Proteolytic profiles and angiotensin-I converting enzyme and alpha-glucosidase inhibitory activities of selected lactic acid bacteria

ABSTRACT:  This study was conducted to examine the growth, proteolytic profiles as well as angiotensin-I converting enzyme (ACE) and α-glucosidase (α-glu) inhibitory potentials of selected strains of lactic acid bacteria (LAB). Two strains each of yogurt bacteria ( Streptococcus thermophilus —1275 and 285, and Lactobacillus delbrueckii ssp. bulgaricus —1092 and 1368), and probiotics ( L. acidophilus —4461 and 33200, and L. casei —2607 and 15286, and 1 strain of Bifidobacterium longum 5022), were cultivated in reconstituted skim milk (RSM) at 37 °C and their proteolytic profiles and ACE as well as α-glu inhibitory activities were determined. Among all the strains of lactic acid bacteria studied, yogurt bacteria grew very well, with the exception of L. delbrueckii ssp. bulgaricus 1368 which showed a slower growth during the initial 3 h of incubation. The growth pattern corresponded well with the decrease in pH for the organisms. All the organisms showed an increase in proteolysis with time. The variations in proteolytic capabilities translated into corresponding variations in ACE inhibitory potential of these organisms. Bifidobacterium longum 5022 showed the highest ACE inhibitory potential followed by L. delbrueckii ssp. bulgaricus 1368, L. casei 15286, S. thermophilus 1275, and L. acidophilus 4461. Organisms with high intracellular enzymatic activities grew well. Also, aminopeptidases of strains of L. acidophilus 4461 and S. thermophilus 1275 that could better utilize proline containing substrates showed enhanced ACE inhibitory potential. Lactic acid bacteria possessed the ability to inhibit α-glu activity, which endowed them an antidiabetic property as well.     

The viability of three probiotic organisms grown with yoghurt starter cultures during storage for 21 days at 4°C

This study investigated the viability of probiotic ( Lactobacillus acidophilus LA5, Lactobacillus rhamnosus LBA and Bifidobacterium animalis subsp . lactis BL-04) in milk fermented with Lactobacillus delbrueckii subsp . bulgaricus LB340 and Streptococcus thermophilus TAO (yoghurt – Y). Each probiotic strain was grown separately in co-culture with Y and in blends of different combinations. Blends affected fermentation time(s), pH and firmness during storage at 4°C. The product made with Y plus B. animalis subsp . lactis and L. rhamnosus had counts of viable cells at the end of shelf life that met the minimum required to achieve probiotic effect. However, L. acidophilus and L. delbrueckii subsp . bulgaricus were inhibited.     

腊鱼中优势乳酸菌的分离、纯化及性质鉴定

为了改进传统腊鱼生产工艺并研究腊鱼生产中优势乳酸菌的种类及性质,利用鉴别性培养基从传统腊鱼制品中分离得到25株疑似菌株,根据生理生化实验确定其性质,获得4株优势菌株分别为干酪乳杆菌、德氏乳杆菌保加利亚种、香肠乳杆菌、乳酸乳杆菌,并考查了4株菌株的24h产酸能力及生长特性。     

内蒙古酸马奶中乳酸菌多样性的研究

采用16S rRNA基因全序列测定和聚类分析技术,对酸马奶中的乳酸菌进行了准确鉴定并构建了乳酸菌的系统发育树.然后对乳酸菌菌群进行了多样性分析,结果显示,酸马奶中的优势乳酸菌分别为:Lactobacillus plantarum(10%),Lactobacillus brevis(8%),Lactobacillus casei(7.8%),Enterococcus faecium(17%),Enterococcus faecalis(14%),Lactococcuslactis(19%),Lactobacillus acidlophilus(5%),Lactobacillus paracasei(2%),Lactobacillus delbrueckii subsp.bulgaricus(4%),Lactobacillus helveticus(4%),Enterococcus durans(4%),Leuconostoc mesenteroides (4%),Leuconostoc garlicum(1%),Streptococcus thermophilus(1%).     

