, 欧阳嘉 南京林业大学 化学工程学院，江苏 南京 210037
收稿日期：2020-06-10；接收日期：2020-08-06；网络出版时间：2020-09-01
基金项目：国家重点研发计划(No. 2018YFA0902200) 资助
摘要：木质纤维素是地球上储量最为丰富的可再生有机碳资源，但由于其结构的复杂性，必须经过一系列预处理过程才能被微生物高效利用，这就不可避免地带来了呋喃醛等典型抑制物，严重阻碍了微生物的生长和后续发酵过程。认知微生物的呋喃醛代谢途径，并基于此开发耐受性和转化能力强的微生物菌株是生物炼制领域的重要研究内容。文中综述了呋喃醛抑制物的来源、呋喃醛对微生物的抑制机理以及微生物降解呋喃醛的代谢途径，并重点讨论了基于生物法降解呋喃醛抑制物的研究进展，涉及的主要技术手段包括传统的适应性进化工程和代谢工程，以及近年来新兴的微生物共培养系统和功能化材料辅助微生物脱除呋喃醛等。
关键词：木质纤维素呋喃醛降解途径生物法
Biodegradation of furan aldehydes in lignocellulose hydrolysates
Huimin Sun, Lihua Zou, Zhaojuan Zheng
, Jia Ouyang College of Chemical Engineering, Nanjing Forestry University, Nanjing 210037, Jiangsu, China
Received: June 10, 2020; Accepted: August 6, 2020; Published: September 1, 2020
Supported by: National Key Research and Development Program of China (No. 2018YFA0902200)
Corresponding author: Zhaojuan Zheng. Tel: +86-25-85427587; E-mail: zhengzj@njfu.edu.cn.
Abstract: Lignocellulose is the most abundant renewable organic carbon resource on earth. However, due to its complex structure, it must undergo a series of pretreatment processes before it can be efficiently utilized by microorganisms. The pretreatment process inevitably generates typical inhibitors such as furan aldehydes that seriously hinder the growth of microorganisms and the subsequent fermentation process. It is an important research field for bio-refining to recognize and clarify the furan aldehydes metabolic pathway of microorganisms and further develop microbial strains with strong tolerance and transformation ability towards these inhibitors. This article reviews the sources of furan aldehyde inhibitors, the inhibition mechanism of furan aldehydes on microorganisms, the furan aldehydes degradation pathways in microorganisms, and particularly focuses on the research progress of using biotechnological strategies to degrade furan aldehyde inhibitors. The main technical methods include traditional adaptive evolution engineering and metabolic engineering, and the emerging microbial co-cultivation systems as well as functional materials assisted microorganisms to remove furan aldehydes.
Keywords: lignocellulosefuran aldehydesdegradation pathwaybiotechnological strategy
木质纤维素是地球上储量最为丰富的可再生资源，也是唯一的可再生有机碳资源。全球每年的木质纤维素产量约为10 000亿t，但其中被利用的生物质不足10%[1]，木质纤维素的高效利用对缓解全球能源危机、减少温室效应和环境污染、实现人类社会的绿色可持续发展具有重要意义。我国作为农业大国，每年农业废弃物大约有8–10亿t[2]，以这些廉价的非粮木质纤维资源制备生物燃料和大宗化学品，来替代部分不可再生的化石资源，将会带来显著的环境效益和社会效益。
木质纤维素是植物细胞壁的主要组成部分，一般由纤维素、半纤维素和木质素组成，它们通过共价键或非共价键相互结合。由于木质纤维素结构的复杂性，必须经过一系列预处理过程才能被高效糖化利用[3]。预处理过程能脱除其中的木质素，水解半纤维素，同时降低纤维素结晶度使原料结构疏松，从而提高纤维素酶的酶解效率[4]。此外，预处理会导致木质纤维原料发生一系列复杂的理化反应，不可避免地生成呋喃醛、弱酸类、酚类等结构多样的抑制物，严重阻碍了微生物的生长和后续发酵过程[5]。
为了去除木质纤维素水解产物中抑制物，研究者开发了多种物理、化学和生物方法[6-9]，其中生物法是以微生物或酶作为催化剂，从水解产物中针对性去除抑制物。相较于物理法和化学法，生物法不需要特殊设备、不会引入新的对后续发酵有影响的化合物、对环境友好[10]。近年来，科研人员围绕抑制物对微生物的抑制机理、微生物代谢抑制物的生化途径以及增强微生物抑制物代谢能力等方面开展了一系列研究工作。本文针对呋喃醛抑制物，综述了其来源、抑制机理和在微生物中的降解途径，并讨论了利用生物法降解呋喃醛抑制物的研究进展，包括传统的利用微生物和酶降解转化呋喃醛抑制物、适应性进化工程和代谢工程增强微生物降解呋喃醛抑制物的能力，以及近年来新兴的微生物共培养系统和功能化材料辅助微生物脱除呋喃醛抑制物等。
1 呋喃醛抑制物的来源及抑制机理图 1展示了木质纤维素经预处理后产生抑制物的来源，这些抑制物显著影响细胞活力和生长速度，延长滞后期，导致发酵过程的低产量和低生产力[11-14]。呋喃醛是最重要的微生物抑制剂之一，主要包括糠醛和5-羟甲基糠醛(5-hydroxymethylfurfural，HMF)，它们分别是戊糖和己糖的脱水产物[15]。不同原料及不同预处理方式产生的糠醛和HMF浓度不同，糠醛浓度大约在0.05–4.0 g/L[16-17]；HMF浓度大约在0.05–3.3 g/L[18-19]。
 |
| 图 1 木质纤维素水解液中的抑制物来源[5] Fig. 1 Source of inhibitors in lignocellulose hydrolysates[5]. |
| 图选项 |
糠醛和HMF对微生物的抑制机理相似，但糠醛对细胞生长和代谢的抑制作用较HMF更强，这可能与其疏水性相关[20]。呋喃醛通常会抑制糖酵解途径和三羧酸循环(TCA循环) 等重要碳源代谢途径中的相关酶，如己糖激酶、3-磷酸甘油醛脱氢酶、丙酮酸脱氢酶、乙醛脱氢酶和乙醇脱氢酶等[21]。呋喃醛还会导致细胞能量通量的重定向，降低细胞生成ATP和NAD(P)H的能力[3]。Hadi等研究发现呋喃醛的高疏水性会破坏细胞膜的完整性，渗入细胞造成DNA链断裂成单链，抑制RNA和蛋白质合成[22]。此外，进入胞内的呋喃醛还会引起细胞内活性氧(Reactive oxygen species，ROS) 的积累，ROS含量的增加进而引起线粒体、液泡膜、肌动蛋白细胞骨架和细胞核染色体的损伤[23]。上述抑制作用最终会导致低生物量和低发酵产率，严重时还会造成细胞完全死亡。
2 呋喃醛抑制物的降解途径某些细菌和真菌可以彻底氧化降解糠醛和HMF。早在1969年，Trudgill就报道了恶臭假单胞菌Pseudomonas putida F2可以完全代谢糠醛为α-酮戊二酸，并基于菌株胞内酶活力提出了有氧条件下糠醛的降解路径[24]。之后，与该菌株同种属的P. putida Fu1也被报道可以完全代谢糠醛。但是，能完全降解HMF的菌株及相应的代谢途径一直未见报道。直到2010年，Wierckx等分离鉴定了一株新的革兰氏阴性菌巴塞尔贪铜菌Cupriavidus basilensis HMF14，其兼具糠醛和HMF的降解能力[25]。基于转座子突变筛选，Koopman等发现C. basilensis HMF14基因组上存在两个与呋喃醛代谢相关的操纵子，其具体催化路径如图 2所示[26]。糠醛首先在非特异性脱氢酶催化下还原为糠醇，之后经HMF/糠醛氧化还原酶的作用氧化生成糠酸，糠酸在糠酰-CoA合成酶催化下与CoA连接生成糠酰-CoA，后者经糠酰-CoA脱氢酶羟基化得到5-羟基-2-糠酰-CoA。5-羟基-2-糠酰-CoA依次通过自发的异构化、内酯水解开环、酮烯醇互变异构化、水解反应转化为α-酮戊二酸。至此，糠醛代谢产物最终进入TCA循环实现彻底氧化分解。HMF的降解路径与糠醛类似，HMF/糠醛氧化还原酶首先氧化HMF为相应的单羧酸5-羟甲基糠酸(5-hydroxymethyl-2- furancarboxylic acid，HMFCA)，随后进一步氧化单羧酸生成2, 5-呋喃二羧酸(2, 5-furandicarboxylic acid，FDCA)。FDCA经2, 5-呋喃二羧酸脱羧酶反应生成糠酸，之后的降解路径与糠醛降解路径重合，最终都以α-酮戊二酸的形式进入TCA循环实现糠醛和HMF的终极降解[26]。
