗醋栗和蔓越莓五种果实的总酚、总花色苷含量,比较了其抗氧化及抗α-淀粉酶活性,并进行了相关性分析,结果表明:不同寒地果实的总酚、总花色苷含量之间存在显著差异(P<0.05)。总酚含量为13.21~66.43 mg GAE/100 g,其中黑果腺肋花楸含量最高,白穗醋栗次之;总花色苷含量为0.15~46.45 mg C3G/100 g,黑果腺肋花楸含量最高,草原樱桃次之。另外5种果实的抗氧化能力也有所差异。用DPPH法、ABTS法、ORAC法、FRAP法测得白穗醋栗的抗氧化活性最强。用高通量淀粉浊度法测得白穗醋栗的抗α-淀粉酶活性最强,IC₅₀为0.122 mg/mL,高于阿卡波糖(0.16 mg/mL),其AE值为2034.18 mmol AE/g,高于阿卡波糖(1550 mmoL AE/g),且是其他果实的42~127倍。相关性分析发现五种寒地果实的总酚、总花色苷含量和抗氧化活性之间存在极显著正相关(P<0.01),而与抗α-淀粉酶活性之间无显著性相关关系。     

黑果腺肋花楸多酚的抑菌效果及对α-淀粉酶活性的抑制作用

为探究黑果腺肋花楸多酚的抑菌效果及对α-淀粉酶活性的抑制作用,采用滤纸片法测定黑果腺肋花楸多酚对大肠杆菌、金黄色葡萄球菌、黄曲霉、烟曲霉、米根霉、岛青霉、康宁木霉的抑制效果,并考察温度和pH值对黑果腺肋花楸多酚抑制金黄色葡萄球菌和烟曲霉活性的影响。通过建立动力学模型研究黑果腺肋花楸多酚对α-淀粉酶活性的抑制作用。结果表明：黑果腺肋花楸多酚对7 种供试菌均有一定的抑制作用,对烟曲霉的抑制效果最强,最低抑菌浓度为0.171 9 mg/mL;黑果腺肋花楸多酚对金黄色葡萄球菌的抑菌效果在pH 5.0条件下最强;而对于烟曲霉,pH值的变化对抑菌效果有破坏作用。黑果腺肋花楸多酚对α-淀粉酶活性具有显著的抑制作用,并在0.10～0.70 mg/mL质量浓度范围内,对α-淀粉酶的抑制活性强于阳性对照阿卡波糖,该抑制作用属于可逆非竞争性抑制,抑制常数为0.193 2 mg/mL。在202 nm波长处,黑果腺肋花楸多酚能够引起α-淀粉酶紫外吸收的增强和吸收峰的红移。通过深入开发研究,黑果腺肋花楸多酚可作为具有辅助防腐抑菌功能的α-淀粉酶抑制剂。     

响应面法优化黑果腺肋花楸酒酿造工艺及成分研究

以黑果腺肋花楸(Aronia melanocarpa)为原料，通过单因素实验和响应面法优化提取工艺，确定黑果腺肋花楸酒最优工艺条件为：发酵液含糖量21%、发酵温度25.5℃、发酵时间7 d以及酵母接种量0.8 g/L，此时酒精度为8.5%vol，多酚类物质含量为3.580 mg/mL。MS/MS结果显示黑果腺肋花楸中多酚类物质主要有矢车菊-3-O-半乳糖苷、矢车菊-3-O-葡萄糖苷、矢车菊-3-O-阿拉伯糖苷、矢车菊-3-O-木糖苷、槲皮素-3-O-芸香糖苷、金丝桃苷、槲皮素-3-O-葡萄糖苷，其中矢车菊-3-O-半乳糖苷含量最高。气相色谱结果表明，该果酒主要包含12种香气成分,苯甲醇含量最高。     

高效液相色谱法测定黑果腺肋花楸提取物中氯化矢车菊素的含量及抗氧化性研究

建立黑果腺肋花楸提取物中氯化矢车菊素的含量测定方法,并优化黑果腺肋花楸提取物供试品的制备方法。以氯化矢车菊素的含量为指标,采用高效液相色谱法测定黑果腺肋花楸提取物中氯化矢车菊素的含量,色谱柱:伊利特ODS2 C18(4.6 mm×250 mm,5μm);流动相以甲醇-0.2%磷酸水溶液(v/v);进样量20μL;检测波长276 nm;柱温30℃。在建立含量测定方法的同时探索黑果腺肋花楸提取物水解提取氯化矢车菊素的工艺条件。试验结果表明氯化矢车菊素在3.6～36.0μg.mL~(-1)(r=0.9995)线性关系良好;氯化矢车菊素的最佳水解提取工艺为:水解温度100℃、水解时间2 h、溶剂酸浓度3%盐酸甲醇。在优化水解提取条件下,氯化矢车菊素的含量为1.8570 mg.g~(-1)。该含量测定方法具有良好的精密度,结果准确可靠;优化的提取方法快速、简单,为黑果腺肋花楸果的开发利用提供了依据。体外抗氧化活性的试验结果表明,黑果腺肋花楸提取物对DPPH自由基具有清除活性,且随着其质量浓度的增加,清除活性明显增强;利用FRAP法和钼酸铵法测得黑果腺肋花楸提取物具有一定的抗氧化活性。     