DNA fingerprinting of thermophilic lactic acid bacteria using repetitive sequence-based polymerase chain reaction

DNA fingerprints of lactic acid bacteria were generated by polymerase chain reaction using a primer based on the repetitive elements found in the genome of Streptococcus pneumoniae (BOX-PCR). The method made it possible to identify 37 isolates from raw milk. industrial starters and yogurt. Differentiation at species, subspecies and strain level was possible for Lactobacillus delbrueckii subsp. lactis, Lb. delbrueckii subsp bulgaricus and Str. thermophilus. BOX-PCR was also applied to studying the strain composition of a starter culture and the direct detection of strains in commercial fermented milk.     

Antibiotic resistance of lactic acid bacteria and Bifidobacterium spp. isolated from dairy and pharmaceutical products

The outlines of antibiotic resistance of some probiotic microorganisms were studied. This study was conducted with the double purpose of verifying their ability to survive if they are taken simultaneously with an antibiotic therapy and to increase the selective properties of suitable media for the isolation of samples containing mixed bacterial populations. We isolated from commercial dairy and pharmaceutical products, 34 strains declared as probiotics, belonging to the genera Bifidobacterium and Lactobacillus, and 21 strains of starter culture bacteria. All the microorganisms have been compared by electrophoresis of the soluble proteins for the purpose of identifying them. A Multiplex-PCR with genus- and species-specific primers was used to detect for Bifidobacterium animalis subsp. lactis presence. All bifidobacteria were B. animalis subsp. lactis except one Bifidobacterium longum. Sometimes the identification showed that the used strain was not the one indicated on the label. The lactobacilli were Lactobacillus acidophilus, Lactobacillus casei, and Lactobacillus delbrueckii subsp. bulgaricus. The streptococci were all Streptococcus thermophilus. The minimal inhibitory concentration (MIC) of 24 common antibiotic substances has been valued by the broth microdilution method. All tested strains were susceptible to ampicillin, bacitracin, clindamycin, dicloxacillin, erytromycin, novobiocin, penicillin G, rifampicin (MIC(90) ranging from 0.01 to 4 microg/ml); resistant to aztreonam, cycloserin, kanamycin, nalidixic acid, polymyxin B and spectinomycin (MIC(90) ranging from 64 to >1000 microg/ml). The susceptibility to cephalothin, chloramphenicol, gentamicin, lincomycin, metronidazole, neomycin, paromomycin, streptomycin, tetracycline and vancomycin was variable and depending on the species.     

乳酸菌发酵大豆糖蜜生产乳酸及糖代谢变化

选用德氏乳杆菌保加利亚亚种KLDS1.8501、嗜酸乳杆菌KLDS1.0327、嗜酸乳杆菌ATCC11975、植物乳杆菌植物亚种CICC23168、干酪乳杆菌ATCC393、植物乳杆菌NAU322分别接种于大豆糖蜜,用高效液相色谱法测定乳酸的产生以及碳水化合物的利用情况,分析不同乳酸菌发酵大豆糖蜜生产乳酸能力及糖代谢能力。结果表明,在15 °Brix大豆糖蜜中,37?℃、pH?6.0条件下发酵24?h,植物乳杆菌植物亚种CICC23168的活细胞数达到6.66×109?CFU/mL,乳酸产生量为12.18?g/L,总糖消耗量为22.48?g/L,与其他菌株相比有明显优势。因此,植物乳杆菌植物亚种CICC23168是能利用大豆糖蜜发酵产乳酸的潜力菌株。     

不同乳酸菌对椰子水饮料的发酵特性研究

以椰子水为原料,利用6株乳酸菌:发酵乳杆菌L20、面包乳杆菌32-2-2、短乳杆菌64-1、植物乳杆菌A33、嗜热链球菌IFFI6038和德氏乳杆菌保加利亚亚种CICC6045发酵椰子水制备饮料,分析测定发酵椰子水主要成分、抑菌活性和感官评价的变化。试验结果表明,发酵48 h时,面包乳杆菌32-2-2发酵椰子水饮料总酸含量最高,达10.85 g/L;植物乳杆菌A33发酵椰子水饮料pH值最低为3.1;德氏乳杆菌保加利亚亚种CICC6045发酵椰子水饮料还原糖含量最低为0.03 g/L;植物乳杆菌A33的抑菌谱最广;抑菌活性与pH值和乳酸含量呈显著相关性。植物乳杆菌、嗜热链球菌和德氏乳杆菌保加利亚亚种发酵椰子水感官评分较高。     

