Improved quantification of UV-B absorbing compounds in Pinus sylvestris L. pollen grains using an internal standard methodology
Improved quantification of UV-B absorbing compounds in Pinus sylvestris L. pollen grains using an internal standard methodology
复制标题
DOI：
10.1016/j.revpalbo.2017.08.007
复制
发表时间：
2017-12-01
影响因子：
1.9
通讯作者：
Willis, Kathy J.
中科院分区：
文献类型：
--
作者：
Seddon, Alistair W. R.;Jokerud, Mari;Willis, Kathy J.
UV-B 吸收化合物（例如对香豆酸）是花粉粒孢子花粉外壁的主要成分。最近的研究表明，在暴露于较高水平 UV-B 辐射的植物花粉中发现这些化合物的浓度较高，并且研究提出化石花粉中对香豆酸的变化可以作为到达地球表面的 UV-B 量变化的代表。然而，使用热辅助水解和甲基化热解气相色谱质谱法 (THM-GC/MS) 的既定方法的分析精度较低，这意味着孢粉质中 UV-B 吸收化合物的定量仍然是一个重大挑战。在这里，我们结合 THM-GC/MS 测试了多种归一化程序，以找到一种可以提高樟子松花粉 UV-B 吸收化合物定量分析精度的方法。将 UV-B 吸收化合物与孢子花粉中发现的非 UV-B 吸收化合物的标准化与基于外部和内部标准的方法进行比较。在测试的不同方法中，用作内标的香草酸（4-羟基-3-甲氧基苯甲酸）提供了改善性能的最佳潜力，当使用次佳标准化程序时，分析精度提高了约 43%。使用这种方法，我们估计从西班牙加泰罗尼亚的 P. sylvestris 个体收集的样品中对香豆酸的丰度为 0.34 +/- 0.02 nggrain(-1)（95% 置信区间，n = 20）。这项研究的结果为之前提出的用于定量花粉粒中对香豆酸的 THM-GC/MS 程序提供了优势，提高了分析精度并增强了稳健性。这将提高长期分析的批次样品的一致性，并能够在不同实验室运行的样品组之间进行比较。 (C) 2017 Elsevier B.V. 保留所有权利。
UV-B absorbing compounds such as para-coumaric acid are a major constituent of the sporopollenin-based exines of pollen grains. Recent research indicates that these compounds are found in higher concentrations in the pollen of plants exposed to higher levels of UV-B radiation and studies have proposed that variations of para-coumaric acid within fossil pollen could act as a proxy for changes in the amount of UV-B reaching the Earth's surface. However, the low analytical precision in the established method using Thermally Assisted Hydrolysis and Methylation with pyrolysis Gas-Chromatography Mass-Spectrometry (THM-GC/MS) means that quantification of UV-B absorbing compounds within sporopollenin remains a major challenge. Here, we test a variety of normalisation procedures combined with THM-GC/MS to find a method that can provide improved analytical precision in the quantification of UV-B absorbing compounds for Pinus sylvestris L. pollen. Normalisation of UV-B absorbing compounds against non-UV-B absorbing compounds found within sporopollenin was compared to external and internal standard-based approaches. Of the different methods tested, vanillic acid (4-hydroxy-3-methoxybenzoic acid) used as an internal standard provided the best potential for improved performance, with analytical precision improving by approximately 43% when the next-best normalisation procedures were used. Using this method, we estimate the abundance of para-coumaric acid to be 0.34 +/- 0.02 ng grain(-1) (95% confidence intervals, n = 20) from a sample collected from a P. sylvestris individual from Catalunya, Spain. The findings from this study provide advantages to previous THM-GC/MS procedures proposed for quantification of para-coumaric acid in pollen grains in terms of improved analytical precision and increased robustness. This will result in improved consistency for batches of samples analysed over long time periods and will enable comparisons between sample sets run in different laboratories. (C) 2017 Elsevier B.V. All rights reserved.