靛蓝的绿色生物制造：酶法及发酵生产靛蓝的研究进展及挑战

doi:10.19586/j.2095-2341.2025.0182

生物技术进展 ›› 2026, Vol. 16 ›› Issue (3): 535-548.DOI: 10.19586/j.2095-2341.2025.0182

靛蓝的绿色生物制造：酶法及发酵生产靛蓝的研究进展及挑战

卢志豪(), 张永辉, 郭泽望, 陈俊, 庄小燕()

集美大学海洋食品与生物工程学院，福建厦门 361021

收稿日期:2025-12-16 接受日期:2026-01-21 出版日期:2026-05-25 发布日期:2026-07-14
通讯作者: 庄小燕
作者简介:卢志豪 E-mail: 19954375274@163.com；
基金资助:
国家自然科学基金项目(32301252)

Green Biomanufacturing of Indigo: Enzymatic and Fermentation Routes-advances and Challenges

Zhihao LU(), Yonghui ZHANG, Zewang GUO, Jun CHEN, Xiaoyan ZHUANG()

College of Ocean Food and Biological Engineering，Jimei University，Fujian Xiamen 361021，China

Received:2025-12-16 Accepted:2026-01-21 Online:2026-05-25 Published:2026-07-14
Contact: Xiaoyan ZHUANG

摘要/Abstract

摘要：

靛蓝作为一种历史悠久且应用广泛的蓝色色素，传统生产方法存在植物提取效率低、化学合成污染大等问题，推动其绿色生物制造成为研究热点。目前，酶法主要集中在黄素单加氧酶和细胞色素P450单加氧酶的挖掘与改造，通过定向进化与理性设计提升其催化效率；发酵法则侧重于利用野生菌株或基因工程菌株，结合代谢工程与发酵过程优化以提高产量。系统梳理了两类方法的关键酶、菌种、改造策略及优化手段，并总结了当前工业化应用中存在的产率低、稳定性不足等挑战，旨在为靛蓝的绿色、高效生物制造提供理论参考与技术支撑，推动其在可持续发展领域的应用。

关键词: 靛蓝, 色素, P450单加氧酶, 黄素单加氧酶, 绿色制造

Abstract:

Indigo， a historically significant and widely used blue pigment， faces challenges in its traditional production， such as low extraction efficiency from plants and substantial pollution from chemical synthesis. These limitations have propelled the green biomanufacturing of indigo. Currently， enzymatic strategies mainly focus on the discovery and engineering of flavin-containing monooxygenases and cytochrome P450 monooxygenases， with efforts directed toward improving catalytic efficiency through directed evolution and rational design. Meanwhile， fermentation methods emphasize the use of wild-type or genetically engineered microbial strains， coupled with metabolic engineering and fermentation process optimization to enhance yield. This paper systematically outlined the key enzymes， microbial hosts， engineering strategies， and optimization techniques associated with both routes. It also discussed major challenges hindering industrial application， including low productivity and insufficient operational stability. The review aims to provide theoretical insights and technical support for the green and efficient biomanufacturing of indigo， thereby facilitating its sustainable development and application.

Key words: indigo, pigment, P450 monooxygenase, flavin-containing monooxygenase, green manufacturing

中图分类号:

Q815

卢志豪, 张永辉, 郭泽望, 陈俊, 庄小燕. 靛蓝的绿色生物制造：酶法及发酵生产靛蓝的研究进展及挑战[J]. 生物技术进展, 2026, 16(3): 535-548.

Zhihao LU, Yonghui ZHANG, Zewang GUO, Jun CHEN, Xiaoyan ZHUANG. Green Biomanufacturing of Indigo: Enzymatic and Fermentation Routes-advances and Challenges[J]. Current Biotechnology, 2026, 16(3): 535-548.

