生物工程学报  2023, Vol. 39 Issue (5): 1976-1986
http://dx.doi.org/10.13345/j.cjb.230010
中国科学院微生物研究所、中国微生物学会主办
0

文章信息

曾彩婷, 纪俊宾, 丁方慧, 李周坤, 曹慧, 崔中利, 闫新
ZENG Caiting, JI Junbin, DING Fanghui, LI Zhoukun, CAO Hui, CUI Zhongli, YAN Xin
一株聚酯型聚氨酯降解菌高地芽孢杆菌YX8-1的分离及鉴定
Isolation and identification of a polyester-polyurethane degrading bacterium Bacillus altitudinis YX8-1
生物工程学报, 2023, 39(5): 1976-1986
Chinese Journal of Biotechnology, 2023, 39(5): 1976-1986
10.13345/j.cjb.230010

文章历史

Received: January 5, 2023
Accepted: March 20, 2023
一株聚酯型聚氨酯降解菌高地芽孢杆菌YX8-1的分离及鉴定
曾彩婷 , 纪俊宾 , 丁方慧 , 李周坤 , 曹慧 , 崔中利 , 闫新     
南京农业大学生命科学学院, 江苏 南京 210095
摘要:聚氨酯(polyurethane, PUR)塑料在日常生活中发挥着重要作用,但同时PUR废弃物也带来严重的环境污染问题。生物(酶)降解是一种环境友好、成本低廉的PUR废弃物回收方法,其关键在于获得高效的降解菌株或酶。本研究从垃圾填埋场的聚氨酯废弃物表面分离出了一株聚酯型PUR降解菌株YX8-1。基于菌落和显微形态观察、16S rDNA和DNA旋转酶(DNA gyrase)基因gyrA系统发育分析及基因组序列比对,将该菌鉴定为高地芽孢杆菌(Bacillus altitudinis)。高效液相色谱(high performance liquid chromatography, HPLC)及液相色谱-质谱联用(liquid chromatography-tandem mass spectrometry, LC-MS/MS)结果显示菌株YX8-1能降解自行合成的聚酯型PUR寡聚物(PBA-PU),并产生单体化合物4, 4′-亚甲基二苯胺。此外,菌株YX8-1能在30 d内使商品化的聚酯型PUR海绵失重32%。本研究为PUR废弃物的生物降解提供了菌株资源,也为挖掘相关降解酶打下了基础。
关键词聚氨酯塑料    生物降解    塑料废弃物回收    高地芽孢杆菌    
Isolation and identification of a polyester-polyurethane degrading bacterium Bacillus altitudinis YX8-1
ZENG Caiting , JI Junbin , DING Fanghui , LI Zhoukun , CAO Hui , CUI Zhongli , YAN Xin     
College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, Jiangsu, China
Abstract: Although polyurethane (PUR) plastics play important roles in daily life, its wastes bring serious environmental pollutions. Biological (enzymatic) degradation is considered as an environmentally friendly and low-cost method for PUR waste recycling, in which the efficient PUR-degrading strains or enzymes are crucial. In this work, a polyester PUR-degrading strain YX8-1 was isolated from the surface of PUR waste collected from a landfill. Based on colony morphology and micromorphology observation, phylogenetic analysis of 16S rDNA and gyrA gene, as well as genome sequence comparison, strain YX8-1 was identified as Bacillus altitudinis. The results of high performance liquid chromatography (HPLC) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) showed that strain YX8-1 was able to depolymerize self-synthesized polyester PUR oligomer (PBA-PU) to produce a monomeric compound 4, 4′-methylene diphenylamine. Furthermore, strain YX8-1 was able to degrade 32% of the commercialized polyester PUR sponges within 30 days. This study thus provides a strain capable of biodegradation of PUR waste, which may facilitate the mining of related degrading enzymes.
Keywords: polyurethane plastics    biodegradation    plastic waste recycling    Bacillus altitudinis    

