Biorefinery Laboratory, Latvian State Institute of Wood Chemistry, LV-1006 Riga, Latvia.
PolyLabs SIA, Mukusalas iela 46, LV-1004 Riga, Latvia.
Molecules. 2023 Feb 27;28(5):2227. doi: 10.3390/molecules28052227.
Suberin is a complex polyester biopolymer, and it is practically impossible to estimate the real content of suberin in suberised plant tissues. This indicates the importance of the development of instrumental analytical methods for the comprehensive characterisation of suberin derived from plant biomass for the successful integration of suberinic products into biorefinery production chains. In this study, we optimised two GC-MS methods-one with direct sylilation, and the second with additional depolymerisation, using GPC methods with RI detector and polystyrene calibration and with a three-angle light scattering detector and an eighteen-angle light scattering detector. We also performed MALDI-Tof analysis for non-degraded suberin structure determination. We characterised suberinic acid (SA) samples obtained from birch outer bark after alkaline depolymerisation. The samples were particularly rich in diols, fatty acids and their esters, hydroxyacids and their corresponding esters, diacids and their corresponding esters, as well as extracts (mainly betulin and lupeol) and carbohydrates. To remove phenolic-type admixtures, treatment with ferric chloride (FeCl) was used. The SA treatment with FeCl allows the possibility to obtain a sample that has a lower content of phenolic-type compounds and a lower molecular weight than an untreated sample. It was possible to identify the main free monomeric units of SA samples by GC-MS system using direct silylation. By performing an additional depolymerisation step before silylation, it was possible to characterise the complete potential monomeric unit composition in the suberin sample. For the molar mass distribution determination, it is important to perform GPC analysis. Even though chromatographic results can be obtained using a three- laser MALS detector, they are not fully correct because of the fluorescence of the SA samples. Therefore an 18-angle MALS detector with filters was more suitable for SA analysis. MALDI-Tof analysis is a great tool for the polymeric compound structural identification, which cannot be done using GC-MS. Using the MALDI data, we discovered that the main monomeric units that makes up the SA macromolecular structure are octadecanedioic acid and 2-(1,3-dihydroxyprop-2-oxy)decanedioic acid. This corresponds with GC-MS results, showing that after depolymerisation hydroxyacids and diacids were the dominant type of compounds found in the sample.
化感物质是一种复杂的聚酯生物聚合物，实际上几乎不可能估计被化感物质包裹的植物组织中化感物质的真实含量。这表明开发仪器分析方法对于全面表征植物生物量衍生的化感物质的重要性，以便成功将化感物质产品整合到生物炼制生产链中。在这项研究中，我们优化了两种 GC-MS 方法——一种是直接硅烷化，另一种是在额外解聚后进行硅烷化，使用带有 RI 检测器和聚苯乙烯校准的 GPC 方法，以及带有三个角度光散射检测器和十八个角度光散射检测器的 GPC 方法。我们还进行了 MALDI-ToF 分析，以确定非降解化感物质的结构。我们对桦木外树皮经过碱性解聚后得到的化感酸（SA）样品进行了表征。这些样品特别富含二醇、脂肪酸及其酯、羟基酸及其相应的酯、二酸及其相应的酯，以及提取物（主要是白桦脂醇和羽扇豆醇）和碳水化合物。为了去除酚型杂质，使用三氯化铁（FeCl）进行处理。用 FeCl 处理 SA 可以获得一种比未处理样品酚型化合物含量更低、分子量更小的样品。通过使用 GC-MS 系统进行直接硅烷化，我们可以识别 SA 样品的主要游离单体单元。通过在硅烷化前进行额外的解聚步骤，可以对化感物质样品中的完整潜在单体单元组成进行表征。对于摩尔质量分布的测定，进行 GPC 分析非常重要。尽管可以使用三激光 MALS 检测器获得色谱结果，但由于 SA 样品的荧光，结果并不完全正确。因此，带有滤波器的 18 角 MALS 检测器更适合用于 SA 分析。MALDI-ToF 分析是一种用于聚合物化合物结构鉴定的强大工具，而 GC-MS 无法完成这种鉴定。使用 MALDI 数据，我们发现构成 SA 大分子结构的主要单体单元是十八烷二酸和 2-(1,3-二羟基丙-2-氧基)癸二酸。这与 GC-MS 结果一致，表明解聚后，羟基酸和二酸是样品中主要的化合物类型。