超声提取-离子色谱-脉冲积分安培检测法检测黑果腺肋花楸果中的8种单糖和双糖

目的建立超声提取-离子色谱-脉冲积分安培检测法测定黑果腺肋花楸果中的8种单糖和双糖的方法。方法采用水浸提超声提取及膜过滤法前处理黑果腺肋花楸果样品,以Dionex CarboPac PA10 Analytical Column(4 mm×250 mm)阴离子交换色谱柱为分析柱,以Dionex CarboPac PA10 Guard Column(4 mm×50 mm)为保护柱,以NaOH与CH3COONa的组合溶液作为流动相,采用梯度淋洗分离程序,脉冲积分安培方法检测。结果 8种单糖和双糖的检出限分别为5、8、3、4、7、2、5、2μg/L,且均具有较宽的线性范围(0.05～20 mg/L)。黑果腺肋花楸果中单糖和双糖测定的相对标准偏差在0.96%～6.12%之间,8种单糖和双糖的加标回收率达到90.5%～102.1%。结论本方法同时检测黑果腺肋花楸果中的单糖和双糖,其方法简便快捷、分离效果好、基体干扰少、无需衍生、灵敏度高,适用于黑果腺肋花楸果中的常见单糖和双糖组分的分析。     

黑果腺肋花楸中花色苷超声辅助提取工艺优化研究

目的优化超声辅助提取黑果腺肋花楸中花色苷工艺的方法。方法在单因素的基础上,研究提取温度、液料比和提取时间对花色苷提取率的影响,并利用响应面法对花色苷的提取条件进行优化。结果黑果腺肋花楸中花色苷提取的最佳工艺条件为:乙醇溶液(含0.5%的乙酸)浓度为40%,提取温度为43℃,超声时间为23 min,料液比为89:1(V:m),此条件下,黑果腺肋花楸中花色苷提取得率为(0.79±0.010)g/100g。结论本方法可以快速有效地提取黑果腺肋花楸中的花色苷。     

Berry fruit teas: Phenolic composition and cytotoxic activity

A detailed phenolic composition analysis of dry chokeberry, bilberry and black currant fruit teas prepared on the most common ways — decoction and infusion, was performed, together with evaluation of cytotoxic activity. The most abundant in phenolics were chokeberry teas, followed by bilberry and black currant teas, while the highest anthocyanin amount was determined in bilberry samples. LC/DAD/MS method was used for identification of 17 anthocyanins, 11 flavonoids and 4 hydroxycinnamic acid derivatives. Quantitative analysis of investigated teas was carried out by HPLC analysis, and major phenolic compounds in berry fruit teas were chlorogenic acids, quercetin glycosides and anthocyanin glycosides. Berry teas were investigated for cytotoxic activity against cervix carcinoma, melanoma, colon, and chronic myelogenous leukemia cell lines, and chokeberry decoction was the most effective against all cell lines. According to the results obtained in this study, pure chokeberry, bilberry and black currant teas could be valuable sources of flavonoids and anthocyanins.     

Simultaneous quantification of flavonoids and phenolic acids in Herba Scutellariae barbatae and its confused plants by high performance liquid chromatography-tandem mass spectrometry

A sensitive and selective high performance liquid chromatography–tandem mass spectrometric method (HPLC–MS/MS) has been developed for simultaneous analysis of 11 components (7 flavonoids and 4 phenolic acids) in Herba Scutellariae barbatae. Simultaneous separation of these 11 compounds was achieved on a C₁₈ analytical column with gradient elution of methanol and 0.1‰ acetic acid (v/v) at a flow rate of 0.8 mL/min. The precursor and product ions of analytes were monitored on a hybrid quadrupole linear ion trap mass spectrometer equipped with a turbo ion spray interface in negative mode using multiple-reaction monitoring (MRM). Full validation of the assay was carried out including linearity, precision, accuracy, limits of detection and quantification. The results demonstrated that the method developed was reliable, rapid and specific. Then the method was successfully applied to differentiate 18 batches of Herba Scutellariae barbatae samples and 3 batches of its confused plants from different sources.     