 |
| 图 2 C. basilensis HMF14菌株降解呋喃醛的途径[26] (图中的五边形和三角形分别代表不同的酶：五边形，糠醛/ HMF氧化还原酶；三角形1，2, 5-呋喃二羧酸脱羧酶；三角形2，糠酰-CoA合成酶；三角形3，糠酰-CoA脱氢酶；三角形4，α-酮戊二酰-CoA水解酶. 标记有圆形的反应过程可以由HmfH/非特异性脱氢酶催化。标记有正方形的反应过程表示可能自发发生的内酯水解，或可能由内酯水解酶催化的内酯水解. 双箭头表示酮-烯醇互变异构) Fig. 2 The furan aldehydes metabolic pathway in C. basilensis HMF14[26]. The pentagons and triangles represent different enzymes: pentagon, furfural/HMF oxidoreductase; triangle 1, 2, 5-furan-dicarboxylic acid decarboxylase; triangle 2, 2-furoyl-CoA synthetase; triangle 3, furoyl-CoA dehydrogenase; triangle 4, 2-oxoglutaryl-CoA hydrolase. The reaction process marked with a circle can be catalyzed by HmfH or non-specific dehydrogenase. The reaction process marked with a square indicates a lactone hydrolysis that may occur spontaneously, or may be catalyzed by a generic lactone hydrolase. Double-pointed arrows indicate keto-enol tautomerizations. |
| 图选项 |
除了上述细菌外，研究者还分离到了多株能够完全代谢呋喃醛的真菌。Nichols等从被糠醛污染过的土壤中分离到了一株可以代谢呋喃醛、有机酸以及酚类化合物的真菌Coniochaeta ligniaria NRRL30616，其可以把糠醛以及HMF转化为毒性弱的相应的醇，即糠醇和2, 5-呋喃二甲醇(2, 5-bis(hydroxymethyl)furan，BHMF)，然后再将醇氧化为相应的酸。最终，FDCA也转化为糠酸，之后呋喃环裂解成可以进入TCA循环的中间物质[27]。我国华东理工大学鲍杰团队筛选的丝状真菌树脂枝孢霉Amorphotheca resinae ZN1也能够完全代谢糠醛和HMF。王晓凤对A. resinae ZN1的呋喃醛代谢产物进行实时监测分析，并推测其代谢途径为：糠醛首先还原为低毒性的糠醇，在有氧条件下，糠醇又被氧化为低浓度的糠醛，糠醛被进一步氧化为糠酸；HMF的降解路径与糠醛类似，也是首先还原为对微生物毒性小的BHMF，在有氧条件下，BHMF再度生成不会对微生物产生危害的低浓度HMF，后者继续氧化为HMFCA[28-29]。之后该团队进一步完善了糠醛和HMF的代谢途径，发现该真菌中的呋喃醛代谢途径与前人提出的细菌代谢途径一致[30-31]。据此可以推测，无论细菌还是真菌，微生物代谢呋喃醛的路径均与C. basilensis HMF14非常类似。
上述提及的微生物在有氧条件下可以将糠醛或HMF氧化至α-酮戊二酸进入TCA循环，因此，这些微生物都能以糠醛或HMF作为唯一碳源生长。此外，许多微生物虽不能利用糠醛或HMF作为唯一碳源生长，但能够将呋喃醛转化为毒性较小的呋喃醇(如糠醇和BHMF) 或呋喃酸(如糠酸和HMFCA) 为终产物，从而降低呋喃醛的抑制影响。例如，在呋喃醛还原方面，大肠杆菌Escherichia coli可利用NADPH依赖的醛还原酶YqhD将糠醛还原为毒性较小的糠醇[32-33]；酿酒酵母Saccharomyces cerevisiae H. 则利用NADH依赖的醇脱氢酶Adh1将糠醛还原为糠醇[34]；谷氨酸棒杆菌Corynebacterium glutamicum中NADPH依赖的醇脱氢酶基因fudC是将糠醛还原为糠醇的主要基因，值得注意的是，fudC与yqhD没有同源性，却与E. coli中的另一种醇脱氢酶基因yahK有高达61%的相似性[35]；季也蒙毕赤酵母Meyerozyma guilliermondii SC1103也能将呋喃醛还原为呋喃醇，且底物耐受性强，但其中行使催化功能的酶尚未报道[36]；笔者所在课题组前期也分析了不同种属菌株在呋喃醛还原方面的转化潜力，发现S. cerevisiae NL22[37]和凝结芽孢杆菌Bacillus coagulans NL01[38]的转化能力非常强。在呋喃醛氧化方面，睾丸酮丛毛单胞菌Comamonas testosteroni SC1588[39]、氧化葡萄糖酸杆菌Gluconobacter oxydans DSM 50049[40]、乌鲁木齐奇异球菌Deinococcus wulumuqiensis R12[41]以及笔者所在课题组报道的P. putida KT2440[42]等菌株都能够快速氧化糠醛和HMF分别至糠酸和HMFCA，但令人遗憾的是，这些菌株中行使呋喃醛氧化功能的酶尚未得到鉴定。Zhang等将C. testosteroni SC1588基因组上的6个不同的醛脱氢酶依次在大肠杆菌中异源表达，发现其中的4个具有不同程度的呋喃醛氧化活力[43]。此外，很多具有宽底物谱的醛氧化还原酶，也被报道可在体外转化呋喃醛，例如马肝醇脱氢酶[44-45]、Baeyer-Villiger单加氧酶[46]等都被报道用于糠醛和HMF的氧化。
3 生物法脱除预处理液中的呋喃醛生物法脱除预处理液中的呋喃醛是利用微生物或酶完全降解或部分转化呋喃醛从而解除或降低呋喃醛对后续发酵的抑制。与其他抑制物脱除方法相比较，生物法反应条件温和、抑制物去除彻底、可发酵糖损失和废水排放均较少[47-48]。除了传统的微生物法和酶法，研究者还利用适应性进化工程、代谢工程，以及新兴的微生物共培养系统和材料辅助等技术手段进一步提高生物法的脱除效率(表 1)。
表 1 生物法脱除预处理液中的呋喃醛Table 1 Summary of biological strategies to remove furan aldehydes from pretreatment hydrolysates
| Strategy | Approach | Biocatalyst | Effect | References |
| Microorganism | Screening microorganisms | I. orientalis and I. occidentalis | Low consumption of sugar, removing 100% furfural, 100% and 84.0% HMF | [50] |
| Whole cell catalysis | P. putida KT2440 | Tolerance of 190 mmol/L HMF, convert HMF to HMFCA as valuable product | [52] | |
| Enzyme | Fixed on cellulose nanofiber | Laccase | Degradation of 12 mmol/L furfural within 36 h | [57] |
| Adaptive evolution | Ultraviolet mutagenesis combined with adaptive evolution | S. cerevisiae | Improve resistance to furfural, shorten the lag period, improve the growth rate, the ethanol yield and the conversion rate of furfural | [61] |
| Adaptive evolution | Z. mobilis | Tolerance of 3.0 g/L furfural | [62] | |
| Metabolic engineered microorganism | Expression of adh1 | E. coli | Enhancement in furfural tolerance and furfural degradation ability | [21] |
| Expression of zmo1771 and udhA | Z. mobilis ZM4 | The conversion rate of furfural and HMF increased by 51.9% and 92.0% | [31] | |
