Accelerated Maturation of Cheddar Cheese: Microbiology of Cheeses Supplemented with Lactobacillus casei subsp. casei L2A

Growth of lactic acid bacteria (LAB) and lactobacilli was studied in Cheddar cheeses supplemented with live and heat-shocked Lactobacillus casei subsp. casei L2A and with Neutrase® to accelerate maturation. Bacterial counts of treated cheeses rapidly reached maximal values within 1 wk, whereas the control cheese reached comparable values only after 2 mo. Addition of 1.0% heat-shocked lactobacilli led to an excellent quality Cheddar cheese with a 50% increase in flavor development, as determined by sensory evaluation, compared to control cheese. Addition of Neutrase (1 × 10^-5 AU/g cheese) permitted a gain of an additional 10% while addition of higher concentrations (2 and 4 × 10^-5 AU/g cheese) resulted in undesirable bitterness.     

Lactic Acid Bacteria Relation to Accelerated Maturation of Cheddar Cheese

Cheddar cheese was supplemented with cell homogenates and/or live cells of Lactobacillus casei-casei L2A. Two concentrations of cell homogenates under two forms, liquid and lyophilized, were compared as well as two stages of addition, renneting and salting. Growth of lactic acid bacteria and lactobacilli was studied during the period of maturation. Addition of live lactobacilli and lyophilized homogenate at renneting led to a good-quality matured cheese with 40% increase in flavor intensity compared to control cheeses.     

Effect of Added Lactobacilli on Composition and Texture of Cheddar Cheese During Accelerated Maturation

The evolution of cheese composition and texture was studied in maturing Cheddar cheese supplemented with live cells and cell homogenates of Lactobacillus casei-casei L2A in order to accelerate maturation. The pH was significantly modified by the lactic acid of the bacterial additives. The Theological properties showed the same general pattern of evolution in experimental as in control cheeses. The process we developed has led to a good-quality matured cheese with 40% increase in flavor intensity compared to control cheeses.     

Quality Attributes of Cheddar Cheese Containing Added Lactobacilli

Various strains of homo- and heterofermentative lactobacilli isolated from cheddar cheese were added to milk with a commercial streptococci culture to produce cheddar cheese. The heterofermatative lactobacilli L. brevis and L. fermentum almost always led to the development of fruity flavors, openness and late-gassing within 10 months of aging. Cheddar cheese produced using combined cultures of heterofermentative lactobacilli and L. casei-casei or L. casei-pseudoplantarum did not exhibit gas formation and openness. The overall grading scores of cheese containing added lactobacilli were not higher than those for the control cheese (without lactobacilli). A definite correlation was found between the lactobacilli used and the flavors of the cheese. The controlled acidity development during cheese making, the fat and the salt in moisture content of the cheeses were not affected.     

Microbial and chemical composition of Cheddar cheese supplemented with prebiotics from pasteurized milk to aging

Microbial and chemical properties of cheese is crucial in the dairy industry to understand their effects on cheese quality. Microorganisms within this fat, protein, and water matrix are largely responsible for physiochemical characteristics and associated quality. Prebiotics can be used as an energy source for lactic acid bacteria in cheese by altering the microbial community and provide the potential for value-added foods, with a more stable probiotic population. This research focuses on the addition of fructooligosaccharides (FOS) or inulin to the Cheddar cheese-making process to evaluate the effects on microbial and physicochemical composition changes. Laboratory-scale Cheddar cheese produced in 2 replicates was supplemented with 0 (control), 0.5, 1.0, and 2.0% (wt/wt) of FOS or inulin using 18 L of commercially pasteurized milk. A total of 210 samples (15 samples per replicate of each treatment) were collected from cheese-making procedure and aging period. Analysis for each sample were performed for quantitative analysis of chemical and microbial composition. The prevalence of lactic acid bacteria (log cfu/g) in Cheddar cheese supplemented with FOS (6.34 ± 0.11 and 8.99 ± 0.46; ± standard deviation) or inulin (6.02 ± 0.79 and 9.08 ± 1.00) was significantly higher than the control (5.84 ± 0.27 and 8.48 ± 0.06) in whey and curd, respectively. Fructooligosaccharides supplemented cheeses showed similar chemical properties to the control cheese, whereas inulin-supplemented cheeses exhibited a significantly higher moisture content than FOS and the control groups. Streptococcus and Lactococcus were predominant in all cheeses and 2% inulin and 2% FOS-supplemented cheeses possessed significant amounts of nonstarter lactic acid bacteria found to be an unidentified group of Lactobacillaceae, which emerged after 90 d of aging. In conclusion, this study demonstrates that prebiotic supplementation of Cheddar cheese results in differing microbial and chemical characteristics.     

