发酵剂及猕猴桃蛋白酶促进马肉发酵香肠蛋白质的降解

为了提高马肉发酵香肠的品质,添加发酵剂和猕猴桃蛋白酶,促进其发酵成熟和肌肉蛋白质的降解。以新鲜马肉和马脂肪为原料,分别设计三个试验组,一组为对照组,另两组为处理组;对照组不采取处理,而两个处理组添加2%的发酵剂和0.05%的猕猴桃蛋白酶,并在相应条件下进行发酵,检测其总氮、非蛋白氮等指标,并通过SDS-PAGE电泳分析其肌肉蛋白质的降解情况。结果表明,灌肠后(0 d)各组总氮含量为3.09 g/100 g左右,而加工结束后(28 d),对照组(CK组)、发酵组(A组)、猕猴桃蛋白酶组(B组)总氮含量分别是3.41 g/100 g、3.85 g/100 g及4.15 g/100 g,说明总氮含量明显上升,并且发酵剂和猕猴桃蛋白酶的影响显著;28 d CK组、A组、B组非蛋白氮含量分别为0.42 g/100 g、0.52 g/100 g、0.65 g/100 g,与0 d非蛋白氮含量0.22 g/100 g相比,CK组上升1.9倍,A组上升2.4倍,B组上升2.9倍;SDS-PAGE电泳结果表明,在整个加工过程中A组与B组肌浆蛋白逐渐发生降解,大分子条带发生了明显的变化,发酵剂对马肉发酵香肠肌浆蛋白和肌原纤维蛋白的降解有显著的促进作用,猕猴桃蛋白酶对肌浆蛋白的影响显著,并比发酵剂大,但对肌原纤维蛋白的作用不大。     

发酵剂对发酵香肠蛋白质降解及多肽抗氧化能力的影响

以植物乳杆菌(Lactobacillus plantarum)CD101和模仿葡萄球菌(Staphylococcus simulans)NJ201作为混合发酵剂制作发酵香肠,以自然发酵为对照。通过测定理化指标、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(sodium dodecylsulfate-polyacrylamidegelelectrophoresis,SDS-PAGE)、多肽含量、游离氨基酸含量等指标研究混合发酵剂对发酵香肠蛋白质降解情况,并以体外1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基清除活性、2,2’-联氮双(3-乙基苯并噻唑啉-6-磺酸)(2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid),ABTS)阳离子自由基清除活性及铁离子还原/抗氧化能力(ferric ion reducing antioxidant power,FRAP)值评价发酵香肠粗肽及小分子多肽(3 kDa)的抗氧化能力。结果表明:混合发酵剂接种组非蛋白氮含量显著高于对照组(P0.05);根据SDS-PAGE结果分析,2组发酵香肠的肌浆蛋白和肌原纤维蛋白均发生降解,接种组的蛋白降解程度高于对照组,尤其是在低分子质量(20 kDa)条带处出现明显累积;接种组粗肽及小分子多肽的DPPH自由基清除活性、ABTS阳离子自由基清除活性及FRAP值均显著高于对照组(P0.05),其中小分子多肽可能是发酵香肠多肽抗氧化能力的主要贡献者;同时发酵剂促进发酵香肠中游离氨基酸的释放。接种该混合发酵剂制作发酵香肠能促进蛋白质降解,得到更多具有抗氧化活性的多肽,从而有助于通过内源性抗氧化肽抑制发酵香肠的氧化,降低生产成本,延长货架期。     

接种混合发酵剂生产的低酸度川味香肠加工过程中的蛋白质降解

接种发酵剂并在控温控湿条件下生产低酸度川味香肠,对蛋白质降解情况进行研究,并与自然条件下生产的传统川味香肠(对照组)相比较,揭示低酸度川味香肠加工过程中蛋白质变化规律。结果显示:干燥结束时,低酸度川味香肠的总氮、非蛋白氮和氨基酸态氮分别为4.40%、0.426%和0.366%,均高于对照组;低酸度川味香肠的游离氨基酸总量为605.06mg/100g,高于对照组的497.47mg/100g(p0.01);肌浆蛋白和肌原纤维蛋白在成熟过程中逐渐降解,大分子条带在干燥过程中发生了明显降解(p0.05),低酸度川味香肠的蛋白质降解更为显著(p0.05)。     