Viability of probiotic (Bifidobacterium, Lactobacillus acidophilus and Lactobacillus casei) and nonprobiotic microflora in Argentinian Fresco cheese

We evaluated the suitability of Argentinian Fresco cheese as a food carrier of probiotic cultures. We used cultures of Bifidobacterium bifidum (two strains), Bifidobacterium longum (two strains), Bifidobacterium sp. (one strain), Lactobacillus acidophilus (two strains), and Lactobacillus casei (two strains) in different combinations, as probiotic adjuncts. Probiotic, lactic starter (Lactococcus lactis and Streptococcus thermophilus), and contaminant (coliforms, yeasts, and molds) organisms were counted at 0, 30, and 60 d of refrigerated storage. Furthermore, the acid resistance of probiotic and starter bacteria was determined from hydrochloric solutions (pH 2 and 3) of Fresco cheese. The results showed that nine different combinations of bifidobacteria and L. acidophilus had a satisfactory viability (count decreases in 60 d <1 log order) in the cheese. Both combinations of bifidobacteria and L. casei cultures assayed also showed a satisfactory survival (counts decreased <1 log order for bifidobacteria but no decrease was detected for L. casei). On the other hand, the three combinations of bifidobacteria, L. acidophilus, and L. casei tested adapted well to the Fresco cheese environment. When a cheese homogenate at pH 3 was used to partially simulate the acidic conditions in the stomach, the probiotic cultures had an excellent ability to remain viable up to 3 h. At pH 2, the cell viability was more affected; B. bifidum was the most resistant organism. This study showed that the Argentinian Fresco cheese could be used as an adequate carrier of probiotic bacteria.     

藤茶中二氢杨梅素对乳酸菌生长及活性的影响

该研究通过生长曲线和流式细胞术实验初步探索了藤茶中二氢杨梅素对嗜酸乳杆菌（Lactobacillus acidophilus）、德氏乳杆菌保加利亚亚种（Lactobacillus delbrueckii subsp. bulgaria）、嗜热链球菌（Streptococcus thermophilus）三种乳酸菌生长的影响。结果表明,二氢杨梅素添加量在100～1 000 μg/mL时,对三种乳酸菌均有显著促进其生长并提高其菌液活菌数量的作用（P＜0.05）;当二氢杨梅素添加量＞1 000 μg/mL时,对三种菌的生长和菌液中活菌的数量均表现出不同程度的抑制,其中德氏乳杆菌保加利亚亚种对二氢杨梅素最为敏感。在发酵后的菌液分析中,未明显观察到二氢杨梅素含量的降低,说明二氢杨梅素通过对生长环境进行调节影响乳酸菌生长。     

Production of pyroglutamic acid by thermophilic lactic acid bacteria in hard-cooked mini-cheeses

Pyroglutamic acid is present in high amounts (0.5g/ 100g) in many cheese varieties-and particularly in extensively ripened Italian cheeses such as Grana Padano and Parmigiano Reggiano. An in vivo model system for cooked mini-cheese production and ripening acceleration was set up to demonstrate the ability of thermophilic lactic acid bacteria, used as a starter, to produce pyroglutamic acid (pGlu). In mini-cheeses stored at 38 and 30 degrees C for up to 45 d, all starters tested produced different amounts of pGlu. In descending order of pGlu production, the bacteria analyzed were: Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and Lactobacillus delbrueckii subsp. lactis. Evidence for the presence of glutamine to pGlu cyclase activity in lactic acid bacteria was provided. Cell lysates obtained from cultures of L. helveticus, L. delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis, and S. thermophilus showed the ability to cyclize glutamine to pGlu, resulting in processing yields from 1.4 to 30.3%, depending on the subspecies. Formation of pGlu from free glutamine appeared to be similar to that observed using a glutamine-glutamine dipeptide substrate. Under the experimental conditions applied, pGlu aminopeptidase activity was only detected in L. helveticus. Thus, pGlu formation in long-ripened cooked cheese may depend on the activity of thermophilic lactic acid bacteria.     