超高效液相色谱-四极杆/轨道阱高分辨质谱法快速筛查功能性保健食品中19种特征性成分

目的 建立超高效液相色谱-四极杆/轨道阱高分辨质谱法(UPLC-Q/ORBITRAP HRMS)快速筛查功能性保健食品中19种特征性成分的分析方法。方法 以甲醇为提取液,经超声、高速离心、过滤,通过XDB-C₁₈色谱柱分离,以0.1%甲酸溶液(含0.02 mol/L醋酸铵)和甲醇作为流动相洗脱,选择正负离子实时切换和一级离子/自动触发二级离子扫描模式,利用保留时间、一级离子和二级离子信息建立质谱数据库,完成定性分析,利用一级分子离子峰完成定量测定。结果 19种特征性成分的实测离子质荷比与理论值相对偏差均小于8.10×10^_-6,19种特征性成分在其相应的线性范围内存在线性关系,相关系数均大于0.99,检出限为0.25 ~ 1.00 mg/kg,加标回收率为75.16% ~ 97.93%,精密度为1.09% ~ 4.86%。结论 该方法抗干扰能力强,准确度高,可用于保健食品中19种特征性成分的快速筛查,同时结合产品标签标识,鉴别其使用原料的真伪性。     

杜仲雄花化学成分的液相色谱-电喷雾三重四极杆飞行时间质谱分析

采用液相色谱-电喷雾三重四极杆飞行时间质谱法对杜仲雄花中化学成分进行分析。用反相C18色谱柱,以0.1%甲酸溶液（A）-乙腈（B）为流动相进行梯度洗脱;采用电喷雾离子源负离子扫描方式对样品进行分析。根据高分辨质谱提供的准分子离子和碎片离子的精确分子质量信息,结合标准品对照与参考相关文献数据,共鉴定出32?种化学成分,包括2?种木脂素类、9?种环烯醚萜类、8?种苯丙素类、12?种黄酮类和1?种酚苷类成分。上述结果可为探究杜仲雄花的药效物质基础及深度开发利用提供基础资料。     

超高效液相色谱-四级杆/静电场轨道阱高分辨质谱联用快速测定水产品及干制水产品制品中的116种农药和24种生物毒素残留

采用了一种新型脂肪吸附剂去除样品基质中脂肪和磷脂等杂质的干扰,利用超高效液相色谱-四级杆/静电场轨道阱质谱法(UPLC-Q Exactive Orbitrap MS)的同时定性定量功能,建立了水产品及干制水产品制品中116种农药和24种生物毒素残留量的检测方法。样品前处理采用QuEChERS方法,经乙腈/水(90:10,V/V)溶液提取,高效基质脂肪吸附剂(EMR-Lipid)净化,平行定量浓缩仪浓缩,C18色谱柱分离,5 mmol甲酸水溶液(含0.1%甲酸铵)和5 mmol甲酸甲醇溶液(含0.1%甲酸铵)梯度洗脱;质谱数据采集使用Q Exactive高分辨质谱的Full MS/dd-MS2监测模式,以Full MS一级质谱全扫描提取母离子精确质量数所得的色谱峰面积进行定量,以保留时间和dd-MS2数据依赖子离子扫描所得的二级子离子质谱图进行定性确证。140种目标物的精确质量数偏差不大于3×10^-6,浓度与母离子峰面积的线性关系良好,相关系数≥0.991,检出限为0.02~0.4μg/kg。基质加标回收率在70.1%~109.1%之间,相对标准偏差(RSD)为1.0%~14.1%。该方法学结果满足GB/T 27417-2017《合格评定化学分析方法确认和验证指南》的技术要求。本方法具有操作简单快捷、灵敏度高等优点。     

Optimization on anthocyanins extraction from wine grape skins using orthogonal test design

To optimize the extraction of anthocyanins from grape skins, orthogonal test with 4 factors and 3 levels was designed and the analysis of extracts was conducted using high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). The results showed that the optimal extraction for obtaining the highest amounts of anthocyanins was carried out by formic acid/methanol solvent (5/95, v/v), with ultrasonication for 10 min, extraction temperature at 25°C, and extraction time for 1.5 hr. The precision and accuracy of this method were established. This work will provide a basis for further study involving in anthocyanins of grape berries.     