聚氨酯(polyurethane, PUR)由多元醇和异氰酸酯聚合而成,改变多元醇、异氰酸酯的种类及其比例将合成不同的PUR,根据多元醇类型的不同可大致将PUR区分为聚酯型和聚醚型两类[1-2]。PUR塑料因其耐用性和生产成本低等优点被广泛应用于建筑、生物医药、服饰、家电和交通等领域。此外PUR塑料常用于生产海绵,良好的保护和缓冲特性使其成为包装、汽车、建筑、医疗保健和运动等领域的优良选择[3]。PUR塑料的生产始于20世纪40年代,据统计2020年我国PUR产量达到1 450万t[4]。伴随着PUR的大量使用,产生的PUR废弃物堆积在自然环境中使得海洋和土壤受到严重污染,危害动植物及人类健康。由于PUR废弃物在自然环境中降解缓慢且难以回收利用,导致绝大部分PUR废弃物被送往垃圾场进行填埋及焚烧,造成了资源的浪费且产生的有毒气体将危害人体健康[5]。此外,PUR废弃物经磨损或某些化学过程处理后将产生直径小于5 mm的微塑料。据报道,微塑料被某些动物食用后将通过食物链传播至人体内威胁海洋生物及人类的生存[6],目前已在人类器官及胎盘中检测到微塑料的存在。因此,急需寻找高效的PUR废弃物处理方法。

常用的PUR废弃物处理方法主要包括焚烧、填埋、机械回收、化学处理及生物降解[7]。其中利用微生物或酶进行生物降解是公认的安全环保的方法[8],目前已报道多株PUR降解菌株。例如,Khan等[9]分离出一株塔宾曲霉(Aspergillus tubingensis),将该菌与聚酯型PUR薄膜在含2% (质量体积分数)葡萄糖的无机盐培养基中孵育2个月后,聚酯型PUR薄膜被降解成小块,降解率高达90%。Álvarez-Barragán等[10]分离出8株PUR降解菌株,其中细枝孢霉(Cladosporium tenuissimum) A2.PP.5在21 d内降解了65.3%不含阻燃剂的聚酯型PUR海绵。此外,酵母、黄曲霉和青霉等也被报道具有PUR降解能力[11-14]。然而,目前分离到的PUR降解菌株中大部分为真菌,有关细菌降解PUR的报道相对较少,且降解效果相对较差。Shah等[15]分离出枯草芽孢杆菌(Bacillus subtilis) MZA-75及铜绿假单胞菌(Pseudomonas aeruginosa) MZA-85,将2株菌单独培养时30 d内聚酯型PUR降解率为20%,混合培养时聚酯型PUR降解率为40%。Roy等[16]从固体废弃物处理厂采集的土样中分离出假单胞菌(Pseudomonas sp.) AKS31,该菌能在10 d内降解约70%的聚酯型PUR薄膜,且通过高效液相色谱可检测到产物二甘醇。

在PUR降解酶的挖掘方面,Akutsu等[17]从菌株食酸丛毛单胞菌(Comamonas acidovorans) TB-35克隆出基因pudA,其编码的酯酶能够水解聚酯型PUR的酯键,生成二甘醇和己二酸。Stern等[18]和Howard等[19]分别从绿针假单胞菌(Pseudomonas chlororaphis)中成功纯化出能够水解聚酯型PUR中酯键的脂肪酶PueA和PueB。此外,Schmidt等[20]发现角质酶LCC及TfCut2不仅能够降解聚对苯二甲酸乙二醇酯(polyethylene terephthalate, PET),还具有聚酯型PUR降解能力。Magnin等[21]研究了南极念珠菌脂肪酶B (Candida antarctica lipase B, CALB)对4种不同聚酯型PUR的降解能力,并将50%降解产物与50% 6-羟基丙酸混合获得了74 000 g/mol无毒聚合物,实现了PUR塑料的循环利用。

尽管已有多种PUR降解菌株及降解酶被报道,但这些菌株和酶资源仍难以满足PUR降解的应用需求,还需要进一步挖掘或改造新的降解菌株及酶。本研究从广德垃圾处理厂的PUR废弃物表面分离出一株PUR高效降解菌株YX8-1,通过系统发育分析、降解产物检测、失重率测定和电子显微镜观察等方法,探究了该菌的系统发育地位并表征了其对合成PUR塑料PBA-PU及商业化PUR的降解特性。

1 材料与方法 1.1 材料 1.1.1 菌株及PUR塑料

本研究所用菌株分离自广德垃圾处理厂的PUR废弃物表面。筛选聚氨酯降解菌株所用的水性聚氨酯Impranil DLN购买自科思创聚合物有限公司,PBA-PU由南京工业大学周杰老师惠赠[22],商业化PUR塑料购自南通大工有限公司。

1.1.2 培养基

低盐LB培养基:10 g/L蛋白胨,5 g/L酵母粉,5 g/L NaCl,固体培养基添加15 g/L琼脂粉。

无机盐培养基:0.92 g/L K2HPO4.3H2O,0.7 g/L KH2PO4,0.7 g/L MgSO4.7H2O,1.0 g/L NH4NO3,0.005 g/L NaCl,0.002 g/L FeSO4.7H2O,0.002 g/L ZnSO4.7H2O,0.001 g/L MnSO4.H2O,pH 7.0。