Growth of Probiotic and Traditional Yogurt Cultures in Milk Supplemented with Whey Protein Hydrolysate

ABSTRACT: Growth of some probiotic bacteria was significantly improved in milk supplemented with whey protein hydrolysate (WPH). However, WPH had no effect on the growth of Lactobacillus delbrueckii ssp. bulgaricus 18, L. delbrueckii ssp. bulgaricus 10442, and Streptococcus thermophilus 1. When the probiotic bacteria were grown in combination with different yogurt cultures in milk, WPH caused significant increases in growth of Bifidobacterium longum S9, L. acidophilus O16, and L. acidophilus L-1. However, by day 28 of refrigerated storage, the populations of the probiotic cultures that had been grown in samples supplemented with WPH were similar or below those in the control samples.     

Yoghurt fermented by Lactobacillus delbrueckii subsp. bulgaricus H+ -ATPase-defective mutants exhibits enhanced viability of Bifidobacterium breve during storage

Persistent acid production by Lactobacillus delbrueckii subsp. bulgaricus during refrigerated storage is a major cause of reduced viability of probiotic strains such as Bifidobacterium breve in yoghurt. It was established that H+ -ATPase-defective mutants of lactic acid bacteria have reduced growth and metabolism in low pH environments. Therefore, the aim of this study was to evaluate inhibition of post-acidification and maintenance of B. breve viability in yoghurt fermented by L. delbrueckii subsp. bulgaricus mutants with reduced membrane-bound H+ -ATPase activity during refrigerated storage. Spontaneous neomycin mutants of L. delbrueckii subsp. bulgaricus that had a significantly (P < or = 0.05) reduced H+ -ATPase activity were successfully isolated. Yoghurt fermented using L. delbrueckii subsp. bulgaricus SBT0164 No. 55-1 (mutant) starter culture had markedly reduced post-acidification and maintained viability (> or = 10(8) CFU/ml) of both Bifidobacteruim breve JCM 1192(T) and Bifidobacteruim breve JCM 7017 during storage at 10 degrees C for 21 days. These results clearly showed that yoghurt fermented by mutants of L. delbrueckii subsp. bulgaricus with reduced membrane-bound H+ -ATPase activity has reduced post-acidification that prolongs viability of B. breve in yoghurt during refrigerated storage.     

Study of the possible mechanisms involved in the mucosal immune system activation by lactic acid bacteria.

The induction of a mucosal immune response is not easy due to the development of oral tolerance, but under some conditions, bacteria can activate this immune system. Antigens administered orally can interact with M cells of Peyer's patches or bind to the epithelial cells. We have demonstrated that certain lactic acid bacteria are able to induce specific secretory immunity, and others will enhance the gut inflammatory immune response. The aim of this work was to establish the reason for these different behaviors and to define possible mechanisms involved in the interaction of lactic acid bacteria at the intestinal level. We studied IgA+ and IgM+ B cells comparatively in bronchus and intestine and CD4+ T cells and IgA anti-lactic acid bacteria antibodies in the intestinal fluid, induced by oral administration of Lactobacillus casei, Lb. delbrueckii ssp. bulgaricus, Lb. acidophilus, Lb. plantarum, Lb. rhamnosus, Lactococcus lactis, and Streptococcus salivarius ssp. thermophilus. The increase in the IgA+ B cells in the bronchus means that these lactic acid bacteria were able to induce the IgA cycle by interaction with M cells from Peyer's patches or intestinal epithelial cells. The IgM+ cells increased when the stimulus did not induce the switch from IgM+ to IgA+. The increase in the CD4+ cells suggests interaction of Peyer's patches and enhancement of the B- and T-cell migration. The anti-lactic acid bacteria antibody is related to the processing and presentation of the microorganisms to the immune cells. We demonstrated that Lb. casei and Lb. plantarum were able to interact with Peyer's patch cells and showed an increase in IgA-, CD4+ cells, and antibodies specific for the stimulating strain. Lactobacillus acidophilus induced gut mucosal activation by interaction with the epithelial cells without increase in the immune cells associated with the bronchus. Although Lb. rhamnosus and Strep. salivarius ssp. thermophilus interact with epithelial cells, they also induced an immune response against their epitopes. Lactococcus lactis and Lb. delbrueckii ssp. bulgaricus induced an increase of IgA+ cells entering the IgA cycle but not CD4+ cells; thus, these bacteria would have been bound to epithelial cells that activated B lymphocytes without processing and presenting of their epitopes. We did not determine specific antibodies against Lc. lactis or Lb. bulgaricus.     