羊肉火腿加工过程中蛋白质的降解

对羊肉火腿加工过程中蛋白质的降解变化进行了研究。结果表明,总氮的含量在加工过程中先降低后升高,非蛋白氮含量和蛋白质水解指数在加工过程中持续增加;通过SDS-PAGE图谱分析,肌浆蛋白和肌原纤维蛋白均发生了降解,其中肌浆蛋白的降解主要发生在腌制期和发酵中期,肌原纤维蛋白则是在腌制期和发酵初期;各种游离氨基酸含量随着加工工艺的进行呈现不同程度增加,其中大部分游离氨基酸含量变化最显著的时期是在发酵初期和中期。     

风干肠加工和贮藏过程中蛋白质的降解规律

以总氮、非蛋白氮、氨基酸态氮、挥发性盐基氮、游离氨基酸以及不同种类蛋白质（肌浆蛋白、肌原纤维蛋白、碱溶蛋白和碱不溶蛋白）为指标,分析研究风干肠加工和贮藏过程中蛋白质的降解规律。结果表明：风干肠加工和贮藏过程中总氮含量没有显著变化;非蛋白氮含量持续升高,从原料肉的469.4 mg/100 g增加到成品肠的786.4 mg/100 g,贮藏240 d后增加至1 256.1 mg/100 g;多肽氮和氨基酸态氮含量在加工和贮藏过程中均持续增加,挥发性盐基氮含量在风干过程中没有显著变化,在贮藏180 d后迅速增加;大部分游离氨基酸含量经过加工和贮藏均有所提高,只有半胱氨酸含量明显降低;在加工过程中,肌浆蛋白和肌原纤维蛋白含量均因降解而减少,基质蛋白含量明显增加;在贮藏过程中,肌浆蛋白含量持续降低,肌原纤维蛋白含量在贮藏前180 d无明显变化,贮藏至240 d明显减少,基质蛋白含量没有显著变化。     

发酵剂对熏马肠脂肪酶活力的影响

主要研究发酵剂对熏马肠发酵成熟过程中脂肪酶活力的影响。结果表明,酸性脂肪酶活力、中性脂肪酶活力、磷脂酶活力在发酵成熟过程中持续下降,并且发酵剂组的三种酶活力在添加发酵剂后显著高于空白组(p0.05)。通过相关性分析表明,在熏马肠发酵成熟过程中,发酵剂组中的三种酶活力与p H、水分含量及盐分含量呈极显著相关性(p0.01)。     

中国对虾冷藏过程中肌肉蛋白质的降解规律

分析肌肉中不同结构的蛋白含量的变化,采用SDS-PAGE考察不同溶解性肌肉蛋白质的降解情况,研究中国对虾在不同冷藏温度下(4,0,-2℃)肌肉蛋白质的降解规律。结果显示:贮藏过程中对虾的肌原纤维蛋白、肌基质蛋白含量随着贮藏时间的延长均显著下降,肌浆蛋白含量变化不大,碱溶性蛋白含量呈增加趋势。MHC(肌动球蛋白重链)、肌动蛋白和原肌球蛋白、肌钙蛋白I、肌钙蛋白C和分子质量分别为30~31 ku,6.5 ku的蛋白质均发生不同程度的降解。总蛋白中分子质量23~25 ku的蛋白,水溶性蛋白中分子质量176~178 ku的蛋白,低盐溶性蛋白中分子质量67~68 ku的蛋白丰度变化与贮藏时间呈良好线性关系,有望作为中国对虾新鲜度的指示蛋白。     

木糖葡萄球菌对风鸭蛋白质降解的影响

以木糖葡萄球菌(Staphylococcus xylosus)为发酵剂加工风鸭,研究其在风鸭加工过程中对肌肉蛋白降解的影响.用SDS-PAGE电泳分析蛋白质的降解规律,同时比较加工初期及末期游离氨基酸的变化,并以不接菌样品为对照.结果表明:与对照相比,木糖葡萄球菌促进了肌浆蛋白与肌原纤维蛋白的降解,显著增加了游离氨基酸的总量;产鲜氨基酸的总量和比例均显著增加;必需氨基酸、甜味氨基酸和苦味氨基酸总量显著增加,比例显著下降.总之,木糖葡萄球菌是一种优良的发酵剂.     