| Microbial co-cultivation system | Co-cultivation | Desulfovibrio B and M. barkeri 227 | Methane production with furfural as substrate | [76] |
| Co-cultivation | Two S. cerevisiae strians | Xylose as substrate, increased tolerance against inhibitors, ethanol yield increased by 41.0% | [77] | |
| Functional material assisting | Entrapped in sodium alginate | M. guilliermondii SC1103 | Conversion rate of HMF can reach 82.0%–85.0% within 7–24 h, the cell activity can still reach 70.0% after reused four times | [81] |
| Entrapped in chitosan | Bordetella sp. BTIITR | 86% HMF and 100% furfural degradation in bagasse hydrolysate, the fixed cells still have good activity after reused seven times | [78] |
表选项
3.1 传统微生物法和酶法目前自然界中发现的可以降解或转化呋喃醛的微生物主要分为两类。一类是能彻底降解呋喃醛为中心代谢中间产物α-酮戊二酸，从而被菌体作为碳源利用；这类菌株甚至更偏好以呋喃醛为碳源，在呋喃醛完全代谢后才开始利用可发酵糖。Okuda等研究了Ureibacillus thermosphaercus对废弃木材水解产物的生物解毒作用，经色谱分析证实，U. thermosphaercus可降解水解物中存在的糠醛、HMF以及酚类化合物。U. thermosphaercus生长迅速，在抑制物消除过程中，消耗的可发酵糖不到5.0%[49]。Guedes等研究发现，用东方伊萨酵母Issatchenkia orientalis和西方伊萨酵母Issatchenkia occidentalis降解甘蔗水解液中的抑制物时，24 h后两种酵母菌在几乎不利用木糖和阿拉伯糖的情况下都可以将糠醛完全降解，HMF的降解率分别为100%和84.0%[50]。Singh等用富集培养的方法从土壤中分离出一株具有降解木质纤维素水解液中糠醛和HMF能力的博德特氏菌Bordetella sp. BTIITR，其在甘蔗渣水解液中培养16 h后，能去除100%的糠醛、94.0%的HMF和82.0%的乙酸。此外，Bordetella sp. BTIITR在刚开始降解糠醛和HMF时，对可发酵糖的消耗很慢，一旦糠醛和HMF被降解到较低的浓度，可发酵糖的消耗就会明显加速[10]。总体而言，已报道的优先以呋喃醛为碳源的抑制物降解菌株还非常稀缺，仍需要研究者们不断地筛选、鉴定及开发应用。
另一类微生物只能将呋喃醛转化为抑制能力相对较弱的还原或氧化衍生物，这一类微生物除了用于抑制物脱除外，还可以用于一系列高附加值的呋喃醇或呋喃羧酸化合物的生产。Tsuge等研究了C. glutamicum ATCC 13032在有氧和厌氧条件下对糠醛的转化能力，发现在有氧条件下，糠醛的转化产物为糠醇和糠酸，且产物的比例随糠醛初始浓度的不同而不同；而在厌氧条件下，大部分糠醛被转化为糠醇[51]。Xu等报道了P. putida KT2440选择性生物氧化HMF制备HMFCA的研究，在最佳催化条件下，该菌对HMF的耐受性最高可达190 mmol/L，且在HMF为70–150 mmol/L之间时几乎将HMF全部氧化，获得了非常高的HMFCA收率；此外，它还可以选择性地氧化其他呋喃醛制备相应的呋喃羧酸，产率可达100%[52]。王娟将皮状丝孢酵母Trichosporon cutaneum ACCC 20271培养在玉米秸秆水解液中，发现其可耐受5.0 g/L的HMF，但0.5 g/L的糠醛即可显著抑制其生长，该菌株仅能将糠醛和HMF降解为醇和酸，不能实现终极降解[53]。此外还有树干毕赤酵母Pichia stipitis[54]、运动发酵单胞菌Zymomonas mobilis等[55]很多微生物虽然不能完全代谢呋喃醛，但都可以转化为低毒性的呋喃醇和呋喃酸，在一定程度上解除了呋喃醛的抑制效应。
采用酶法脱除抑制物时，常用的酶主要是漆酶和过氧化物酶。现有报道中，漆酶和过氧化物酶多用于酚类化合物的去除，它们对呋喃醛的去除作用不显著[56]。不过，Saravanakumar等将漆酶固定在木葡糖酸醋杆菌Gluconacetobacter xylinus生产的纤维素纳米纤维上，他们发现固定化漆酶具有脱除糠醛和松柏醛的能力[57]。利用酶法进行抑制物脱除最大的问题在于酶制剂生产成本高，因此酶制剂的回收利用尤为重要。
3.2 微生物适应性进化进化工程主要是指通过对菌株进行诱变和驯化以提高菌株耐受目标抑制物的能力，获得抑制物耐受菌株，其优势在于不会引入给菌株代谢带来负担的异源蛋白[58]。诱变就是用物理或化学方法对菌株进行诱变处理，目前多采用复合诱变，目的是使多种优良性状集于一株菌株上[59]。驯化就是将菌株长期置于一定的环境中，并逐渐增强环境的恶劣程度以获得对目标抑制物高耐受性的菌株[60]。
丁明珠以酿酒酵母为出发菌种，通过紫外诱变结合驯化的方法筛选出一株在含1.3 g/L糠醛、0.5 g/L苯酚和5.3 g/L乙酸的高糖培养基中仍有很强生长能力和较高代谢速率的耐受性菌株[61]。Shui等通过从1.0 g/L逐级提高糠醛浓度至3.0 g/L，对Z. mobilis进行适应性驯化，最终获得一株对3.0 g/L的糠醛仍具有很好耐受性的菌株[62]。进化工程菌株一般周期较长，且耐受机理复杂不清，后续需要结合基因组、转录组、蛋白质组或代谢组等组学数据分析，才能系统阐明进化菌株的耐受机制。
3.3 代谢工程改造微生物基于不同微生物对抑制物的耐受能力差异及其转录、蛋白表达和代谢差异分析，从基因组上寻找能增强微生物抑制物脱除能力的靶基因，并从耐受菌株的遗传文库中扩增该基因以改造目标宿主，从而实现增强目标菌株抗逆性的目的[4]。
细菌的呋喃醛抑制物抗逆研究表明，呋喃醛的转化主要涉及的氧化还原酶有FucO[63-64]、YqhD[32-33, 64]、DkgA[32, 64-65]、UcpA[66]等，这些蛋白的过量表达能够提高微生物的呋喃醛转化能力。真菌的呋喃醛抗逆工程研究中，研究者们对酿酒酵母关注较多，主要涉及醇脱氢酶[67]、醛还原酶[68-69]、葡萄糖-6-磷酸脱氢酶[70]、木糖还原酶[54]和NADPH依赖型的还原酶[71]等，处于呋喃醛环境中的酵母通过过表达上述相关酶蛋白以提高自身的呋喃醛耐受能力。Wang等发现在大肠杆菌中异源表达热带念珠菌Candida tropicalis的adh1可增强其糠醛耐受性和糠醛降解能力[21]。Park等发现S. cerevisiae的醛脱氢酶6既能促进呋喃醛的直接氧化，又能为呋喃醛的降解反应提供NADPH辅因子，通过过表达醛脱氢酶6可降低糠醛对酿酒酵母的抑制作用，并促进酿酒酵母的细胞生长和乙醇合成[72]。王霞在Z. mobilis ZM4中异源表达了来自A. resinae ZN1的内源醇脱氢酶基因zmo1771，增强菌株转化糠醛和HMF的能力，后又在上述重组菌株中进一步过表达了来自E. coli的转氢酶基因udhA，所得重组菌株在玉米秸秆水解液中发酵12 h后，其糠醛、HMF的转化率分别较单一过表达zmo1771的菌株提高了51.9%和92.0%[31]。Xiao等基于全基因组RNA干扰技术发现E3 SUMO-蛋白连接酶与酿酒酵母对糠醛的耐受性紧密相关，敲除编码该蛋白的siz1基因能够显著增强细胞对糠醛的耐受能力[73]。
微生物菌株对抑制物的耐受或去除往往不是一个或两个基因决定的，而是一个复杂的调控系统。代谢工程菌株在过表达目标蛋白时应避免异源蛋白质表达的代谢负担、外排泵的毒性和诱导剂的成本等问题[74-75]。
3.4 微生物共培养系统发酵前脱除抑制物会增加处理时间和过程成本，同步进行抑制物脱除和产品生产无疑可以进一步提高这一过程的经济性，并避免额外的处理步骤。为了实现这一目标，近年来，研究者尝试同时培养两种或两种以上的微生物，它们分工合作，在抑制物脱除的同时，实现目标产品的生产。例如，Boopathy等将硫酸盐还原菌脱硫弧菌Desulfovibrio B和产甲烷古菌巴氏甲烷八叠球菌Methanosarcina barkeri 227联合培养，实现了以糠醛为底物生产甲烷。Desulfovibrio B可以以糠醛为碳源进行生长，其代谢产物为乙酸，而M. barkeri 227则能以乙酸为底物，将之进一步转化为甲烷气体[76]。Zhu等将一株代谢工程改造过的含有木糖利用途径的工程酿酒酵母和一株抑制物耐受性强的酿酒酵母共培养于未脱毒的玉米秸秆水解液中，不仅同时解决了木糖利用和抑制物问题，而且乙醇产量相较于单一菌株，最高提升了41.0%[77]。利用菌株代谢途径和代谢能力差异，使其互惠共生或偏利共生、各司其职，不仅能实现抑制物的原位降解，也能实现目标产物的同步合成，这种微生物共培养方式正成为抑制物脱除领域的新研究方向。