Molecular methods for identification of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus using methionine biosynthesis and 16S rRNA genes

Yoghurt and starter culture producers are still searching strains of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus to produce healthier yogurt with longer shelf life, better texture, taste and quality. However, selective identification of Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus from a mixed population using microbiological and biochemical methods is difficult, time consuming and may not be accurate. In this study, a quick, sensitive and accurate method is proposed to identify both Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus using PCR. The method is comprised of two parts. In the first part, methionine biosynthesis genes, known to be present in both species were partially amplified by designed primers (cysmet2F and cysmet2R). Partial amplification of the methionine biosynthesis gene which gives 700 bp fragment resulted in selective identification of Lb. bulgaricus and Strep. thermophilus. All 16 Lb. bulgaricus and 6 Strep. thermophilus isolates assessed by this method gave the expected amplification. On the other hand, further analysis of other closely related species with the same primers have indicated that the same product was also amplified in two more lactobacilli namely, Lb. delbrueckii subsp. lactis and Lb. helveticus species. Thus, in the second part of the method, further differentiation of Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus from each other and these species was achieved using restriction analysis of 16S rRNA gene with EcoRI.     

定量PCR技术检测杀菌型产品中乳酸菌

目的利用定量PCR(quantitativePCR,qPCR)技术检测杀菌型产品中乳酸菌,并与标签标示相比较,查探产品中各类菌种的异同性及数量。方法以市售的发酵乳及含乳饮料作为研究对象,设计可区分产品中常见的6种乳酸菌特异性引物及探针,利用qPCR技术对方法的特异性、灵敏度进行验证,并建立标准扩增曲线;利用模拟样品对方法进行检验,最后对市场上购买的实际样品进行检测。结果除去干酪乳杆菌,其他5种引物都能特异性扩增出其目标菌,并且对其他的乳酸菌均无扩增现象;DNA检测的灵敏度可达到2～4pg;单一标准菌种扩增曲线线性较好,相关系数r2值在0.954～0.992之间;对模拟样品的检测,与平板法差异性比较,P值均大于0.05,无显著性差异。对市售样品进行检测,产品中标记的干酪乳杆菌、双歧杆菌和嗜热链球菌与检测结果存在不匹配现象。结论本研究建立的qPCR方法可快速、准确地对产品中德氏乳杆菌保加利亚亚种、嗜热链球菌、乳酸乳球菌、嗜酸乳杆菌、双歧杆菌的鉴定和定量的分析。     

Manufacture of Fior di Latte cheese by incorporation of probiotic lactobacilli

This work aimed to select heat-resistant probiotic lactobacilli to be added to Fior di Latte (high-moisture cow milk Mozzarella) cheese. First, 18 probiotic strains belonging to Lactobacillus casei, Lactobacillus delbrueckii ssp. bulgaricus, Lactobacillus paracasei, Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus reuteri were screened. Resistance to heating (65 or 55°C for 10 min) varied markedly between strains. Adaptation at 42°C for 10 min increased the heat resistance at 55°C for 10 min of all probiotic lactobacilli. Heat-adapted L. delbrueckii ssp. bulgaricus SP5 (decimal reduction time at 55°C of 227.4 min) and L. paracasei BGP1 (decimal reduction time at 55°C of 40.8 min) showed the highest survival under heat conditions that mimicked the stretching of the curd and were used for the manufacture of Fior di Latte cheese. Two technology options were chosen: chemical (addition of lactic acid to milk) or biological (Streptococcus thermophilus as starter culture) acidification with or without addition of probiotics. As determined by random amplified polymorphic DNA-PCR and 16S rRNA gene analyses, the cell density of L. delbrueckii ssp. bulgaricus SP5 and L. paracasei BGP1 in chemically or biologically acidified Fior di Latte cheese was approximately 8.0 log(10)cfu/g. Microbiological, compositional, biochemical, and sensory analyses (panel test by 30 untrained judges) showed that the use of L. delbrueckii ssp. bulgaricus SP5 and L. paracasei BGP1 enhanced flavor formation and shelf-life of Fior di Latte cheeses.     