米酒乳杆菌对风鸭肌肉蛋白质降解的影响

以米酒乳杆菌(Lactobacillus sake)为发酵剂加工风鸭,研究其在风鸭加工过程中对肌肉蛋白降解的影响。用SDS-PAGE电泳分析蛋白质的降解规律,同时比较加工初期及末期游离氨基酸的变化,并以不接菌样品为对照。结果表明,与对照样相比,米酒乳杆菌促进了肌浆蛋白与肌原纤维蛋白的降解,显著增加了游离氨基酸的总量;产鲜氨基酸、苦味氨基酸的总量和比例均显著增加;必须氨基酸总量显著增加,比例未发生显著变化;甜味氨基酸的总量未发生显著变化,比例显著下降。总之,米酒乳杆菌是一种优良的发酵剂。     

乳酸菌发酵对草鱼肉理化特性和蛋白质降解变化的影响

研究了乳酸片球菌、乳杆菌-Lo3、乳杆菌-B5407和清酒乳杆菌4种不同乳酸菌在发酵草鱼肉时鱼肉理化特性及蛋白质降解变化的规律和特征。结果表明:添加乳酸菌后,鱼肉pH值迅速由最初的6.25降至2d时的5.06～5.40,对照组则为5.98;接种发酵剂的草鱼肉的TBA和TVB-N值的上升较缓慢,整个发酵过程中显著低于对照组(P<0.05);腌制草鱼肉的蛋白质在发酵期发生了轻微的降解,游离氨基酸略有增加;贮藏期间游离氨基酸量显著增加,肌浆蛋白和肌原纤维蛋白均发生了强烈的水解,接菌组的蛋白降解显著多于对照组,其中,接种乳酸片球菌和乳杆菌-B5407组降解程度最大。     

Characterization of Proteolytic Effect of Lactic Acid Bacteria Starter Cultures on Thai Fermented Sausages

The objective of this study was to investigate the effect of starter culture addition on proteolysis of Thai fermented sausages. Sausages inoculated with six different external starter cultures—Pediococcus pentosaceous, Pediococcus acidilactici, Weissella cibaria, Lactobacillus plantarum, Lactobacillus pentosus, and Lactobacillus sakei—were compared with naturally fermented sausages. The results of microbiological analysis indicated that the dominance of lactic acid bacteria (LAB) could inhibit the growth of pathogens and spoilage. Proteolysis was observed during fermentation by the reduction of myofibrillar and sarcoplasmic proteins and the increase in nonprotein nitrogen (NPN) and total free amino acids. The highest increase in concentration of NPN and free amino acids was obtained from sausages inoculated with LAB. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) showed a similar pattern of proteolysis of sarcoplasmic proteins in all sausages, while that of the inoculated sausages with L. plantarum, L. pentsus, and L. sakei exhibited increased degradation of myofibrillar protein bands at 200 and 45 kDa.     

Proteomic and peptidomic insights on myofibrillar protein hydrolysis in a sausage model during fermentation with autochthonous starter cultures

The hydrolysis of myofibrillar proteins during fermentation of sausage models by an autochthonous starter culture was investigated. In order to provide a whole map of the generated products, proteomic and peptidomic were used and complemented with the amino acid profile. Beaker sausages (BS) were used as models which were inoculated or not with Lactobacillus curvatus CRL705 and Staphylococcus vitulinus GV318 as starter cultures. The hydrolysis of actin, myosin light chain 1/3 (MLC 1/3), myosin regulatory light chain-2 (MRLC-2) and myosin heavy chain (MHC) was evidenced by two-dimensional gel electrophoresis (2-DE). In addition, a total of 33 peptides arisen from troponin T, MRLC-2 and particularly from actin were identified by LC–MS/MS. These results showed that the starter culture significantly enhanced the proteolysis of the proteins named above, even when the endogenous enzymes induced a clear breakdown. L. curvatus CRL705 highly enriched both peptide pattern and amino acid concentrations. When the autochthonous starter culture was inoculated, although proteolysis was remarkably reinforced, a reduction in peptide and amino acid composition was observed. Regarding actin primary structure, three regions of this protein were highly susceptible to degradation by the starter culture. Additionally, the essential role of exopeptidases – from meat and bacteria – in diversity of actin peptides during fermentation was shown. This study improved the knowledge of the proteolysis of myofibrillar proteins and the involved enzymes, as well as, completed the previously reported degradation of sarcoplasmic proteins by the same autochthonous starter culture. The singular peptides and amino acids pattern generated might contribute to the uniqueness of produced fermented sausages while they may be used as quality markers.     

