PTR-ToF-MS结合自动采样系统和食品研究定制数据分析：生物过程监测，筛选和鼻空间分析
Summary
Proton Transfer Reaction Time of Flight Mass Spectrometry allows high-sensitivity, rapid and non-invasive analysis of volatile organic compounds. To demonstrate its potential, we give three examples: lactic acid fermentation of yogurt (on-line bioprocess monitoring), different apple genotypes (large-scale screening), and retronasal space after drinking coffee (nosespace analysis).
Abstract
质子转移反应（PTR），结合飞行时间（ToF）质谱（MS）是基于直接注入质谱（DIMS）技术的基于化学电离的分析方法。这些技术可以快速测定挥发性有机化合物（VOC），确保高灵敏度和准确度。一般来说，PTR-MS既不需要样品制备也不需要样品破坏，可以对样品进行实时和非侵入性分析。 PTR-MS在许多领域得到开发，从环境和大气化学到医学和生物科学。最近，我们开发了一种基于PTR-ToF-MS联合自动采样器和定制数据分析工具的方法，以增加自动化程度，从而提高该技术的潜力。这种方法允许我们监测生物过程（ 例如酶促氧化，酒精发酵），筛选大样品组例如不同的来源，整个胚泡），并分析几种实验模式（ 例如 ，给定成分的不同浓度，特定技术参数的不同强度）在VOC含量方面。在这里，我们报告了实验方案，例示了我们的方法的不同可能的应用： 即酸奶发酵过程中释放的VOCs（在线生物过程监测），与不同苹果品种（大规模筛选）相关的VOC的监测， ，以及咖啡饮用期间维生素VOC释放的体内研究（鼻孔空间分析）。
Introduction
直接注入质谱（DIMS）技术代表了一类分析仪器方法，具有高灵敏度和鲁棒性的相当大的质量和时间分辨率，可以快速检测和定量挥发性有机化合物（VOC） 1 。这些工具方法包括MS-e-noses，大气压力化学电离质谱（APCI-MS），质子转移反应质谱（PTR-MS）和选定离子流管质谱（ SIFT-MS） 1 。每种方法的优点和缺点取决于：样品注入的种类，前体离子的来源和控制，电离过程的控制以及质量分析仪1,2 。
质子转移反应质谱（PTR-MS）是二十多年前开发的，用于实时监测和检测在空气中的最低挥发性有机化合物（VOC）的低检测限（通常为几ppbv，体积百分之十）。目前PTR-MS的应用范围从医疗应用，食品控制到环境研究5,6 。该技术的主要特点是：快速连续测量的可能性，前体离子的强烈和纯净的来源，以及控制电离条件（压力，温度和漂移电压）的可能性。这些功能允许将多功能用途与高标准化组合1,4 。事实上，该方法是基于水合氢离子（H 3 O + ）的反应，其在大多数挥发性化合物（特别是以质子亲和力高于水的特征）中诱导非解离质子转移，质子化中性化合物（M）根据反应：H 3 O + + M→H 2 O + MH + 。与其他技术相比， 例如 APCI-MS，前体离子产生和样品电离分成两个不同的仪器室（ 图1中给出了PTR-MS仪器的示意图 ）。中空阴极离子源中的水蒸汽放电产生一束水合氢离子。在此阶段之后，离子穿过漂移管，其中VOC的电离发生7 。离子然后进入脉冲提取部分并加速到TOF部分。通过飞行时间，可以确定离子8的质荷比。每个提取脉冲导致所选m / z范围的完整质谱8 。离子光谱由快速数据采集系统7记录。一个完整的频谱是典型的在一秒钟内获得，尽管可以根据信号噪声水平实现更高的时间分辨率，并且即使没有校准，也可以提供VOC顶空浓度的定量估计9,10 。
图1： PTR-MS的示意图。 PTR-MS仪器的示意图。 HC：具有中空阴极的外部离子源; SD：源漂移;六，文丘里型入口; EM，电子倍增器; FC1-2，流量控制器。转载自Boschetti 等人的许可7 。 请点击此处查看此图的较大版本。
与食品基质相关的挥发性有机化合物在食品科学和技术方面具有突出的兴趣，因为它们在与气味和风味感知相关的生物现象的分子基础中起重要作用，因此在食品接受度方面。因此，我们对VOC的实时和非侵入性检测的兴趣主要涉及食品的感官品质。此外，如果我们考虑通过释放的VOC 13检测腐败和致病微生物的可能性，和/或监测挥发性有机化合物作为标记物机翼技术过程（ 例如热处理中的美拉德副产品） 14 ，VOC识别和定量是食品质量管理中感兴趣的领域。几种近来PTR-MS技术用于快速监测和定量食品基质中VOCs的应用证明了这些分析方法的广泛应用（ 表1 ）。
| 食物矩阵 | 种类适用 | 简要描述;简介 | 参考 |
| 牛油 | 筛选/表征 | 欧洲的地理起源 | 15 |
| 酸奶 | 生物过程监控 | 乳酸过程中的进化心理状态 | 16 |
| 谷物酒吧 | 体内测量 | 在食用不同糖成分的谷物酒的食物空间 | 17 |
| 液体模型系统 | 模拟口服条件 | 口腔压力和口腔状况的评估 | 18 |
| 苹果 | 体内测量 | 消费苹果中的空白具有不同的遗传，结构和物理化学参数 | 19 |
| 咖啡 | 筛选/表征 | 专业咖啡的区别 | 20 |
| 葡萄必须 | 筛选/表征 | 烹饪过程的影响 | 21 |
| 调味糖果 | 体内测量 | 决定使用不同的小组成员直接质谱法 | 22 |
| 火腿 | 筛选/表征 | 养猪系统的效果 | 23 |
| 面包 | 模拟口服条件 | 在咀嚼期间模拟面包香气 | 24 |
| 牛奶 | 筛选/表征 | 监测牛奶中光氧化诱导的动态变化 | 25 |