3.5 功能化材料辅助法将微生物固定在惰性载体上或包封在多孔材料中形成珠状结构以增加微生物代谢性能已引起研究者的广泛关注。与自由细胞相比，固定化细胞具备许多优点，如重复批量操作中易于回收、防止细胞冲洗流失、提高生物反应器的灵活性和操作稳定性等。珠状结构还可增加对被包裹的微生物的保护，使其免受诸如温度变化、pH变化和有毒化学品等外界不利条件的影响[78]。
Nguyen等设计了用于固定化细胞的中空纤维膜生物反应器(ICHFMB)，收集指数生长末期的Z. mobilis细胞，并将其重新悬浮在0.3%的NaCl溶液中，然后使用蠕动泵以10 mL/min的流速在膜组件中循环6 h以固定细胞。固定化后的Z. mobilis细胞在香草醛、丁香醛、HMF和4-羟基-3-甲氧基肉桂醛存在时的葡萄糖摄取率和生长状况都较游离细胞有所改善，其中HMF的去除率可达89.0%，其乙醇产量可达到理论值的95.0%。通过增加水解液在膜上的流速和增加膜的数量，还可以进一步提高ICHFMB的性能[79]。
Kumar尝试将厌氧污泥与海藻酸钙、壳聚糖以及活性炭混合制作了杂交固定化细胞(H-IC)，将其用于去除稀酸预处理后的红藻水解液中的HMF并转化可发酵糖产氢。以糖含量为16.6 g/L的50.0% (V/V) 的藻类水解液为底物产氢时，其HMF去除率达到100%[80]。
Xu等将驯化后的M. guilliermondii SC1103细胞包裹在海藻酸钠微球中，以其为生物催化剂还原HMF为BHMF。当底物浓度为200–300 mmol/L时，在7–24 h内BHMF得率能达到82.0%–85.0%，选择性达到99.0%。此外，固定化后的细胞重复使用4次后，细胞存活率仍能达到70.0%，细胞的催化活性及其HMF耐受性水平在驯化和固定化时显著提高[81]。
Singh等从土壤中分离出一株Bordetella sp. BTIITR，它可以选择性地从木质纤维素水解液中降解HMF和糠醛且耗糖量少[10]。随后其所在团队尝试将Bordetella sp. BTIITR固定在壳聚糖珠中，并利用固定化后的Bordetella sp. BTIITR对水解液进行解毒，发现固定的细胞能够在pH 8.0和40 ℃的条件下，在20 h内分别从模拟水解液和甘蔗渣水解液中降解93.0% HMF+100%糠醛和86.0% HMF+100%糠醛。此外，固定的细胞较游离的细胞能够在操作范围更广的pH、温度和抑制剂浓度下解毒木质纤维素水解液，固定的细胞在经过7次的循环使用后仍具有较好的生物活性[78]。
由以上实例可以看出，借助各种功能化材料的辅助，可创建更适合微生物生长的微环境，增强微生物对呋喃醛等木质纤维素水解液抑制物的脱除能力。目前，功能材料辅助微生物脱毒正处于研究起步阶段，但势必会成为未来研究热点。
4 展望利用可再生的木质纤维生物质资源替代化石能源生产生物燃料和高值化学品是当前新能源领域研究的重点，也是实现社会可持续发展的需求，而预处理后产生的呋喃醛等多种抑制物严重制约着木质纤维素的生物转化效率，是木质纤维素生物炼制的主要瓶颈之一[82]。生物法脱除抑制物具有很多化学法和物理法无可比拟的优点，多年来一直是生物炼制领域的研究热点。传统的进化工程和代谢工程为获得具有呋喃醛耐受能力和降解能力的优良菌株提供了强大的技术支撑平台，而近年来逐步受到重视的微生物共培养系统和功能化材料辅助微生物脱除呋喃醛抑制物也必将发展为进一步提高菌株的抑制物代谢能力的有效方法。此外，糠醛和HMF的不完全代谢产物(呋喃醇和呋喃羧酸) 也都是具有重要应用价值的化合物，在关注微生物脱除呋喃醛抑制物的同时，实现抑制物的价值提升也是一个前景广阔的研究领域。
参考文献
| [1] | 陈洪章. 纤维素生物技术: 理论与实践. 北京: 化学工业出版社, 2014. Chen HZ. Biotechnology of Lignocellulose: Theory and Practice. Beijing: Chemical Industry Press, 2014 (in Chinese). |
| [2] | 刘彬. 碱性预处理抑制物对酶解和发酵抑制作用机理[D]. 济南: 齐鲁工业大学, 2019. Liu B. Inhibitory mechanism of alkaline pretreatment inhibitors on enzymatic hydrolysis and fermentation[D]. Ji'nan: Qilu University of Technology, 2019 (in Chinese). |
| [3] | Almeida JRM, Modig T, Petersson A, et al. Increased tolerance and conversion of inhibitors in lignocellulosic hydrolysates by Saccharomyces cerevisiae. J Chem Technol Biotechnol, 2007, 82(4): 340-349. DOI:10.1002/jctb.1676 |
| [4] | Wang SZ, Sun XX, Yuan QP. Strategies for enhancing microbial tolerance to inhibitors for biofuel production: a review. Bioresour Technol, 2018, 258: 302-309. DOI:10.1016/j.biortech.2018.03.064 |
| [5] | 张小欢, 张宇, 王琼, 等. 微生物耐受糠醛机制的研究进展. 新能源进展, 2017, 5(3): 89-196. Zhang XH, Zhang Y, Wang Q, et al. Research and development on the microbial furfural-tolerance mechanism. J Circ Syst, 2017, 5(3): 89-196 (in Chinese). |
| [6] | López MJ, Nichols NN, Dien BS, et al. Isolation of microorganisms for biological detoxification of lignocellulosic hydrolysates. Appl Microbiol Biotechnol, 2004, 64(1): 125-131. DOI:10.1007/s00253-003-1401-9 |
| [7] | Shibuya M, Sasaki K, Tanaka Y, et al. Development of combined nanofiltration and forward osmosis process for production of ethanol from pretreated rice straw. Bioresour Technol, 2017, 235: 405-410. DOI:10.1016/j.biortech.2017.03.158 |
| [8] | Trinh LTP, Kundu C, Lee JW, et al. An integrated detoxification process with electrodialysis and adsorption from the hemicellulose hydrolysates of yellow poplars. Bioresour Technol, 2014, 161: 280-287. DOI:10.1016/j.biortech.2014.03.042 |
| [9] | Xie R, Tu MB, Carvin J, et al. Detoxification of biomass hydrolysates with nucleophilic amino acids enhances alcoholic fermentation. Bioresour Technol, 2015, 186: 106-113. DOI:10.1016/j.biortech.2015.03.030 |
| [10] | Singh B, Verma A, Pooja, et al. A biotechnological approach for degradation of inhibitory compounds present in lignocellulosic biomass hydrolysate liquor using Bordetella sp. BTⅡTR. Chem Eng J, 2017, 328: 519-526. DOI:10.1016/j.cej.2017.07.059 |
| [11] | Taherzadeh MJ, Karimi K. Fermentation inhibitors in ethanol processes and different strategies to reduce their effects. Biofuels, 2011, 287-311. DOI:10.1016/B978-0-12-385099-7.00012-7 |
| [12] | Ximenes E, Kim Y, Mosier N, et al. Deactivation of cellulases by phenols. Enzyme Microb Technol, 2011, 48(1): 54-60. DOI:10.1016/j.enzmictec.2010.09.006 |