富集培养基:含5% (体积分数) LB的无机盐培养基中添加1% (体积分数) Impranil DLN。

筛选培养基:LB固体培养基中添加1% (体积分数) Impranil DLN。

固体PBA-PU降解培养基:0.1 g PBA-PU溶于5 mL二甲基亚砜(dimethyl sulfoxide, DMSO)后,加至100 mL LB固体培养基中。

液体PBA-PU降解培养基:0.1 g PBA-PU溶于5 mL DMSO后,加至含5% (体积分数) LB的无机盐培养基中。

商业化PUR降解培养基:称取0.2 g PUR海绵置于含5% (体积分数) LB的无机盐培养基中,121 ℃灭菌30 min。

1.2 方法 1.2.1 聚氨酯降解菌株的分离

取适量广德垃圾处理厂的PUR废弃物用10 mL超纯水冲洗表面土样,随后按10% (体积分数)的接种量接种至富集培养基中,置于30 ℃、180 r/min摇床振荡培养1周后取10 mL富集培养液转接至新鲜100 mL富集培养基中继续富集1周,重复转接2次。富集完成后取富集液稀释至10−4−10−7倍梯度,涂布于筛选平板,30 ℃培养约1周后取出,挑取有水解圈的菌株接种至新鲜的筛选平板上观察是否再次出现水解圈。将再次出现水解圈的菌株进行稀释涂布纯化,最终纯化获得Impranil DLN降解菌株,命名为YX8-1。

1.2.2 菌株形态观察

观察菌株YX8-1在Impranil DLN平板上的颜色、形状、透明度、隆起程度和边缘状况,并通过革兰氏染色观察菌株形态。

1.2.3 16S rDNA序列鉴定及系统发育分析

用细菌基因组DNA提取试剂盒[生工生物工程(上海)股份有限公司]提取降解菌株的基因组DNA,通过16S rDNA通用引物27F (5′-AGA GTTTGATCCTGGCTCAG-3′)和1492R (5′-TAC GGCTACCTTGTTACGACTT-3′)扩增其16S rDNA序列,将获得的PCR产物送至公司测序。最终将测序结果提交至NCBI数据库进行比对,并通过软件MEGA-X的邻接法(neighbor-joining, NJ)方法构建系统发育进化树。

通过引物gyrA-F (5′-GCDGCHGCNATGC GTTAYAC-3′)和gyrA-R (5′-ACAAGMTCWGC KATTTTTTC-3′)扩增gyrA序列,将获得的PCR产物送至公司测序,最终将得到的gyrA序列提交至NCBI数据库进行比对,通过软件MEGA-X的NJ法构建系统发育进化树。

对比分析后下载与分离菌株最相近的模式菌株的基因组序列,计算2个基因组之间的平均核酸统一性(average nucleic acid uniformity, ANI) (http://www.ezbiocloud.net/tools/ani)[23]和基因组间的DNA-DNA杂交值(dDDH) (http://ggdc.dsmz.de/ggdc.php)[24]

1.2.4 PBA-PU降解特性分析

将菌株YX8-1划至固体PBA-PU降解平板30 ℃过夜培养后取出观察是否产生水解圈;将菌悬液接入30 mL液体PBA-PU降解培养基中至OD600=1,30 ℃、180 r/min摇床振荡培养15 d后取5 mL菌液,10 000×g离心10 min取上清。用盐酸调节至pH 2.0后用2倍体积乙酸乙酯进行萃取,取上层溶液置于通风橱风干,最后用400 μL甲醇重悬,有机相滤头过滤后利用高效液相色谱(high performance liquid chromatography, HPLC)及液相色谱-质谱联用(liquid chromatography-tandem mass spectrometry, LC-MS/MS)进行检测。

HPLC所用仪器为Thermo Dionex UltiMate 3000,检测条件如下:C18反相色谱柱(4.6 nm×250 nm, 3 µm),检测波长为240 nm,柱温40 ℃,流速0.6 mL/min,进样量5 µL,流动相为乙腈: 水(80:20, 体积分数),进样时间8 min。

LC-MS/MS所用仪器为以上HPLC仪配备电喷雾电离探针,该探针连接Thermo LTQ Orbitrap XL混合质谱仪。测定条件与HPLC相同,样品数据分析在正离子模式下进行。