Mixed starter cultures to control biogenic amine production in dry fermented sausages

Several combinations of an amine-negative Lactobacillus sakei strain, along with proteolytic Staphylococcus carnosus or Staphylococcus xylosus strains, were used to study the influence of mixed starter cultures on biogenic amine production during the manufacture of dry fermented sausages. Changes in pH, water content, proteolysis, microbial counts, and biogenic amine contents were simultaneously examined in a spontaneously fermented batch and in three mixed starter-mediated batches. A double-controlled microbial charge initially inoculated as mixed starter culture of L. sakei and Staphylococcus spp. (all amine-negative strains) drastically reduced tyramine, cadaverine, and putrescine accumulation. No production of other aromatic amines such as histamine, phenylethylamine, or tryptamine was observed in any batch. The polyamines, spermine and spermidine, were found in raw materials and their levels decreased slightly in the spontaneously fermented batch. No correlation between proteolysis and biogenic amine production was observed. The use of proper technological conditions favoring starter development and the use of the raw materials with good hygienic quality make it possible to produce fermented sausages nearly free of biogenic amines.     

Characterization of proteolysis during the ripening of semi-dry fermented sausages

The respective contribution of indigenous enzymes and enzymes from starter bacteria to proteolysis in fermented sausages were determined by comparing the proteolytic changes occurring in sausages resulting from the presence of a proteolytic strain of Staphylococcus carnosus, i.e. S. carnosus MC 1 to the proteolytic changes occurring in control sausages containing glucono-δ-lactone (GDL) and an antibiotic mixture. Proteolysis was quantified by assaying for non-protein nitrogen (NPN) and free amino acids. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and reversed phase high performance liquid chromatography (RP-HPLC) were used to qualitatively assess the proteolytic changes in the sarcoplasmic and myofibrillar proteins as ripening progressed. The concentration of NPN and free amino acids increased in both sausages initially, but subsequently decreased towards the end of ripening in sausages inoculated with the starter culture. SDS-PAGE showed a similar pattern of proteolysis of sarcoplasmic proteins in both sausages, while of the two sausage types; the S. carnosus MC 1 inoculated sausages exhibited the most intense degradation of myofibrillar proteins, especially myosin and actin. RP-HPLC profiles of 2% trichloroacetic acid (TCA)-soluble peptides for the two sausage types were similar, with the production of numerous hydrophilic peptides. N-Terminal amino acid sequence analysis and sequence homology with proteins of known primary structure showed that six of the TCA-soluble peptides were released from the sarcoplasmic (myoglobin and creatine kinase) and myofibrillar (troponin-I, troponin-T and myosin light chain-2) proteins. In addition, the initial degradation of sarcoplasmic proteins was due to the activity of indigenous proteinases, while both indigenous and bacterial enzymes contributed to the initial degradation of myofibrillar proteins. Furthermore, indigenous enzymes were responsible for the release of TCA-soluble peptides, which, were further hydrolysed by bacterial enzymes.     

Effect of processing methods and starter culture (Staphylococcus xylosus and Pediococcus pentosaceus) on proteolytic changes in Turkish sausages (sucuk) during ripening and storage

Proteolytic changes in Turkish sausages (sucuk) produced by two methods (heat processing and traditional) were determined during processing and storage for 90 days. The sausages were produced with or without starter culture in both methods. A mixture of Staphylococcus xylosus and Pediococcus pentosaceus was used as starter culture for their acidic and proteolytic characteristics.The major changes in proteolytic characteristics of sucuk took place during the fermentation stage, with an increase in non-protein nitrogen (NPN) and a decrease in protein solubility. Proteolytic activity was observed in both starter-inoculated and non-inoculated (control) sausages during processing. Moreover, a slight increase in proteolytic activity was detected during storage in both starter-inoculated and control traditional sausages, and also in heat-processed sausages due to some heat-resistant proteolytic enzymes. Protein solubility was significantly affected by processing time and heat treatment. Sarcoplasmic and myofibrillar proteins were also affected by starter addition, fermentation, drying and heat processing. During fermentation, starter-inoculated and control sausages showed intense proteolysis in both the traditional and heat processing methods. After heating, intensive degradation of both sarcoplasmic and myofibrillar proteins due to denaturation was observed in heat-processed samples.     