Effect of Versagel on the growth and metabolic activities of selected lactic acid bacteria

ABSTRACT:  In this study, the influence of a protein-based fat replacer, Versagel ^® added at 1% and 2% (w/v) to reconstituted skim milk (RSM), on the growth and metabolic activities of selected strains of yogurt starters ( Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus ) as well as probiotic organisms ( Lactobacillus casei , Lactobacillus acidophilus , and Bifidobacterium longum ) was examined. Addition of Versagel resulted in significantly improved growth of S. thermophilus and B. longum but inhibited that of L. casei , L. acidophilus , and L. delbrueckii ssp. bulgaricus . This is also reflected in the extent of reduction of pH in RSM with added Versagel by these organisms. Among the biochemical activities, proteolytic activity of all the organisms except B. longum was adversely affected by the presence of Versagel, although the angiotensin-I converting enzyme (ACE) inhibitory and α-glucosidase (α-glu) inhibitory activities were improved. Versagel at 1% level influenced the growth, while ACE inhibitory and α-glu inhibitory activities of the organisms were better at 2% level.     

Effect of fermented milk containing probiotic bacteria in the prevention of an enteroinvasive Escherichia coli infection in mice

This study investigated the protective capacity of the oral administration of fermented milk containing the probiotic strains; Lactobacillus casei, Lb. delbrueckii subsp. bulgaricus and Streptococcus thermophilus, against enteroinvasive Escherichia coli infection in a murine (BALB/ c mice) model. Mice were fed for 2, 5 or 7 consecutive days with fermented milk diluted to a concentration of viable Lb. casei, Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus of 10(7) cfu/ml. Phagocytic activity of peritoneal macrophages and the number of IgA+ cells in small and large intestine were determined at the end of the feeding periods. For the preventive effect against Esch. coli, animals were fed for 5 days (selected dose). Mice were challenged with an infective dose of enteroinvasive Esch. coli of 10(8) cfu/mouse. The colonization of liver and spleen and the secretory IgA specific for the pathogen in the intestinal fluid were determined (ELISA test). Results showed that the unspecific immune response enhanced itself after 5 consecutive days of the administration of this fermented milk (increase in the percentage of phagocytosis and number of IgA+ cells in the small intestine). Treated animals showed less Esch. coli colonization of liver than control mice and a higher secretory anti-Esch. coli IgA in the intestinal fluids. These results suggest that the protection against enteroinvasive Esch. coli infection observed for the fermented milk containing probiotic bacteria may be associated with an enhance of the intestinal mucosa immunity.     

Effect of milk base and starter culture on acidification,texture, and probiotic cell counts in fermented milk processing

In the present work, the compared effect of milk base and starter culture on acidification, texture, growth, and stability of probiotic bacteria in fermented milk processing, was studied. Two strains of probiotic bacteria were used, Lactobacillus acidophilus LA5 and L. rhamnosus LR35, with two starter cultures. One starter culture consisted only of Streptococcus thermophilus ST7 (single starter culture); the other was a yogurt mixed culture with S. thermophilus ST7 and L. bulgaricus LB12 (mixed starter culture). For the milk base preparation, four commercial dairy ingredients were tested (two milk protein concentrates and two casein hydrolysates). The resulting fermented milks were compared to those obtained with control milk (without enrichment) and milk added with skim milk powder. The performance of the two probiotic strains were opposite. L. acidophilus LA5 grew well on milk but showed a poor stability during storage. L. rhamnosus LR35 grew weakly on milk but was remarkably stable during storage. With the strains tested in this study, the use of the single starter culture and the addition of casein hydrolysate gave the best probiotic cell counts. The fermentation time was of about 11 h, and the probiotic level after five weeks of storage was greater than 106 cfu/ml for L. acidophilus LA5 and 10(7) cfu/ml for L. rhamnosus LR35. However, an optimization of the level of casein hydrolysate added to milk base has to be done, in order to improve texture and flavor when using this dairy ingredient.