1.2.5 商品化PUR海绵降解特性分析

称取大小约为2 cm×1 cm×0.5 cm (约0.2 g)的海绵,置于含5% (体积分数) LB的无机盐培养基中,121 ℃灭菌30 min。将菌悬液接入50 mL商业化PUR降解培养基中至OD600=1,30 ℃、180 r/min摇床振荡培养,每组处理设置3组重复。15 d和30 d后(中途补加培养基至恒重)取出海绵,超纯水清洗3遍后用0.4% (体积分数) NaClO浸泡过夜,取出继续用超纯水清洗3遍,最后超声10 min除去菌体置于烘箱中烘干至恒重取出称重并计算失重率。

通过扫描电镜观察菌株YX8-1处理后PUR海绵表面形态变化。

2 结果与分析 2.1 水性聚氨酯DLN降解菌株的分离

为分离聚氨酯降解菌株,以水性聚氨酯Impranil DLN作为底物进行初步筛选。将样品富集、传代后稀释涂布至Impranil DLN平板,获得Impranil DLN降解菌株YX8-1。该菌株在Impranil DLN平板上菌落呈白色,不透明,形态呈圆形,边缘整齐,表面光滑湿润、凹陷;菌株YX8-1在30 ℃、pH 7.0条件下48 h内可在Impranil DLN平板上产生水解圈(图 1A)。革兰氏染色结果显示,菌株YX8-1营养体细胞为杆状、革兰氏阳性;此外,该菌株可以形成不着色的芽孢(图 1B)。

图 1 菌株YX8-1菌落和显微形态 Fig. 1 Colony and micromorphology of strain YX8-1. A: Colony morphology of strain YX8-1 on Impranil DLN plate. B: Gram staining of strain YX8-1 (1 000× magnification).
2.2 菌株YX8-1的系统发育分析

基于菌株的16S rDNA相似性比对分析发现,菌株YX8-1与高地芽孢杆菌(Bacillus altitudinis)、沙福芽孢杆(B. safensis)、短小芽孢杆菌(B. pumilus)等芽孢杆菌同源性大于99%,与模式菌株B. altitudinis 41KF2b相似性最高(99.24%)。但由于高地芽孢杆菌、沙福芽孢杆菌和短小芽孢杆菌亲缘关系较近,很难通过16S rDNA序列将其区分开(图 2)。

图 2 基于16S rDNA序列构建的系统发育树 Fig. 2 Phylogenetic tree based on 16S rDNA sequences.

据报道[25]gyrA序列具有高变异性,使其与16S rDNA相比有更好的系统发育分辨率。将菌株YX8-1的gyrA序列提交至数据库比对,发现菌株YX8-1与模式菌株B. altitudinis 41KF2b的gyrA相似性为98.12%,且与模式菌株B. altitudinis 41KF2b在系统发育树上处于同一个分支(图 3)。此外,基因组数据分析发现,菌株YX8-1基因组序列与B. altitudinis 41KF2b基因组序列的平均核酸同一性为98.5%,高于95%−96%的物种界限[23];基因组间的DNA-DNA杂交值为94.29%,显著高于物种划分阈值(70%)[24];菌株YX8-1基因组G+C含量为41.28%,与模式菌株B. altitudinis 41KF2b基因组G+C含量(41.26%)相当。结合菌株YX8-1的菌落形态,最终确定菌株YX8-1为B. altitudinis中的一员,命名为B. altitudinis YX8-1。

图 3 基于gyrA序列构建的系统发育树 Fig. 3 Phylogenetic tree based on gyrA sequences.
2.3 菌株YX8-1对PBA-PU的降解特性

PUR塑料通常由异氰酸酯和多元醇聚合而成,其中普遍使用的异氰酸酯和多元醇分别为4, 4′-二苯基甲烷二异氰酸酯(4, 4′-diphenylmethane diisocyanate, MDI)和聚己二酸丁二醇酯[poly(1, 4-butylene adipate), PBA]。为进一步确认菌株YX8-1的聚氨酯降解效果,以MDI与PBA聚合形成的PBA-PU作为底物(图 4A)。研究发现,菌株YX8-1在30 ℃、pH 7.0条件下24 h内可在PBA-PU平板上产生水解圈(图 4B)。此外,以PBA-PU为底物培养15 d后,通过HPLC检测到新的产物峰(保留时间5.133 min),且该产物峰的保留时间与标准样品4, 4′-亚甲基二苯胺(4, 4′-methylenedianiline, MDA)的保留时间(5.130 min)基本一致(图 4C)。

图 4 菌株YX8-1的PBA-PU降解特性 Fig. 4 PBA-PU degradation properties of strain YX8-1. A:PBA-PU合成线路图. B:PBA-PU平板上菌株YX8-1的菌落形态. C:菌株YX8-1降解PBA-PU的HPLC图谱. D:菌株YX8-1降解PBA-PU的LC-MS/MS图谱 A: Synthesis and degradation route of PBA-PU. B: Colony morphology of strain YX8-1 on PBA-PU plate. Liquid chromatogram (C) and LC-MS/MS chromatogram (D) of PBA-PU treated by strain YX8-1.