Effect of starter culture on proteolytic changes during processing of fermented beef sausages

Fermented beef sausages inoculated with four different starter cultures (Pediococcus acidilactici,Lactobacillus curvatus, Lactobacillus sake, or Streptomyces griseus) were evaluated for proteolysis during process stages (prefermentation, fermentation, drying and heating). Increases (p ? 0.05) in the nonprotein nitrogen (NPN) fraction were found at sequential stages of processing, while starter cultures had no major effects on NPN content. Concentrations of most free amino acids increased (p ? 0.05) during fermentation and drying, and culture effects were found for differences among concentrations of some individual free amino acids. From SDS–PAGE analysis of sarcoplasmic and myofibrillar protein fractions after fermentation and drying, myosin heavy and light chains, actin and troponin were degraded during processing. However, starter culture effects were absent from SDS–PAGE protein patterns.     

Protein Degradation by Lactobacillus plantarum and Lactobacillus casei in a Sausage Model System

ABSTRACT: :
The proteolytic activity of a starter culture involving Lactobacillus plantarum and Lactobacillus casei towards meat sarcoplasmic and myofibrillar proteins during the fermentation of a sausage-like system was studied. After 96 h of incubation the proteolytic system of L. plantarum CRL681 caused a degradation of both sarcoplasmic and myofibrillar proteins, whereas L. casei CRL705 showed a different affinity to meat proteins. The inoculation of both strains showed a higher activity toward sarcoplasmic fraction. These results correlated with a high rate of sarcoplasmic protein degradation observed in SDS-PAGE analysis. The generation of free amino acids as well as the pH drop at the end of the incubation period was maximal in presence of the mixed starter culture, thereby demonstrating the suitability of these strains to be used in the fermentation of meat products.     

Use of Fourier transform infrared spectroscopy to evaluate the proteolytic activity of Yarrowia lipolytica and its contribution to cheese ripening

The interaction between tyrosine-decarboxylase and proteolytic activities of a Lactobacillus curvatus and Staphylococcus xylosus, respectively, on biogenic amine production during the ripening and the storage of dry fermented sausages was investigated. Water content, pH, proteolysis parameters, microbial counts, and biogenic amine contents were monitored in spontaneously and starter fermented sausages. The use of proteolytic staphylococci as starter resulted in a higher content of non-protein nitrogen and total free amino acids. Tyramine was the main amine produced in all batches. However, tyrosine-decarboxylase activity of the L. curvatus starter strain was weak and yielded lower amounts of tyramine than those produced by the wild mioroflora in the control batch. Association between tyramine production and proteolysis could only be established in a defectively dried batch. Putrescine and cadaverine accumulation was efficiently reduced in the starter-mediated fermentation, in agreement with the lower development of enterobacteria. Phenylethylamine and tryptamine were only detected in the spontaneously fermented sausages, while histamine, spermine and spermidine did not vary during the ripening. Biogenic amine levels and related parameters showed significant changes during the storage of dry sausages depending on the temperature and the batch. As a general rule, changes in the pH, proteolysis, microbial counts, and biogenic amine contents were stronger at 19 degrees C than at 4 degrees C. The results suggest that refrigeration would be advisable for preventing further accumulation of biogenic amines during the storage of dry fermented sausages.     

Reduction of biogenic amine formation using a negative amino acid-decarboxylase starter culture for fermentation of Fuet sausages

The ability of Lactobacillus sakei CTC494, a negative amino acid-decarboxylase starter culture, to reduce biogenic amine accumulation during sausage fermentation and storage at 4 and 19 degrees C was studied. The effect on the amine formation of the tyramine producer Lactobacillus curvatus CTC371, as a positive strain, was also examined in comparison to a spontaneous fermentation process without starter culture (control batch). The polyamines spermine, spermidine, and diaminopropane were not influenced by the ripening, and their levels slightly decreased in all the batches throughout the storage. Tyramine, cadaverine, and putrescine were the main amines formed during the ripening. The addition of starter culture resulted in a decrease on the biogenic amine formation, depending on the strain inoculated. A great reduction in tyramine content was achieved when L. sakei CTC494 was inoculated, whereas L. curvatus CTC371 only attenuated tyramine accumulation compared with the control batch. Both starters were able to significantly limit the production of putrescine and cadaverine, and they inhibited tryptamine and phenylethylamine formation by the wild microbial flora. Tyramine levels of the control sausages rose during the storage at both temperatures, whereas those of cadaverine only increased at 19 degrees C. On the contrary, sausages manufactured through the starter controlled fermentation did not show changes of amine contents during the storage. The addition of a proper selected starter culture is advisable to produce safer sausages with low contents of biogenic amines.