[16- FAPESP-BE] An integrated approach to explore a novel paradigm for biofuel production from lignocellulosic feedstocks

气候变化至少在一定程度上是由化石燃料的燃烧和随之而来的二氧化碳排放到环境中造成的。为了缓解这种情况，我们需要从可再生资源中生产更多的燃料/化学品。一种与全球相关的丰富资源是木质纤维素（存在于木材、秸秆、草和许多废物流中），目前正在努力有效地利用这种资源。然而，由于酵母（生物燃料发酵中最常用的生物）的限制，目前的工艺具有固有的低效率。酵母擅长将葡萄糖和蔗糖等单糖转化为乙醇，但天然菌株不能代谢木糖，木糖在木质纤维素或长链糖（低聚糖）中含量丰富。这意味着对于酵母发酵，有必要将木质纤维素分解为简单的单体糖，以便有效地利用它们。这种方法通常需要苛刻的物理化学预处理方法，这增加了过程的能量需求，并产生可以抑制后续发酵的化合物。因此，通常需要去除这些抑制剂，这增加了过程的费用。在这个项目中，我们打算证明不这么严厉地预处理木质纤维素是更明智的（逻辑和经济的），并且对该过程采用更“全面”的方法：通过优化产生部分分解产物并确保随后的发酵生物能够将这些直接转化为产品，从而在提供所需产品的同时最大限度地降低整个过程的能量和成本。最常用的一类发酵生物——酵母——将被设计成能够直接转化低聚糖。然而，有一类生物——地杆菌——已经被英国的一个研究小组广泛地研究过，它已经自然地有利用低聚糖的倾向，也可以很容易地设计来优化关键的代谢途径。因此，在这个项目中，我们将使用这组细菌的代表与工程酵母的性能进行比较。我们还建议在本研究中考虑三种不同的木质纤维素原料，它们都具有用于可持续燃料和化学品生产的潜力：巴西的甘蔗秸秆——收获后留在田野里；芒草——在英国种植，用于发电站燃烧（共烧），与甘蔗秸秆有很多相似之处；桉树林业残留物，在巴西储量丰富，代表了不同类型的机会和材料进行评估。一些团队成员将专注于开发方法，将这些转化为可以被这些新菌株吸收的低聚糖。这将是一个不那么严格（比目前）的预处理和使用选定的酶来生产所需的低聚糖的组合。团队的另一部分将专注于生产将这些转化为低聚糖所需的酶，而第三组将设计酵母菌株，使用木糖和葡萄糖的低聚糖。为了提高新的木质纤维素工厂原料的能源效率，我们将通过将厌氧消化（AD）整合到该过程中，从液体流出物、酒液和半纤维素水解物中回收化学品和沼气。混合培养的AD发酵可以提高能量比，产生沼气和肥料。所有这一切的基础是需要确保这项工作的产出与它们可能进入的工业过程保持相关。因此，我们有一个LCA专家团队，确保原料/产品选择是合适的，建议的过程优化方法对过程性能产生积极影响，并确定可以进行进一步更改/修改的地方。

Climate change is being driven, at least partly, by the burning of fossil fuels and consequent CO2 release into the environment. To mitigate this we need to produce more fuels/chemicals from renewable resources. One globally relevant abundant resource is lignocellulose (present in wood, straw, grasses and in many waste streams) and efforts are being made to exploit this efficiently. However, current processes have inherent inefficiencies due to the limitations of yeast, the most common organism used in biofuel fermentations. Yeasts are good at converting simple sugars such as glucose and sucrose to ethanol, but natural strains cannot metabolise xylose, which is abundant in lignocellulose, or longer chains of sugars (oligosaccharides). This means that for yeast fermentations it is necessary to break down the lignocellulose to simple monomeric sugars for them to be utilised effectively. This approach generally requires harsh physico-chemical pre-treatment methods which, increase the energy demand of the process and produce compounds that can inhibit the subsequent fermentation. Thus it is often necessary to remove these inhibitors, which adds expense to the process. In this project we intend to demonstrate that it is more sensible (logical and economic) not to pre-treat lignocellulose so harshly, and have a more "holistic" approach to the process: delivering the desired products whilst minimising overall process energy and cost by working on the optimisation of generating partial breakdown products and ensuring that the subsequent fermentation organism is able to convert these directly to product.The most commonly employed class of fermentation organisms - yeasts - will be engineered to be able to convert the oligomeric sugars directly. However, there is a class of organisms - Geobacillus - that have been quite extensively studied by one of the UK groups, which already naturally has the propensity to utilise oligomeric sugars and can also be readily engineered to optimise key metabolic pathways. Therefore, in this project we will use a representative of this group of bacteria to compare performance with the engineered yeast.We also propose to consider three different lignocellulosic feedstocks in this study, all of which have the potential to be used for sustainable fuels and chemicals production: Brazilian cane straw - which is current left in the fields after harvesting, Miscanthus - which is grown in the UK for burning in power stations (co-firing) and has a lot of similarities to cane straw, and Eucalyptus forestry residues, which are abundant in Brazil and represent a different type of opportunity and material to evaluate. Some of the team involved will focus on developing methods to convert these to oligosaccharides that can be taken up by these new strains. This will be a combination of less severe (than currently) pre-treatment and the use of selected enzymes to produce the oligo-saccharides required. Another part of the team will focus on producing the enzymes required for these conversions to oligosaccharides, while a third group will engineer the yeast strains to use oligosaccharides of both xylose and glucose.To increase the energy efficiency of the feedstocks in the new lignocelulose mills we are going to recover chemicals and biogas from the liquid effluents, vinasse and hemicellulose hydrolysates, by integrating anaerobic digestion (AD) to the process. AD with mixed culture fermentation will improve the energy ratio bringing biogas production and fertilizers as products.Underpinning all this is the need to ensure that the outputs of this work remains relevant to the industry processes that they potentially feed into. Therefore we have a team of LCA experts ensuring that feedstock/ product choice is appropriate, that the proposed process optimisation approaches are delivering a positive impact on process performance and pinpointing where further changes/modifications could be made.