Elliston Adam, Wood Ian P, Soucouri Marie J, Tantale Rachelle J, Dicks Jo, Roberts Ian N, Waldron Keith W
The Biorefinery Centre, Institute of Food Research, Norwich Research Park, Colney, Norwich, NR4 7UA UK.
The Biorefinery Centre, Institute of Food Research, Norwich Research Park, Colney, Norwich, NR4 7UA UK ; École supérieure d'ingénieurs Réunion Océan Indien, Génie Biologique, Université de La Réunion, Parc Technologique Universitaire, 2 Rue Joseph Wetzell, 97490 Sainte-Clotilde, La Réunion France.
Biotechnol Biofuels. 2015 Jan 22;8(1):2. doi: 10.1186/s13068-014-0181-z. eCollection 2015.
High-throughput (HTP) screening is becoming an increasingly useful tool for collating biological data which would otherwise require the employment of excessive resources. Second generation biofuel production is one such process. HTP screening allows the investigation of large sample sets to be undertaken with increased speed and cost effectiveness. This paper outlines a methodology that will enable solid lignocellulosic substrates to be hydrolyzed and fermented at a 96-well plate scale, facilitating HTP screening of ethanol production, whilst maintaining repeatability similar to that achieved at a larger scale.
The results showed that utilizing sheets of biomass of consistent density (handbills), for paper, and slurries of pretreated biomass that could be pipetted allowed standardized and accurate transfers to 96-well plates to be achieved (±3.1 and 1.7%, respectively). Processing these substrates by simultaneous saccharification and fermentation (SSF) at various volumes showed no significant difference on final ethanol yields, either at standard shake flask (200 mL), universal bottle (10 mL) or 96-well plate (1 mL) scales. Substrate concentrations of up to 10% (w/v) were trialed successfully for SSFs at 1 mL volume. The methodology was successfully tested by showing the effects of steam explosion pretreatment on both oilseed rape and wheat straws.
This methodology could be used to replace large shake flask reactions with comparatively fast 96-well plate SSF assays allowing for HTP experimentation. Additionally this method is compatible with a number of standardized assay techniques such as simple colorimetric, High-performance liquid chromatography (HPLC) and Nuclear magnetic resonance (NMR) spectroscopy. Furthermore this research has practical uses in the biorefining of biomass substrates for second generation biofuels and novel biobased chemicals by allowing HTP SSF screening, which should allow selected samples to be scaled up or studied in more detail.
高通量（HTP）筛选正成为一种越来越有用的工具，用于整理生物数据，否则这些数据的收集将需要耗费过多资源。第二代生物燃料生产就是这样一个过程。HTP筛选能够以更高的速度和成本效益对大量样本进行研究。本文概述了一种方法，该方法可使固体木质纤维素底物在96孔板规模下进行水解和发酵，便于对乙醇生产进行HTP筛选，同时保持与更大规模实验相似的可重复性。
结果表明，使用密度一致的生物质片材（传单）作为纸张，以及可通过移液器移取的预处理生物质浆料，可实现向96孔板的标准化和准确转移（分别为±3.1%和1.7%）。在不同体积下通过同步糖化发酵（SSF）处理这些底物，在标准摇瓶（200 mL）、通用瓶（10 mL）或96孔板（1 mL）规模下，最终乙醇产量均无显著差异。在1 mL体积下成功测试了高达10%（w/v）的底物浓度用于SSF。通过展示蒸汽爆破预处理对油菜籽和小麦秸秆的影响，该方法得到了成功验证。
该方法可用于用相对快速的96孔板SSF分析取代大型摇瓶反应，从而实现HTP实验。此外，该方法与多种标准化分析技术兼容，如简单比色法、高效液相色谱（HPLC）和核磁共振（NMR）光谱法。此外，本研究在第二代生物燃料和新型生物基化学品的生物质底物生物精炼方面具有实际应用价值，通过允许HTP SSF筛选，这应能使选定的样品扩大规模或进行更详细的研究。
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