| [13] | 王丹, 王洪辉, 王競, 等. 糠醛和5-羟甲基糠醛对大肠杆菌产丁二酸的影响. 生物工程学报, 2013, 29(10): 1463-1472. Wang D, Wang HH, Wang J, et al. Effects of furfural and 5-hydroxymethylfurfural on succinic acid production by Escherichia coli. Chin J Biotech, 2013, 29(10): 1463-1472 (in Chinese). |
| [14] | 雍子涵, 张栩, 谭天伟. 糠醛对粘红酵母生长与油脂积累的影响. 生物工程学报, 2015, 31(10): 1484-1491. Yong ZH, Zhang X, Tan TW. Effects of furfural on the growth and lipid production of oleaginous yeast Rhodotorula glutinis. Chin J Biotech, 2015, 31(10): 1484-1491 (in Chinese). |
| [15] | Barakat A, Monlau F, Steyer JP, et al. Effect of lignin-derived and furan compounds found in lignocellulosic hydrolysates on biomethane production. Bioresour Technol, 2012, 104: 90-99. DOI:10.1016/j.biortech.2011.10.060 |
| [16] | 余强, 庄新姝, 袁振宏, 等. 高温液态水中甜高梁渣半纤维素水解及其机理. 化工学报, 2012, 63(2): 599-605. Yu Q, Zhuang XS, Yuan ZH, et al. Hydrolysis of sweet sorghum bagasse hemicellulose with liquid hot water and its mechanism. CIESC J, 2012, 63(2): 599-605 (in Chinese). DOI:10.3969/j.issn.0438-1157.2012.02.037 |
| [17] | 吕双亮, 庄新姝, 余强, 等. 渗滤与间歇高温液态水预处理甘蔗渣的对比研究. 太阳能学报, 2015, 36(3): 664-670. Lv SL, Zhuang XS, Yu Q, et al. Comparison of liquid hot water pretreatment with flow-through and batch reactor for sugarcane bagasse. Acta Energ Solaris Sin, 2015, 36(3): 664-670 (in Chinese). DOI:10.3969/j.issn.0254-0096.2015.03.024 |
| [18] | Villarreal MLM, Prata AMR, Felipe MGA, et al. Detoxification procedures of eucalyptus hemicellulose hydrolysate for xylitol production by Candida guilliermondii. Enzyme Microb Technol, 2006, 40(1): 17-24. DOI:10.1016/j.enzmictec.2005.10.032 |
| [19] | Alriksson B, Cavka A, J?nsson LJ. Improving the fermentability of enzymatic hydrolysates of lignocellulose through chemical in-situ detoxification with reducing agents. Bioresour Technol, 2011, 102(2): 1254-1263. DOI:10.1016/j.biortech.2010.08.037 |
| [20] | Back SW, Kim JS, Park YK, et al. The effect of sugar decomposed on the ethanol fermentation and decomposition reactions of sugars. Biotechnol Bioproc Eng, 2008, 13(3): 332-341. DOI:10.1007/s12257-007-0161-2 |
| [21] | Wang SZ, He ZJ, Yuan QP. Xylose enhances furfural tolerance in Candida tropicalis by improving NADH recycle. Chem Eng Sci, 2017, 158: 37-40. DOI:10.1016/j.ces.2016.09.026 |
| [22] | Hadi SM, Shahabuddin, Rehman A. Specificity of the interaction of furfural with DNA. Mutat Res Lett, 1989, 225(3): 101-106. DOI:10.1016/0165-7992(89)90125-5 |
| [23] | Allen SA, Clark W, McCaffery JM, et al. Furfural induces reactive oxygen species accumulation and cellular damage in Saccharomyces cerevisiae. Biotechnol Biofuels, 2010, 3: 2. DOI:10.1186/1754-6834-3-2 |
| [24] | Trudgill PW. The metabolism of 2-furoic acid by Pseudomonas F2. Biochem J, 1969, 113(4): 577-587. DOI:10.1042/bj1130577 |
| [25] | Wierckx N, Koopman F, Bandounas L, et al. Isolation and characterization of Cupriavidus basilensis HMF14 for biological removal of inhibitors from lignocellulosic hydrolysate. Microb Biotechnol, 2010, 3(3): 336-343. DOI:10.1111/j.1751-7915.2009.00158.x |
| [26] | Koopman F, Wierckx N, De Winde JH, et al. Identification and characterization of the furfural and 5-(hydroxymethyl)furfural degradation pathways of Cupriavidus basilensis HMF14. Proc Natl Acad Sci USA, 2010, 107(11): 4919-4924. DOI:10.1073/pnas.0913039107 |
| [27] | Nichols NN, Sharma LN, Mowery RA, et al. Fungal metabolism of fermentation inhibitors present in corn stover dilute acid hydrolysate. Enzyme Microb Technol, 2008, 42(7): 624-630. DOI:10.1016/j.enzmictec.2008.02.008 |
| [28] | 王晓凤, 张建, 辛秀娟, 等. 丝状真菌Amorphotheca resinae ZN1的糠醛降解代谢分析. 生物工程学报, 2012, 28(9): 1070-1079. Wang XF, Zhang J, Xin XJ, et al. Furfural degradation by filamentous fungus Amorphotheca resinae ZN1. Chin J Biotech, 2012, 28(9): 1070-1079 (in Chinese). |
| [29] | 王晓凤. 生物脱毒真菌Amorphotheca resinae ZN1的生长特性及呋喃衍生物降解途径确定的研究[D]. 上海: 华东理工大学, 2012. Wang XF. Study of growth characteristics and metabolic mechanism of furan degradation by the biodetoxification fungus Amorphotheca resinae ZN1[D]. Shanghai: East China University of Science and Technology, 2012 (in Chinese). |