LC-MS/MS结果(图 4D)显示某产物在阳离子条件下分子量为199.122 30,与MDA的理论分子量(M+H+=199.122 98)之间的误差值为−3.41。由于该误差值在5 ppm范围内,因此可认为是同样大小[26]。此外碎片离子分子量为106.067 30,与MDA质子化衍生物相一致,结合以上结果,将该产物鉴定为MDA。

2.4 菌株YX8-1对商品化PUR的降解特性分析

商业化PUR种类多样,成分复杂,为研究菌株YX8-1对商品化PUR的降解效果,将YX8-1与聚酯型PUR海绵进行孵育;孵育15 d后PUR海绵失重率为21.1%,30 d后PUR海绵失重率达到32.42% (图 5A)。经YX8-1处理后的PUR海绵与对照组相比形态发生明显变化,体积缩小表面塌陷且失去弹性(图 5B)。SEM结果显示(图 5C),对照组PUR海绵呈网状、连接紧密,边缘及表面较光滑;而经菌株YX8-1孵育后部分网状结构断裂,结构变得稀疏,表面有多处明显侵蚀痕迹,放大5 000倍时发现海绵表面变得十分粗糙,且布满较大侵蚀坑。这些结果表明菌株YX8-1具有降解商品化PUR海绵的能力。

图 5 YX8-1对商品化PUR的降解特性 Fig. 5 Degradation properties of commercial PUR by strain YX8-1. A:商业化PUR海绵的失重率. B:与菌株YX8-1孵育30 d前(左)后(右) PUR海绵实物图. C:菌株YX8-1孵育前(左)后(右) PUR海绵电镜图 A: Weight loss of commercial PUR after incubation with strain YX8-1. B: Morphology of commercial PUR before (left) and after (right) incubation with strain YX8-1 for 30 d. C: Electron micrograph of commercial PUR before (left) and after (right) incubation with strain YX8-1.
3 讨论

近年来利用生物法处理PUR废弃物成为了研究热点,但目前分离的大部分PUR高效降解菌为真菌,为丰富PUR塑料生物降解资源库,本研究以水性聚氨酯Impranil DLN为底物成功分离出一株PUR降解菌株YX8-1,该菌株在30 ℃时48 h内可在Impranil DLN平板上产生水解圈。根据生理生化特性、系统发育树分析及基因组数据分析,确定菌株YX8-1为芽孢杆菌属的高地芽孢杆菌(B. altitudinis)。目前已发现芽孢杆菌属中枯草芽孢杆菌、沙福芽孢杆菌、短小芽孢杆菌及短芽孢杆菌等具有PUR降解能力[27-34],但还未有过高地芽孢杆菌降解PUR的报道。Nair等[30]从被聚氨酯污染过的水中分离出B. pumilus NMSN-1d,该菌能以Impranil DLN为唯一碳源生长,并在Impranil DLN平板上产生水解圈。Nakkabi等[33]分离到一株B. safensis,该菌可在10 d内使Impranil DLN完全降解。何洁等[34]分离出短芽孢杆菌(Brevibacillus sp.) P10,该菌在添加5% (体积分数) LB作为碳源时,6 d内降解50 mg聚酯型PUR。然而,目前分离的芽孢杆菌属的菌株中大部分仅以能否降解Impranil DLN为指标来评估其PUR降解能力,未探究其对商品化PUR的降解性能。

为进一步评估菌株YX8-1的PUR降解能力,本研究以合成的成分明确的PBA-PU为底物,并通过HPLC及LC-MS/MS检测到了产物MDA的产生。合成PUR塑料结构上与商业化PUR塑料相似且成分明确,已被广泛应用于评估菌株PUR降解能力。Shah等[22]分离的菌株P. aeruginosa MZA-85可降解PBA和MDI为底物合成的聚酯型PUR,通过气相色谱-质谱检测到产物己二酸和1, 4-丁二醇的产生。Magnin等[35]以MDI和4种不同的多元醇聚合形成TPU用于检测不同酶的PUR降解能力,通过LC-MS/MS检测到6-羟基己酸、MDA及MDA衍生物的产生。此外,将菌株YX8-1与商品化聚酯型PUR进行孵育,30 d内PUR海绵失重率达到32.42%。其降解能力与芽孢杆菌属的其他PUR降解菌不相上下。Shah等[36]分离出菌株B. subtilis MZA-75,以聚酯型PUR为唯一碳源时30 d内失重率为20%。彭瑞婷等[37]从载人航天器冷凝水中分离出Bacillus sp. S10-2,该菌株以线性聚酯型PUR作为唯一碳源时30 d内失重率为19%。但由于培养基成分及PUR种类的不同可比性较差。