| 咖啡 | 筛选/表征 | 来自不同地理起源的烤咖啡的多样性 | 26 |
| 面包 | 生物过程监控 | 不同酵母起泡器在酒精发酵过程中的作用 | 27 |
| 咖啡 | 体内测量 | 在消费不同的焙炒咖啡制品期间的空气 | 28 |
| 筛选/表征 | 生产位置，生产系统和品种的影响 | 29 | |
| 面包 | 生物过程监控 | 酒精发酵过程中面粉，酵母及其相互作用的影响 | 三十 |
| 蘑菇 | 筛选/表征 | 干燥的猪笼草蘑菇的保质期 | 31 |
| 酸奶 | 生物过程监控 | 不同发酵剂培养物在乳酸发酵过程中的作用 | 32 |
| 苹果 | 筛选/表征 | 苹果种质资源多样性 | 33 |
| 咖啡 | 筛选/表征 | 跟踪咖啡来源 | 34 |
| 咖啡 | 体内测量 | 组合a动态感官方法和体内鼻子空间分析来了解咖啡感觉 | 35 |
表1：使用PTR-ToF-MS在食品行业的科学研究清单。使用基于PTR的方法在食品相关实验中监测VOC含量的科学研究非详尽列表。
在最近的研究中，我们报告了PTR-ToF-MS的应用以及自动采样系统和定制的数据分析工具，以提高采样自动化和可靠性，从而提高这种技术的潜力7,10,13。这允许我们在VOC含量方面筛选大样本集（ 例如具有许多重复的不同来源的食物，整个胚泡），以分析几种实验模式对VOC释放的影响（ 例如不同浓度特定技术参数的不同强度），并监测与给定生物过程相关的VOC（ 如酶氧化，酒精发酵）。在这里，为了说明PTR-ToF-MS在农业食品行业的潜力，我们提出了三种范式应用：检测由不同微生物启动子培养物诱导的酸奶乳酸发酵期间释放的VOC（在线生物过程监测），监测与不同苹果栽培品种（大规模筛选）相关的挥发性有机化合物（VOC），以及饮用咖啡时鼻内VOC释放的体内研究（鼻孔空间分析）。
Protocol
Representative Results
Discussion
耦合飞行时间（ToF）质谱分析仪的质子转移反应 – 质谱（PTR-MS）代表了对挥发性有机化合物的鉴定和定量的需要以及快速分析分析的必要性之间的有效折中。表征ToF质量分析仪的高质量分辨率为相关灵敏度和质谱提供了相当大的信息内容。此外，PTR-ToF-MS与自动采样器的配合以及增加自动化程度的定制数据分析工具的应用，增强了这一技术的潜力。
在许多研究和技术领域，挥?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
This work is supported by the European Commission’s 7th Framework Programme under Grant Agreement Number 287382. SY is a beneficiary of a European Commission’s 7th Framework Programme Grant Agreement Number 287382. IK is a beneficiary of a FIRST doctoral school grant from the Fondazione Edmund Mach. For his work at University of Foggia, VC is supported by the Apulian Region in the framework of ‘Future In Research’ program (practice code 9OJ4W81).
Materials
| PTR-TOF 8000 High-Resolution PTR-TOF-MS | Ionicon Analytik Ges.m.b.H. | PTR-TOF 8000 | An detector for volatile organic compounds (VOCs) that allows for continuous VOC quantification with a very high mass resolution |
| GERSTEL MPS 2XL | Gerstel | A multifunctional autosampler | |
| Gas Calibration Unit | Ionicon Analytik Ges.m.b.H. | GCU-s / GCU-a | A dynamic gas dilution system that provides variable but known quantities of different standard compounds in a carrier gas stream |
| TofDaq | Tofwerk AG | free available at http://soft.tofwerk.com/ | A data acquisition software (for spectra acquisition) |
| MATLAB | MathWorks | http://it.mathworks.com/products/matlab/ | A technical computing language and interactive environment for algorithm development, data visualization, and data analysis |
| R | The R Foundation | free available at https://cran.r-project.org/mirrors.html | A language and environment for statistical computing and graphics |
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