Analysis of water-soluble azo dyes in soft drinks by high resolution UPLC-MS

An UPLC-Orbitrap MS system was exploited to develop and validate a method for the simultaneous determination of 11 water-soluble azo dyes (Acid Yellow 17, Acid Red 14, Acid Red 26, Acid Red 73, Acid Orange 52, Acid Orange 7, Acid Orange 12, Acid Yellow 36, Acid Orange 5, Acid Red 88 and Acid Red 9) in soft drinks. Three pairs of isomers and four disulphonated azo dyes were among a total of 11 water-soluble azo dyes obtained and purified using an SPE cartridge. They were well separated using optimized UPLC conditions with a RP18 column and a MS detector with a compatible mobile phase system. All these dyes were detected by the Orbitrap XL mass spectrometer in negative ion mode. HCD tandem MS fragment ions are first reported in this paper, and these fragment ions can be used for identification of isomers of azo dyes. According to SANCO/10684/2009, one quasi-molecular ion in full scan mode as quantification ion and one or two HCD tandem MS fragment ions as identification ions are required for compound confirmation. Matrix-matched calibration was employed for quantification. The linear matrix-matched calibration for the 11 dyes was in the range 5-200?ng?g(-1) with correlation coefficients (r) of 0.9939-0.9988. Recoveries were 68.9-110.8% with coefficients of variation of 0.9-12.0%. Depending on the dye and matrix involved, the LODs were between 1.0 and 3.2?ng?g(-1) and LOQs were between 5.2 and 9.8?ng?g(-1).     

黑果腺肋花楸果中总花色苷含量测定方法的比较

分别探讨摩尔吸光系数和对照品对pH示差法和高效液相色谱（high performance liquid chromatography,HPLC）法测定黑果腺肋花楸果中总花色苷含量的影响,并通过比较优化前后pH示差法和HPLC法的测定结果,确定黑果腺肋花楸果中总花色苷含量的最佳测定方法。采用t检验将优化前后测定方法测得的总花色苷含量进行比较。结果表明,摩尔吸光系数和对照品的选择均会影响测定结果,其中以黑果腺肋花楸果中高含量的矢车菊素-3-O-半乳糖苷为标准的pH示差法和混合标样HPLC法均优于传统方法（以矢车菊素-3-O-葡萄糖苷为标准）（P＜0.05）;以矢车菊素-3-O-半乳糖苷为标准的单标样HPLC法与混合标样HPLC法测定结果比较无显著差异（P＞0.05）;HPLC法测定结果均高于pH示差法。结果表明,混合标样HPLC法为测定黑果腺肋花楸果中总花色苷含量最准确的方法,但成本较高;矢车菊素-3-O-半乳糖苷单标样HPLC法简单、准确而经济,同样适用于黑果腺肋花楸果中总花色苷的含量测定。     

液相色谱-四极杆飞行时间质谱用于快速筛查动物组织中多种激素残留的方法研究和应用

建立了一种采用液相色谱-四极杆飞行时间质谱（LC-Q-TOF/MS）用于快速筛查动物组织中多种激素残留的检测方法。样品通过乙腈和乙酸乙酯分步提取,提取液经增强型脂质去除吸附剂（EMR）净化,盐析后经N-丙基乙二胺（PSA）和官能化聚苯乙烯/二乙烯苯（PEP-2）进一步净化,以0.1%甲酸水?0.1%甲酸乙睛作为流动相进行梯度洗脱,在EclipsePlus-C₁₈（3.0 mm×100 mm,1.8 μm）色谱柱上进行分离,在Q-TOF/MS正离子全扫描模式下采集质谱数据,以保留时间、精确分子质量数、同位素丰度比和二级特征碎片离子定性,待测物准分子离子峰面积定量。结果表明,所有药物在各自浓度范围内线性良好,相关系数大于0.995,在10、50、100 μg/kg添加水平下,平均回收率在70.3%～116.2%之间,相对标准偏差为0.87%～8.97%,方法检测限为1～10 μg/kg,定量限为3～30 μg/kg。该方法通过构建一级精确质量数据库和二级谱库,结合保留时间、精确分子质量数、同位素丰度比和二级特征碎片离子,实现对目标化合物快速筛查和确证,具有简单快速、高灵敏度,高选择性和良好精密度的优势,适用于动物组织多种激素的快速筛查和定量测定。     

Analysis of synthetic antioxidants and preservatives in edible vegetable oil by HPLC/TOF-MS

The application of high performance liquid chromatography time-of-flight mass spectrometry (HPLC/TOF-MS) for the qualitation and quantitation of 11 synthetic antioxidants and preservatives in edible vegetable oil samples is reported here. The qualitation by HPLC/TOF-MS is accomplished with the accurate mass of the deprotonated molecules [M−H]⁻, along with the accurate mass of their main fragment ion. In order to obtain sufficient sensitivity for quantitation purposes (using deprotonated molecule), segment programme of fragmentor voltage is designed in negative ion mode. The mass accuracy typically obtained is routinely better than 5 ppm. The 11 compounds behave linearly in the 0.05–5.0 mg/kg concentration range, with correlation coefficient >0.997. The recoveries at the tested concentrations of 0.1–2.0 mg/kg are 65.8–106.9%, with coefficients of variation <8.1%. The method illustrated is suitable for routine qualitative and quantitative analyses of synthetic antioxidants and preservatives in edible vegetable oils.