| [30] | 冉红. 树脂枝孢霉菌Amorphotheca resinae ZN1生物降解木质纤维素来源呋喃醛和芳香醛的实验研究[D]. 上海: 华东理工大学, 2014. Rang H. Experimental study of biodegradation of furan aldehydes and aromatic aldehydes derived from lignocellulose by Amorphotheca resinae ZN1[D]. Shanghai: East China University of Science and Technology, 2014 (in Chinese). |
| [31] | 王霞. 呋喃醛抑制物的生物脱毒机理解析及高抗逆生物炼制发酵工程菌株的构建[D]. 上海: 华东理工大学, 2017. Wang X. Study on biodetoxification mechanism of furan aldehyde inhibitors and the construction robust biorefinery fermenting strains[D]. Shanghai: East China University of Science and Technology, 2017 (in Chinese). |
| [32] | Miller EN, Jarboe LR, Yomano LP, et al. Silencing of NADPH-dependent oxidoreductase genes (yqhD and dkgA) in furfural-resistant ethanologenic Escherichia coli. Appl Environ Microbiol, 2009, 75(13): 4315-4323. DOI:10.1128/AEM.00567-09 |
| [33] | Jarboe LR. YqhD: a broad-substrate range aldehyde reductase with various applications in production of biorenewable fuels and chemicals. Appl Microbiol Biotechnol, 2011, 89(2): 249-257. DOI:10.1007/s00253-010-2912-9 |
| [34] | Ishii J, Yoshimura K, Hasunuma T, et al. Reduction of furan derivatives by overexpressing NADH-dependent adh1 improves ethanol fermentation using xylose as sole carbon source with Saccharomyces cerevisiae harboring XR-XDH pathway. Appl Microbiol Biotechnol, 2013, 97(6): 2597-2607. DOI:10.1007/s00253-012-4376-6 |
| [35] | Tsuge Y, Kudou M, Kawaguchi H, et al. FudC, a protein primarily responsible for furfural detoxification in Corynebacterium glutamicum. Appl Microbiol Biotechnol, 2016, 100(6): 2685-2692. DOI:10.1007/s00253-015-7115-y |
| [36] | Li YM, Zhang XY, Li N, et al. Biocatalytic reduction of HMF to 2, 5-bis (hydroxymethyl) furan by HMF-tolerant whole cells. ChemSusChem, 2017, 10(2): 304. DOI:10.1002/cssc.201700004 |
| [37] | Yan YX, Bu CY, Huang X, et al. Efficient whole-cell biotransformation of furfural to furfuryl alcohol by Saccharomyces cerevisiae NL22. J Chem Technol Biotechnol, 2019, 94(12): 3825-3831. DOI:10.1002/jctb.6177 |
| [38] | Yan YX, Bu CY, He Q, et al. Efficient bioconversion of furfural to furfuryl alcohol by Bacillus coagulans NL01. RSC Adv, 2018, 8(47): 26720-26727. DOI:10.1039/C8RA05098H |
| [39] | Zhang XY, Zong MH, Li N. Whole-cell biocatalytic selective oxidation of 5-hydroxymethylfurfural to 5-hydroxymethyl-2-furancarboxylic acid. Green Chem, 2017, 19(9): 4544-4551. |
| [40] | Sayed M, Pyo SH, Rehnberg N, et al. Selective oxidation of 5-hydroxymethylfurfural to 5-hydroxymethyl-2-furancarboxylic acid using Gluconobacter oxydans. ACS Sustainable Chem Eng, 2019, 7(4): 4406-4413. DOI:10.1021/acssuschemeng.8b06327 |
| [41] | Cang R, Shen LQ, Yang G, et al. Highly selective oxidation of 5-hydroxymethylfurfural to 5-hydroxymethyl-2-furancarboxylic acid by a robust whole-cell biocatalyst. Catalysts, 2019, 9(6): 526-541. DOI:10.3390/catal9060526 |
| [42] | Liu P, Xie JX, Tan HH, et al. Valorization of Gelidium amansii for dual production of D-galactonic acid and 5-hydroxymethyl-2- furancarboxylic acid by chemo-biological approach. Microb Cell Fact, 2020, 19: 104-113. DOI:10.1186/s12934-020-01357-6 |
| [43] | Zhang XY, Ou XY, Fu YJ, et al. Efficient synthesis of 5-hydroxymethyl-2-furancarboxylic acid by Escherichia coli overexpressing aldehyde dehydrogenases. J Biotechnol, 2020, 307: 125-130. DOI:10.1016/j.jbiotec.2019.11.007 |
| [44] | Jia HY, Zong MH, Yu HL, et al. Dehydrogenase-catalyzed oxidation of furanics: exploitation of hemoglobin catalytic promiscuity. ChemSusChem, 2017, 10(18): 3524-3528. DOI:10.1002/cssc.201701288 |
| [45] | Peng B, Ma CL, Zhang PQ, et al. An effective hybrid strategy for converting rice straw to furoic acid by tandem catalysis via Sn-sepiolite combined with recombinant E. coli whole cells harboring horse liver alcohol dehydrogenase. Green Chem, 2019, 21(21): 5914-5923. DOI:10.1039/C9GC02499A |
| [46] | Kumar H, Fraaije MW. Conversion of furans by Baeyer-Villiger monooxygenases. Catalysts, 2017, 7(6): 179-188. DOI:10.3390/catal7060179 |
| [47] | Cao GL, Ximenes E, Nichols NN, et al. Biological abatement of cellulase inhibitors. Bioresour Technol, 2013, 146: 604-610. DOI:10.1016/j.biortech.2013.07.112 |
