冯娟,罗霜,苏雅健,房耀维,杨光,侯晓月,陈梦,刘姝

• 1:江苏海洋大学海洋食品与生物工程学院
• 2:连云港市食品药品检验检测中心

摘要(Abstract)：

【目的】筛选褐藻胶降解菌，评价其对马尾藻（Sargassum denticarpum）的降解效果，为提高马尾藻的利用率提供理论支持。【方法】运用透明圈法和酶活力测定法获得产褐藻胶裂解酶水平较高的菌株，通过16S rDNA序列分析进行菌株鉴定。利用单因素和响应面优化菌株产酶条件，通过扫描电镜观察马尾藻的降解情况，测定藻渣质量和还原糖质量浓度变化。【结果】获得1株符合条件的菌株MAL001，鉴定为阿氏芽孢杆菌Bacillus aryabhattai。优化后产酶条件为NaCl质量分数1%,pH 7.0，温度23℃，转速180 r/min，接种体积分数2%，培养时间24 h，此条件下褐藻胶裂解酶活力达65.8 U/mL，为优化前（44.7 U/mL）的1.5倍。MAL001的褐藻胶裂解酶最适温度为35℃，最适pH为7.0，在最适温度下保持12 h其酶活力仍能在最初酶活力的70%以上；在pH 6.6～8.6时，其酶活力仍保留初始酶活力的60%以上，金属离子Al~(3+)、Fe~(3+)及Co~(2+)抑制褐藻胶裂解酶活力，Mg~(2+)则具有促进作用；该酶降解海藻酸钠的终产物为二糖、三糖和四糖。扫描电镜结果表明，菌株MAL001通过破坏细胞结构裂解马尾藻，藻渣失重率为42.13%，还原糖增量为0.121 mg/mL。【结论】筛选并鉴定得到褐藻胶降解菌阿氏芽孢杆菌Bacillus aryabhattai MAL001，其对马尾藻有良好的降解效果。

关键词(KeyWords)： 阿氏芽孢杆菌;褐藻胶裂解酶;发酵优化;酶学性质;马尾藻降解

Abstract:

Keywords:

基金项目(Foundation): 江苏海洋大学2023年研究生科研与实践创新计划项目（KYCX2023-39）

作者(Author): 冯娟,罗霜,苏雅健,房耀维,杨光,侯晓月,陈梦,刘姝

参考文献(References)：

• [1] ABKA-KHAJOUEI R, TOUNSI L, SHAHABI N, et al.Structures, properties and applications of alginates[J]. Marine Drugs, 2022, 20(6):364.
• [2]蓝嫄嫄,韦蕾,杜良伟,等.海藻酸钠—银纳米粒子涂膜保鲜香蕉研究[J].南方农业学报, 2020, 51(7):1584-1590.
• [3] LUNA-PéREZ Y, RíOS-LóPEZ L G, OTERO-TEJADA E L,et al. Sargassum filipendula, a source of bioactive compounds with antioxidant and matrix metalloproteinases inhibition activities in vitro with potential dermocosmetic application[J].Antioxidants, 2023, 12(4):876.
• [4] XING M C, CAO Q, WANG Y, et al. Advances in research on the bioactivity of alginate oligosaccharides[J]. Marine Drugs,2020, 18(3):144.
• [5] LIU J, KENNEDY J F, ZHANG X F, et al. Preparation of alginate oligosaccharide and its effects on decay control and quality maintenance of harvested kiwifruit[J]. Carbohydrate Polymers, 2020, 242:116462.
• [6] LI F L, TANG Y, WEI L X, et al. Alginate oligosaccharide modulates immune response, fat metabolism, and the gut bacterial community in grass carp(Ctenopharyngodon idellus)[J]. Fish&Shellfish Immunology, 2022, 130:103-113.
• [7] HAN Y, ZHANG L, YU X, et al. Retraction Note:Alginate oligosaccharide attenuates α2, 6-sialylation modification to inhibit prostate cancer cell growth via the Hippo/YAP pathway[J]. Cell Death&Disease, 2021, 12(12):1143.
• [8] POWELL L C, PRITCHARD M F, FERGUSON E L, et al.Targeted disruption of the extracellular polymeric network of Pseudomonas aeruginosa biofilms by alginate oligosaccharides[J]. NPJ Biofilms and Microbiomes, 2018, 4:13.
• [9] AARSTAD O A, T?NDERVIK A, SLETTA H, et al. Alginate sequencing:an analysis of block distribution in alginates using specific alginate degrading enzymes[J]. Biomacromolecules,2012, 13(1):106-116.
• [10] ZHENG L X, LIU Y, TANG S J, et al. Preparation methods,biological activities, and potential applications of marine algae oligosaccharides:a review[J]. Food Science and Human Wellness, 2023, 12(2):359-370.
• [11] FILIPPINI M, BALDISSEROTTO A, MENOTTA S, et al.Heavy metals and potential risks in edible seaweed on the market in Italy[J]. Chemosphere, 2021, 263:127983.
• [12] DUY N D Q, THUY M N, LAL M M, et al. Assessing potential to improve sandfish(Holothuria scabra)culture in Vietnam using supplemental seaweed feeding[J].Aquaculture Reports, 2024, 35:101945.
• [13] INOUE A, OJIMA T. Functional identification of alginate lyase from the brown alga Saccharina japonica[J]. Scientific Reports, 2019, 9:4937.
• [14]郑志国.产褐藻胶裂解酶菌株HB172198的分类鉴定及产酶相关基因分析[D].海口:海南大学, 2019.
• [15] MA Y, LI J, ZHANG X Y, et al. Characterization of a new intracellular alginate lyase with metal ions-tolerant and pH-stable properties[J]. Marine Drugs, 2020, 18(8):416.
• [16]孙驰翔.褐藻胶裂解酶产生菌的筛选与异源表达[D].无锡:江南大学, 2021.
• [17]尚丹丹.白色噬琼胶菌（Agarivorans albus）B2Z047降解海带能力及其机制探究[D].济南:山东大学, 2023.
• [18] SUN X K, GONG Y, SHANG D D, et al. Degradation of alginate by a newly isolated marine bacterium Agarivorans sp. B2Z047[J]. Marine Drugs, 2022, 20(4):254.
• [19]陆钰.褐藻胶裂解酶的高效表达及酶学性质分析[D].无锡:江南大学, 2022.
• [20]许超,熊亚茹,卢明倩,等.一株具有褐藻胶降解能力的海洋细菌的筛选鉴定及其多糖利用能力研究[J].生物技术通报, 2017, 33(4):198-204.
• [21] NGUYEN T N T, CHATAWAY T, ARAUJO R, et al.Purification and characterization of a novel alginate lyase from a marine Streptomyces species isolated from seaweed[J]. Marine Drugs, 2021, 19(11):590.
• [22] TAVAFI H, ABDI-ALI A, GHADAM P, et al. Screening of alginate lyase-producing bacteria and optimization of media compositions for extracellular alginate lyase production[J]. Iranian Biomedical Journal, 2017, 21(1):48-56.
• [23]李梦,温少红,王坤阳,等.产褐藻胶裂解酶菌株的筛选与鉴定[J].食品科技, 2023, 48(7):1-9.
• [24]周敏.褐藻胶裂解酶产酶菌株的优选及褐藻寡糖的研究[D].青岛:青岛大学, 2019.
• [25] HUANG G Y, WANG Q Z, LU M Q, et al. AlgM4:a new saltactivated alginate lyase of the PL7 family with endolytic activity[J]. Marine Drugs, 2018, 16(4):120.
• [26]刘耀天.肠道菌中褐藻胶裂解酶的克隆表达及酶学性质研究[D].天津:天津科技大学, 2021.
• [27]吴阳.褐藻胶裂解酶产生菌BY-1的筛选、培养及酶学性质研究[D].烟台:烟台大学, 2021.
• [28]韩峰.海洋弧菌QY103褐藻胶裂解酶的研究[D].青岛:中国海洋大学, 2008.
• [29] LI Q, HU F, ZHU B W, et al. Biochemical characterization and elucidation of action pattern of a novel polysaccharide lyase 6 family alginate lyase from marine bacterium Flammeovirga sp. NJ-04[J]. Marine Drugs, 2019, 17(6):323.
• [30] YAN J J, CHEN P, ZENG Y, et al. The characterization and modification of a novel bifunctional and robust alginate lyase derived from Marinimicrobium sp. H1[J]. Marine Drugs, 2019, 17(10):545.
• [31]吴雯.基于全基因组分析的褐藻胶裂解酶基因挖掘与表达[D].无锡:江南大学, 2022.
• [32]王定全.复杂海藻多糖降解菌株的分离培养、基因组学分析与活性优化研究[D].舟山:浙江海洋大学, 2022.
• [33] XUE Z, SUN X M, CHEN C, et al. A novel alginate lyase:identification, characterization, and potential application in alginate trisaccharide preparation[J]. Marine Drugs,2022, 20(3):159.
• [34] ZHANG Y H, SHAO Y, JIAO C, et al. Characterization and application of an alginate lyase, Aly1281 from marine bacterium Pseudoalteromonas carrageenovora ASY5[J].Marine Drugs, 2020, 18(2):95.
• [35] DONG F, XU F, CHEN X L, et al. Alginate lyase Aly36B is a new bacterial member of the polysaccharide lyase family 36 and catalyzes by a novel mechanism with lysine as both the catalytic base and catalytic acid[J]. Journal of Molecular Biology, 2019, 431(24):4897-4909.
• [36] YIN R, YI Y J, CHEN Z, et al. Characterization of a new biofunctional, exolytic alginate lyase from Tamlana sp.s12 with high catalytic activity and cold-adapted features[J]. Marine Drugs, 2021, 19(4):191.
• [37]宋鑫.马尾藻降解微生物的筛选、鉴定及其降解效果初步评价[D].海口:海南大学, 2016.