因商品化PUR成分的复杂性,以商品化PUR为底物时利用HPLC未能检测到其降解产物。且商品化PUR与菌株YX8-1孵育80 d时失重率为36.8%,与孵育30 d的失重率相比仅增加4.38%。这可能是因为随着孵育时间的延长培养基中营养成分的减少导致菌体转变为休眠状态的芽孢从而使得降解效果减弱,也可能是因为商品化PUR中包含的硅油、催化剂等不明物质随着PUR降解被释放到培养基中,从而对PUR降解酶产生了一定抑制作用。因此持续补充碳源,探究功能酶的最适反应条件或许有助于提高其商品化PUR降解能力。后续实验将进一步探究菌株YX8-1在不同培养条件下的商品化PUR降解特性,并挖掘菌株YX8-1中参与PUR降解的功能酶,解析其降解PUR塑料的分子机制。

参考文献
[1]
LIU JW, HE J, XUE R, XU B, QIAN XJ, XIN FX, BLANK LM, ZHOU J, WEI R, DONG WL, JIANG M. Biodegradation and up-cycling of polyurethanes: progress, challenges, and prospects. Biotechnology Advances, 2021, 48: 107730. DOI:10.1016/j.biotechadv.2021.107730
[2]
RUSSELL JR, HUANG J, ANAND P, KUCERA K, SANDOVAL AG, DANTZLER KW, HICKMAN D, JEE J, KIMOVECV FM, KOPPSTEIN D, et al. Biodegradation of polyester polyurethane by endophytic fungi. Appl Environ Microbiol, 2011, 77(17): 6076-6084. DOI:10.1128/AEM.00521-11
[3]
江志通, 陈雪, 闫新, 李周坤, 曹慧, 崔中利. 聚氨酯塑料生物降解的研究现状. 生物加工过程, 2022, 20(4): 439-449.
JIANG ZT, CHEN X, YAN X, LI ZK, CAO H, CUI ZL. Research status of polyurethane biodegradation. Chinese Journal of Bioprocess Engineering, 2022, 20(4): 439-449 (in Chinese). DOI:10.3969/j.issn.1672-3678.2022.04.009
[4]
GROUP T. Plastics-the Facts 2019. RFP: Rubber Fibres Plastics International, 2020(3): 12.
[5]
彭瑞婷, 夏孟丽, 茹家康, 霍毅欣, 杨宇. 聚氨酯塑料的微生物降解. 生物工程学报, 2018, 34(9): 1398-1409.
PENG RT, XIA ML, RU JK, HUO YX, YANG Y. Microbial degradation of polyurethane plastics. Chinese Journal of Biotechnology, 2018, 34(9): 1398-1409 (in Chinese). DOI:10.13345/j.cjb.170532
[6]
JIANG Y, YANG F, HASSAN KAZMI SSU, ZHAO YN, CHEN M, WANG J. A review of microplastic pollution in seawater, sediments and organisms of the Chinese coastal and marginal seas. Chemosphere, 2022, 286: 131677. DOI:10.1016/j.chemosphere.2021.131677
[7]
李昕玥, 刘卓苗, 薛润泽, 代燕辉, 岳同涛, 赵建. 典型塑料的生物降解及其降解机理. 科学通报, 2021, 66(20): 2573-2589.
LI XY, LIU ZM, XUE RZ, DAI YH, YUE TT, ZHAO J. Biodegradation of typical plastics and its mechanisms. Chinese Science Bulletin, 2021, 66(20): 2573-2589 (in Chinese).
[8]
WIERCKX N, POMPOSIELLO P, de LORENZO V, O'CONNOR K, BLANK LM. Plastic waste as a novel substrate for industrial biotechnology. Microbial Biotechnology, 2015, 8(6): 900-903. DOI:10.1111/1751-7915.12312
[9]
KHAN S, NADIR S, SHAH ZU, ALI SHAH A, KARUNARATHNA SC, XU JC, KHAN A, MUNIR S, HASAN F. Biodegradation of polyester polyurethane by Aspergillus tubingensis. Environmental Pollution, 2017, 225: 469-480. DOI:10.1016/j.envpol.2017.03.012
[10]
ÁLVAREZ-BARRAGÁN J, DOMÍNGUEZ- MALFAVÓN L, VARGAS-SUÁREZ M, GONZÁLEZ-HERNÁNDEZ R, AGUILAR-OSORIO G, LOZA-TAVERA H. Biodegradative activities of selected environmental fungi on a polyester polyurethane varnish and polyether polyurethane foams. Applied and Environmental Microbiology, 2016, 82(17): 5225-5235. DOI:10.1128/AEM.01344-16
[11]
WU YH, LI JY, WANG CC, CHEN CY. Effect of oxalate unit on polycarbonate-based polyurethane and biodegradation by fungi. Materials Chemistry and Physics, 2020, 252: 123463. DOI:10.1016/j.matchemphys.2020.123463
[12]