| [48] | Pienkos PT, Zhang M. Role of pretreatment and conditioning processes on toxicity of lignocellulosic biomass hydrolysates. Cellulose, 2009, 16(4): 743-762. DOI:10.1007/s10570-009-9309-x |
| [49] | Okuda N, Soneura M, Ninomiya K, et al. Biological detoxification of waste house wood hydrolysate using Ureibacillus thermosphaericus for bioethanol production. J Biosci Bioeng, 2008, 106(2): 128-133. DOI:10.1263/jbb.106.128 |
| [50] | Guedes FB, Fernandes B, Brambilla RRDCL, et al. Microaerobic biodegradability of sugarcane hemicellulosic hydrolysate toxic compounds by Issatchenkia yeasts strains. New Biotechnol, 2009, 25(S1): S65-S66. |
| [51] | Tsuge Y, Hori Y, Kudou M, et al. Detoxification of furfural in Corynebacterium glutamicum under aerobic and anaerobic conditions. Appl Microbiol Biotechnol, 2014, 98(20): 8675-8683. DOI:10.1007/s00253-014-5924-z |
| [52] | Xu QQ, Zheng ZJ, Zou LH, et al. A versatile Pseudomonas putida KT2440 with new ability: selective oxidation of 5-hydroxymethylfurfural to 5-hydroxymethyl-2-furancarboxylic acid. Bioprocess Biosys Eng, 2020, 43(1): 67-73. DOI:10.1007/s00449-019-02205-7 |
| [53] | 王娟. 皮状丝孢酵母纤维素油脂发酵及抑制物耐受机制研究[D]. 上海: 华东理工大学, 2016. Wang J. Cellulosic lipid fermentation by Trichosporon cutaneum and inhibitors tolerance mechanism study[D]. Shanghai: East China University of Science and Technology, 2016 (in Chinese). |
| [54] | Almeida JRM, Modig T, R?der A, et al. Pichia stipitis xylose reductase helps detoxifying lignocellulosic hydrolysate by reducing 5-hydroxymethyl-furfural (HMF). Biotechnol Biofuels, 2008, 1: 12. DOI:10.1186/1754-6834-1-12 |
| [55] | He MX, Wu B, Shui ZX, et al. Transcriptome profiling of Zymomonas mobilis under furfural stress. Appl Microbiol Biotechnol, 2012, 95(1): 189-199. DOI:10.1007/s00253-012-4155-4 |
| [56] | J?nsson LJ, Palmqvist E, Nilvebrant NO, et al. Detoxification of wood hydrolysates with laccase and peroxidase from the white-rot fungus Trametes versicolor. Appl Microbiol Biotechnol, 1998, 49(6): 691-697. DOI:10.1007/s002530051233 |
| [57] | Saravanakumar T, Park HS, Mo AY, et al. Detoxification of furanic and phenolic lignocellulose derived inhibitors of yeast using laccase immobilized on bacterial cellulosic nanofibers. J Mol Catal B Enzym, 2016, 134: 196-205. DOI:10.1016/j.molcatb.2016.11.006 |
| [58] | Mohamed ET, Wang SZ, Lennen RM, et al. Generation of a platform strain for ionic liquid tolerance using adaptive laboratory evolution. Microb Cell Fact, 2017, 16: 204-219. DOI:10.1186/s12934-017-0819-1 |
| [59] | Sridhar M, Sree NK, Rao LV. Effect of UV radiation on thermotolerance, ethanol tolerance and osmotolerance of Saccharomyces cerevisiae VS1 and VS3 strains. Bioresour Technol, 2002, 83(3): 199-202. DOI:10.1016/S0960-8524(01)00221-8 |
| [60] | Qin DD, Hu YL, Cheng J, et al. An auto-inducible Escherichia coli strain obtained by adaptive laboratory evolution for fatty acid synthesis from ionic liquid-treated bamboo hydrolysate. Bioresour Technol, 2016, 221: 375-384. DOI:10.1016/j.biortech.2016.09.024 |
| [61] | 丁明珠. 酵母对纤维素水解液中复合抑制剂耐受的系统分析与解耦[D]. 天津: 天津大学, 2011. Ding MZ. Systematic analysis and decoupling of yeast tolerance to combined inhibitors in lignocellulose hydrolysates[D]. Tianjin: Tianjin University, 2011 (in Chinese). |
| [62] | Shui ZX, Qin H, Wu B, et al. Adaptive laboratory evolution of ethanologenic Zymomonas mobilis strain tolerant to furfural and acetic acid inhibitors. Appl Microbiol Biotechnol, 2015, 99(13): 5739-5748. DOI:10.1007/s00253-015-6616-z |
| [63] | Wang X, Miller EN, Yomano LP, et al. Increased furfural tolerance due to overexpression of NADH-dependent oxidoreductase fucO in Escherichia coli strains engineered for the production of ethanol and lactate. Appl Environ Microbiol, 2011, 77(15): 5132-5140. DOI:10.1128/AEM.05008-11 |
| [64] | Seo HM, Jeon JM, Lee JH, et al. Combinatorial application of two aldehyde oxidoreductases on isobutanol production in the presence of furfural. J Ind Microbiol Biotechnol, 2016, 43(1): 37-44. DOI:10.1007/s10295-015-1718-2 |
| [65] | Miller EN, Jarboe LR, Turner PC, et al. Furfural inhibits growth by limiting sulfur assimilation in ethanologenic Escherichia coli strain LY180. Appl Environ Microbiol, 2009, 75(19): 6132-6141. DOI:10.1128/AEM.01187-09 |
