【作者】 张雯；

【导师】 叶秀云；

【作者基本信息】 福州大学 ， 分析化学， 2014， 博士

【摘要】 大黄鱼(Pseudosciaena crocea)是我国特有的海产鱼类,大黄鱼销售的主要方式为冰鲜整鱼销售,为了解决冰鲜大黄鱼在贮藏和销售过程中变质腐败和延长其货架期,开展了冰鲜大黄鱼腐败机理以及保鲜新技术研究。主要研究成果：研究发现海产品鲜度评价指标中,针对鱼肉的挥发性盐基氮(TVBN)和细菌菌落计数(TVC)指标判定大黄鱼各级鲜度等级的出现期和维持期明显滞后于感官评定结果。建议以大黄鱼的鱼肠TVBN指标和TVC指标来评判大黄鱼鲜度等级与感官评定结果更趋一致。提出利用鱼体菌群变化确定鲜度变化和预测货架期,分析各部位菌相变化,得出贮藏2-3d和5-6d是鲜度变化的关键时间点。用细菌16S rDNA PCR-变性梯度凝胶电泳(DGGE)与主成分分析(PCA)对贮藏期大黄鱼细菌菌相变化分析结果,冰鲜贮藏前期大黄鱼各部位的菌群以种类多样性为特征,嗜冷杆菌和希瓦氏菌是主要特征菌；贮藏后期鱼体优势菌突出,植物乳杆菌是大黄鱼各部位的共有特征菌,其他重要特征菌有希瓦氏菌、假单胞菌和戊糖片球菌。不同部位的特征菌群结构有区别。PCR-DGGE图谱主成分分析结果,大黄鱼肠道是菌群结构最稳定,腐败最快的部位,以假单胞菌为主要特征菌。鱼肠中的假单胞菌在腐败后期向鱼肉、鱼鳃扩散,鱼鳃和鱼皮中的希瓦氏菌向鱼肠扩散,最终导致鱼体全面变质腐败。利用多重培养基分离、细菌16S rDNA PCR- DGGE测定、细菌总脂肪酸鉴定等技术,从大黄鱼鱼鳞、鱼皮、鱼肉、鱼鳃和鱼肠等部位鉴定出希瓦氏菌属、嗜冷杆菌属、假单胞杆菌属、乳杆菌属、片球菌属、金黄杆菌属等20属的细菌。希瓦氏菌属、嗜冷杆菌属、假单胞杆菌属、乳杆菌属、片球菌属为优势菌。通过接种试验证实了腐败希瓦氏菌、静止嗜冷杆菌和荧光假单胞菌对大黄鱼具有致腐能力。在4℃-25℃随温度增高各菌株的繁殖速度加快和致腐能力增强。腐败希瓦氏菌O是低菌浓度致腐菌,荧光假单胞菌是高菌浓度致腐菌；高菌量条件下对三甲胺的生成具有自我抑制作用,腐败希瓦氏菌A尤为明显。利用气相色谱-嗅觉测量法(GC-O)可以对大黄鱼的鲜度进行区分,并判断菌株的腐败特性。对峙培养试验证实枯草芽孢杆菌BS08对大黄鱼致腐菌具有抑菌活性和较广的抑菌谱。经高效液相色谱、质谱和红外吸收光谱测定,抑菌活性物质有效成份为枯草芽孢杆菌表面活性素,其具有良好的热稳定性、酸碱稳定性和溶血活性；优化了发酵工艺和研制了枯草芽孢杆菌BS08大黄鱼保鲜剂。更多还原

【Abstract】 Large yellow croaker (Pseudosciaena crocea) is an important commercial marine fish of China. Large yellow croaker is sold as iced whole fresh fish in markets in order to meet the demand for freshness, rather than frozen fish. The objectives of this dissertation were to investigate the spoilage microbial ecology of large yellow croaker and develop some preservation technologies to extend the shelf-life of the fish.The main research results are listed as follows:As the traditional indicators of freshness of fish, TVB-N (TotalVolatile Base Nitrogen) and TVC (Total Viable Count)in fish fleshwere less sensitivethan sensory evaluation. The research showed TVB-N and TVC in gills and intestines were qualified indicators, whoseconclusionswere consistent with sensory evaluation.It was suggested that the diversity of bacterial communities of fish could be used to discriminate and predict fishfreshness. The bacterial flora changed significantlyduring storage, and it was showed that 2-3d and 5-6d of storage were key time pointsin the change of bacterial flora and freshness.The diversity of bacterial communities of fish based on 16S rDNA PCR-DGGE analyzedwithprincipal component analysis (PCA) showed the abundantbacterial flora, featuredby Psychrotrophic and Shewanelladuring early storage time, and during later storage time,the bacterial flora were simple, featured by Lactobacillusplantarum, PsychrotrophicShewanella and Pediococcus. Fish intestine spoiled fastest. Intestinalbacterial flora was most stable, featured byPseudomonas. Pseudomonas in the fish intestine diffused to fish flesh and gills, and Shewanella in fish skin and gills spread to the intestine, resulted in freshness decay absolutely.20 genuses of bacteria were identified from scale, skin, flesh, gills and intestine of fish based on 16S rDNA PCR-DGGE, purification and identification with multiple medium and fatty-acid identification technology. The results showed that Shewanella, Psychrotrophic, Pseudomonas, Lactobacillus, Pediococcus and Chryseobacteriumwere dominant flora.Shewanella putrefaciens, Pseudomonas fluorescens and Psychrobacter immobilis were confirmed to be spoilage bacteria by inoculation test The strainsgrew faster and spoilage capacity was enhancedas temperature increased from 4℃ to 25 ℃.Shewanella putrefaciensstrain O showed greaterability of spoilageat lower inoculum concentration, and Pseudomonas fluorescensshowed higher ability of spoilage at higher inoculum concentration. Self-inhibition of TMAwere found at high bacterial concentration, particularly withShewanella putrefaciensstrain A. GC-O method was used to distinguish the freshness of fish and determine the type of spoilage strains.Bacillus subtilis BS08 hadbroad spectrum antibacterial activity against spoilage strains isolated from large yellow croaker. The antibacterial substances wereidentifiedasBacillus subtilis surfactinwithHPLC-MS and IR. The surfactin had good thermal stability, pH stability and hemolytic activity.Fermentation process wasoptimizedfor the yield ofsurfactin, and the biological preservative was used to prevent and control spoilage in large yellow croaker.更多还原