RODRIGO OA, del CASTILLO DAYNET S, JAIME NM, NORA F, JOSé ÃB, ALEJANDRO A, JHONNY SV, MILTON BA. Analysis of the degradation of polyethylene, polystyrene and polyurethane mediated by three filamentous fungi isolated from the Antarctica. African Journal of Biotechnology, 2021, 20(2): 66-76. DOI:10.5897/AJB2020.17200
[13]
SCHAEFER DA, KHAN S, NADIR S, DONG Y, XU J. Biodegradation of polyester polyurethane by Aspergillus flavus G10[J]. BioRxiv, 2020. DOI: 10.1101/2020.06.25.170654.
[14]
HUNG CS, BARLOW DE, VARALJAY VA, DRAKE CA, CROUCH AL, RUSSELL JN Jr, NADEAU LJ, CROOKES-GOODSON WJ, BIFFINGER JC. The biodegradation of polyester and polyester polyurethane coatings using Papiliotrema laurentii. International Biodeterioration & Biodegradation, 2019, 139: 34-43.
[15]
SHAH Z, GULZAR M, HASAN F, SHAH AA. Degradation of polyester polyurethane by an indigenously developed consortium of Pseudomonas and Bacillus species isolated from soil. Polymer Degradation and Stability, 2016, 134: 349-356. DOI:10.1016/j.polymdegradstab.2016.11.003
[16]
ROY R, MUKHERJEE G, GUPTA A, TRIBEDI P, SIL AK. Isolation of a soil bacterium for remediation of polyurethane and low-density polyethylene: a promising tool towards sustainable cleanup of the environment. 3 Biotech, 2021, 11(1): 1-14. DOI:10.1007/s13205-020-02554-1
[17]
AKUTSU Y, NAKAJIMA-KAMBE T, NOMURA N, NAKAHARA T. Purification and properties of a polyester polyurethane-degrading enzyme from Comamonas acidovorans TB-35. Applied and Environmental Microbiology, 1998, 64(1): 62-67. DOI:10.1128/AEM.64.1.62-67.1998
[18]
STERN RV, HOWARD GT. The polyester polyurethanase gene (pueA) from Pseudomonas chlororaphis encodes a lipase. FEMS Microbiology Letters, 2000, 185(2): 163-168. DOI:10.1111/j.1574-6968.2000.tb09056.x
[19]
HOWARD GT, CROTHER B, VICKNAIR J. Cloning, nucleotide sequencing and characterization of a polyurethanase gene (pueB) from Pseudomonas chlororaphis. International Biodeterioration & Biodegradation, 2001, 47(3): 141-149.
[20]
SCHMIDT J, WEI R, OESER T, DEDAVID E SILVA L, BREITE D, SCHULZE A, ZIMMERMANN W. Degradation of polyester polyurethane by bacterial polyester hydrolases. Polymers, 2017, 9(12): 65. DOI:10.3390/polym9020065
[21]
MAGNIN A, ENTZMANN L, BAZIN A, POLLET E, AVÉROUS L. Green recycling process for polyurethane foams by a chem-biotech approach. ChemSusChem, 2021, 14(19): 4234-4241. DOI:10.1002/cssc.202100243
[22]
SHAH Z, HASAN F, KRUMHOLZ L, AKTAS DF, ALI SHAH A. Degradation of polyester polyurethane by newly isolated Pseudomonas aeruginosa strain MZA-85 and analysis of degradation products by GC-MS. International Biodeterioration & Biodegradation, 2013, 77: 114-122.
[23]
YOON SH, HA SM, LIM J, KWON S, CHUN J. A large-scale evaluation of algorithms to calculate average nucleotide identity. Antonie Van Leeuwenhoek, 2017, 110(10): 1281-1286. DOI:10.1007/s10482-017-0844-4
[24]
MEIER-KOLTHOFF JP, AUCH AF, KLENK HP, GÖKER M. Genome sequence-based species delimitation with confidence intervals and improved distance functions. BMC Bioinformatics, 2013, 14(1): 1-14. DOI:10.1186/1471-2105-14-1
[25]