| [66] | Wang X, Miller EN, Yomano LP, et al. Increased furan tolerance in Escherichia coli due to a cryptic ucpA gene. Appl Environ Microbiol, 2012, 78(7): 2452-2455. DOI:10.1128/AEM.07783-11 |
| [67] | Laadan B, Wallace-Salinas V, Carlsson ?J, et al. Furaldehyde substrate specificity and kinetics of Saccharomyces cerevisiae alcohol dehydrogenase 1 variants. Microb Cell Fac, 2014, 13: 112. DOI:10.1186/s12934-014-0112-5 |
| [68] | Bowman MJ, Jordan DB, Vermillion KE, et al. Stereochemistry of furfural reduction by a Saccharomyces cerevisiae aldehyde reductase that contributes to in situ furfural detoxification. Appl Environ Microbiol, 2010, 76(15): 4926-4932. DOI:10.1128/AEM.00542-10 |
| [69] | Zhao XX, Tang J, Wang X, et al. YNL134C from Saccharomyces cerevisiae encodes a novel protein with aldehyde reductase activity for detoxification of furfural derived from lignocellulosic biomass. Yeast, 2015, 32(5): 409-422. DOI:10.1002/yea.3068 |
| [70] | Gorsich SW, Dien BS, Nichols NN, et al. Tolerance to furfural-induced stress is associated with pentose phosphate pathway genes zwf1, gnd1, rpe1, and tkl1 in Saccharomyces cerevisiae. Appl Microbiol Biotechnol, 2006, 71(3): 339-449. DOI:10.1007/s00253-005-0142-3 |
| [71] | Heer D, Heine D, Sauer U. Resistance of Saccharomyces cerevisiae to high concentrations of furfural is based on NADPH-dependent reduction by at least two oxireductases. Appl Environ Microbiol, 2009, 75(24): 7631-7638. DOI:10.1128/AEM.01649-09 |
| [72] | Park S, Koo HM, Park YK, et al. Expression of aldehyde dehydrogenase 6 reduces inhibitory effect of furan derivatives on cell growth and ethanol production in Saccharomyces cerevisiae. Bioresour Technol, 2011, 102(10): 6033-6038. DOI:10.1016/j.biortech.2011.02.101 |
| [73] | Xiao H, Zhao HM. Genome-wide RNAi screen reveals the E3 SUMO-protein ligase gene siz1 as a novel determinant of furfural tolerance in Saccharomyces cerevisiae. Biotechnol Biofuels, 2014, 7: 78. DOI:10.1186/1754-6834-7-78 |
| [74] | Turner WJ, Dunlop MJ. Trade-offs in improving biofuel tolerance using combinations of efflux pumps. ACS Synth Biol, 2014, 4(10): 1056-1063. |
| [75] | Ruegg TL, Kim EM, Simmons BA, et al. An auto-inducible mechanism for ionic liquid resistance in microbial biofuel production. Nat Commun, 2014, 5: 3490. DOI:10.1038/ncomms4490 |
| [76] | Boopathy R, Bokang H, Daniels L. Biotransformation of furfural and 5-hydroxymethyl furfural by enteric bacteria. J Ind Microbiol, 1993, 11(3): 147-150. DOI:10.1007/BF01583715 |
| [77] | Zhu JQ, Li X, Qin L, et al. In situ detoxification of dry dilute acid pretreated corn stover by co-culture of xylose-utilizing and inhibitor-tolerant Saccharomyces cerevisiae increases ethanol production. Bioresour Technol, 2016, 218: 380-387. DOI:10.1016/j.biortech.2016.06.107 |
| [78] | Singh B, Kumar P, Yadav A, et al. Degradation of fermentation inhibitors from lignocellulosic hydrolysate liquor using immobilized bacterium, Bordetella sp. BTⅡTR. Chem Eng J, 2019, 361: 1152-1160. DOI:10.1016/j.cej.2018.12.168 |
| [79] | Nguyen DTT, Praveen P, Loh KC. Zymomonas mobilis immobilization in polymeric membranes for improved resistance to lignocellulose-derived inhibitors in bioethanol fermentation. Biochem Eng J, 2018, 140: 29-37. DOI:10.1016/j.bej.2018.09.003 |
| [80] | Kumar G, Sivagurunathan P, Kobayashi T, et al. Simultaneous removal of 5-hydroxy methyl furfural (5-HMF) and hydrogen production from acid (H2SO4) pretreated red-algal hydrolysate via hybrid immobilized cells. Algal Res, 2015, 11: 326-333. DOI:10.1016/j.algal.2015.07.015 |
| [81] | Xu ZH, Cheng AD, Xing XP, et al. Improved synthesis of 2, 5-bis (hydroxymethyl) furan from 5-hydroxymethylfurfural using acclimatized whole cells entrapped in calcium alginate. Bioresour Technol, 2018, 262: 177-183. DOI:10.1016/j.biortech.2018.04.077 |
| [82] | 顾翰琦, 刘冉, 邵玲智, 等. 酿酒酵母对酚类抑制物耐受性研究. 生物技术通报, 2020, 36(6): 136-142. Gu HQ, Liu R, Shao LZ, et al. Study on the tolerance of Saccharomyces cerevisiae strain to phenolic inhibitors. Biotechnol Bull, 2020, 36(6): 136-142 (in Chinese). |