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图/表 11

参考文献 72

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FMO来源	表达载体	靛蓝产量/(mg·L^-1)	参考文献
噬甲基菌（Methylophaga sp.）	pBlue scriptSK(+)	160	[42]
嗜硫胺噬甲基菌（Methylophaga aminisulfidivorans）	pBlue 1.7	920	[43]
棒状杆菌（Corynebacterium glutamicum）	pMAL-c2x	685	[45]
酶蓼蓝（Polygonum tinctorium）	pET19b	30	[46]

FMO来源	表达载体	靛蓝产量/(mg·L^-1)	参考文献
噬甲基菌（Methylophaga sp.）	pBlue scriptSK(+)	160	[42]
嗜硫胺噬甲基菌（Methylophaga aminisulfidivorans）	pBlue 1.7	920	[43]
棒状杆菌（Corynebacterium glutamicum）	pMAL-c2x	685	[45]
酶蓼蓝（Polygonum tinctorium）	pET19b	30	[46]

FMO来源	改造位点	改造后活性	参考文献
噬甲基菌（Methylophaga sp. SK1）	F170、A210和T326	K_m=0.035 mmol·L^-1， k_cat=0.61 s^-1	[47]
棒状杆菌（Corynebacterium glutamicum ATCC13032）	M185L/V402A（双点突变，位于底物通道与辅酶结合区）	K_m=0.66 mmol·L^-1， k_cat=0.4 s^-1	[48]
嗜硫胺噬甲基菌（Methylophaga aminosulfidivorans）	K223R/D317S	K_m=0.55 mmol·L^-1， k_cat=0.79 s^-1	[50]

FMO来源	改造位点	改造后活性	参考文献
噬甲基菌（Methylophaga sp. SK1）	F170、A210和T326	K_m=0.035 mmol·L^-1， k_cat=0.61 s^-1	[47]
棒状杆菌（Corynebacterium glutamicum ATCC13032）	M185L/V402A（双点突变，位于底物通道与辅酶结合区）	K_m=0.66 mmol·L^-1， k_cat=0.4 s^-1	[48]
嗜硫胺噬甲基菌（Methylophaga aminosulfidivorans）	K223R/D317S	K_m=0.55 mmol·L^-1， k_cat=0.79 s^-1	[50]

P450来源	定向进化突变位点	最后活性（k_cat/K_m）	参考文献
芽孢杆菌（Bacillus megaterium）	F87V/L188Q/A74G（三突变）	1 365 L·mol^-1·s^-1	[55]
芽孢杆菌（Bacillus megaterium）	E435T	577 L·mmol^-1·min^-1	[56]
芽孢杆菌（Bacillus megaterium）	F87G/T268A	1 388 L·mmol^-1·min^-1	[57]

靛蓝的绿色生物制造：酶法及发酵生产靛蓝的研究进展及挑战

Green Biomanufacturing of Indigo: Enzymatic and Fermentation Routes-advances and Challenges

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P450来源	改造位点	最后活性（k_cat/K_m）	参考文献
芽孢杆菌（Bacillus megaterium P450 BM3）	D168位点进行定点饱和突变	278 L·mmol^-1·min^-1	[60]
芽孢杆菌（Bacillus megaterium P450 BM3）	以三突变体A74G/F87V/L188Q为亲本，再叠加定点突变D168L与E435T	16.37 L·mmol^-1·min^-1	[61]
链霉菌SC0581Streptomyces sp. SC0581	单点突变库（21个）以及组合突变体	43.7 U·mg^-1	[59]

菌种来源	生产方式	产量/（mg·L^-1）	参考文献
大肠杆菌（Escherichia coli HB101）	利用萘作为唯一的碳源	25	[28]
不动杆菌属（Acinetobacter sp. ST-550）	采用二苯甲烷-水两相体系	292	[64]
肠杆菌属（Enterobacter sp.）	利用萘作为唯一碳源生长，萘同时作为诱导物，激活菌体内的萘双加氧酶表达	68.1	[66]

菌种来源	基因改造方式	靛蓝产量	参考文献
嗜硫胺噬甲基菌（Methylophaga aminisulfidivorans）	强化色氨酸摄取（“入口”改造）	1 492 mg·L^-1	[67]
大肠杆菌（Escherichia coli）	敲除竞争支路	188.9 mg·L^-1	[69]
噬甲基菌（Methylophaga）	trpR、tnaB、mtr、ptsG竞争途径基因敲除	3.9 g·L^-1	[13]

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