LIU Y, ŠTEFANIČ P, MIAO YZ, XUE YS, XUN WB, ZHANG N, SHEN QR, ZHANG RF, XU ZH, MANDIC-MULEC I. Housekeeping gene gyrA, a potential molecular marker for Bacillus ecology study. AMB Express, 2022, 12(1): 1-12. DOI:10.1186/s13568-021-01341-2
[26]
BLAKE SL, WALKER SH, MUDDIMAN DC, HINKS D, BECK KR. Spectral accuracy and sulfur counting capabilities of the LTQ-FT-ICR and the LTQ-orbitrap XL for small molecule analysis. Journal of the American Society for Mass Spectrometry, 2011, 22(12): 2269-2275. DOI:10.1007/s13361-011-0244-3
[27]
ALDILA F, SUSILOWATI A, SETYANINGSIH R. Polyurethane degrading bacteria isolated from decayed teak wood (Tectona grandis Linn. f.). Journal Biodjati, 2019, 4(2): 225-235. DOI:10.15575/biodjati.v4i2.4525
[28]
BLAKE RC, NORTON WN, HOWARD GT. Adherence and growth of a Bacillus species on an insoluble polyester polyurethane. International Biodeterioration & Biodegradation, 1998, 42(1): 63-73.
[29]
ROWE L, HOWARD GT. Growth of Bacillus subtilis on polyurethane and the purification and characterization of a polyurethanase-lipase enzyme. International Biodeterioration & Biodegradation, 2002, 50(1): 33-40.
[30]
NAIR S, KUMAR P. Molecular characterization of a lipase-producing Bacillus pumilus strain (NMSN-1d) utilizing colloidal water-dispersible polyurethane. World Journal of Microbiology and Biotechnology, 2007, 23(10): 1441-1449. DOI:10.1007/s11274-007-9388-5
[31]
SHAH AA, HASAN F, AKHTER JI, HAMEED A, AHMED S. Degradation of polyurethane by novel bacterial consortium isolated from soil. Annals of Microbiology, 2008, 58(3): 381-386. DOI:10.1007/BF03175532
[32]
NAKKABI A, SADIKI M, FAHIM M, ITTOBANE N, IBNSOUDA KS, BARKAI H, ELABED S. Biodegradation of poly(ester urethane)s by Bacillus subtilis. International Journal Environmental Research, 2015, 9(1): 315-318.
[33]
NAKKABI A, SADIKI M, IBNSSOUNDA KS, FAHIM M. Biological degradation of polyurethane by a newly isolated wood bacterium. International Journal Recent Advanced Multidis Research, 2015, 2: 222-225.
[34]
何洁, 徐安明, 刘嘉唯, 周杰, 崔中利, 董维亮, 姜岷. 一株聚氨酯降解菌的分离及其降解特性解析. 生物工程学报, 2021, 37(10): 3675-3684.
HE J, XU AM, LIU JW, ZHOU J, CUI ZL, DONG WL, JIANG M. Isolation and characterization of a polyurethane-degrading bacterium. Chinese Journal of Biotechnology, 2021, 37(10): 3675-3684 (in Chinese). DOI:10.13345/j.cjb.210411
[35]
MAGNIN A, POLLET E, PERRIN R, ULLMANN C, PERSILLON C, PHALIP V, AVÉROUS L. Enzymatic recycling of thermoplastic polyurethanes: synergistic effect of an esterase and an amidase and recovery of building blocks. Waste Management, 2019, 85: 141-150. DOI:10.1016/j.wasman.2018.12.024
[36]
SHAH Z, KRUMHOLZ L, AKTAS DF, HASAN F, KHATTAK M, ALI SHAH A. Degradation of polyester polyurethane by a newly isolated soil bacterium, Bacillus subtilis strain MZA-75. Biodegradation, 2013, 24(6): 865-877.
[37]
彭瑞婷, 秦利锋, 杨宇. 载人航天器下行细菌对聚氨酯塑料的分解作用[C]//中国生物工程学会第二届青年科技论坛暨首届青年工作委员会学术年会论文集. 广州, 2017: 103-104.
PENG RT, QIN LF, YANG Y. Decomposition of polyurethane plastics by bacteria descending from manned spacecraft[C]//China Society of Biotechnology Young Scientists Forum Ⅱ. Guangzhou, 2017: 103-104 (in Chinese).