Impact of Using Exopolysaccharides (EPS)-Producing Strain on Qualities of Half-Fat Cheddar Cheese

Yield, textural, proteolysis, melting, and sensory properties of exopolysaccharide-producing Lactobacillus paracasei on properties of half-fat (about 16 g fat/100 g cheese) Cheddar cheese during ripening at 8℃ for up to six months were investigated. The results revealed that B-3 cheese, made with 2.0% (v/v) high yield exopolysaccharide-producing L. paracasei in combination with 0.011% (w/w) commercial Cheddar culture (B-3 cheese), had a 10.15, 7.71, and 10.04% separately increase in moisture content and had a 7.70, 5.05, and 6.76% separately increase in yield compared with B-2, B-4, and B-5 cheese, texture and melting characteristics were significantly improved (P < 0.05), sensory score surpassed B-4 and B-5 cheese and was similar to the full-fat one. Any differences of B-3 cheese detected among half-fat Cheddar cheeses were attributed to the presence of high yield exopolysaccharide-producing L. paracasei.     

Size-Exclusion HPLC Separation of Bitter and Astringent Fractions from Cheddar Cheese Made with Added Lactobacillus Strains to Accelerate Ripening

Lactobacillus strains were added as an adjunct to the regular lactic starter in Cheddar cheese manufacture in order to accelerate ripening. Microbial cheese proteolysis resulted in the release of free amino acids which were extracted with the astringent and bitter fractions and separated by size-exclusion and reversed-phase HPLC chromatography. Lactobacillus strains generally increased the degree of proteolysis. L. plantarum and L. brevis produced off-flavors possibly due to an accumulation of medium-size peptides. The control cheese (without lactobacilli) had the most peptides with a mean molecular- weight of < 1000 daltons and had a flavor described as slightly bitter. Addition of L. casei-casei L2A accelerated ripening and yielded a well-aged Cheddar cheese without any bitterness even after 7 months at 6°C.     

Evolution of Free Amino Acids during the Ripening of Cheddar Cheese Containing Added Lactobacilli Strains

Cheddar cheese was produced with different lactobacilli strains added to accelerate ripening. The concentration of proteolytic products was determined as free amino acids in the water-soluble fraction at two, four, seven and nine months of aging and at two different maturation temperatures (6°C, 15°C). All amino acids increased during ripening and were higher in the Lactobacillus- added cheeses than in the control cheese, and higher in cheeses ripened at 15°C than at 6°C. Glutamic acid, leucine, phenylalanine, valine and lysine were generally in higher proportion in all cheeses. The cheeses with added L. casei-casei L2A were classified as having a “strong Cheddar cheese” flavor after only seven months of ripening at 6°C.     

复合酶在pH值自然变化下水解干酪工艺条件的研究

采用Flavourzyme和Neutrase双酶组合水解切达干酪制备乳滋味增强物质,利用单因素优化了双酶组合的作用条件。实验结果表明,在总酶添加量为8%的条件下,复合酶水解的最佳工艺条件为:F酶与N酶比例为5∶3,水解温度45℃,pH值8.0,底物浓度4%,水解12 h。在此工艺条件下水解产物的三氯乙酸-氮(TCA-N)增加百分数为68.3%,并且其滋味评分与添加有相同浓度的市售干酪香精最为接近,乳滋味浓郁,无苦味、异味。     

Enhanced nutty flavor formation in cheddar cheese made with a malty Lactococcus lactis adjunct culture

Nutty flavor in Cheddar cheese is desirable, and recent research demonstrated that 2- and 3-methyl butanal and 2-methyl propanal were primary sources of nutty flavors in Cheddar. Because malty strains of Lac-tococcus lactis (formerly Streptococcus lactis var. malti-genes) are characterized by the efficient production of these and other Strecker aldehydes during growth, this study investigated the influence of a malty L. lactis adjunct culture on nutty flavor development in Cheddar cheese. Cheeses made with different adjunct levels (0, 10⁴ cfu/mL, and 10⁵ cfu/mL) were ripened at 5 or 13°C and analyzed after 1 wk, 4 mo, and 8 mo by a combination of instrumental and sensory methods to characterize nutty flavor development. Cheeses ripened at 13°C developed aged flavors (brothy, sulfur, and nutty fla-vors) more rapidly than cheeses held at 5°C. Additionally, cheeses made with the adjunct culture showed more rapid and more intense nutty flavor development than control cheeses. Cheeses that had higher intensities of nutty flavors also had a higher concentration of 2/3-methyl butanal and 2-methyl propanal compared with control cheeses, which again confirmed that these compounds are a source of nutty flavor in Cheddar cheese. Results from this study provide a simple methodology for cheese manufacturers to obtain consistent nutty flavor in Cheddar cheese.     

Liposome encapsulated nisin Z: optimization,stability and release during milk fermentation

Liposomes prepared from different proliposomes (Pro-lipo^® H, Pro-lipo^® S, Pro-lipo^® C and Pro-lipo^® DUO) were tested for their capacity to encapsulate nisin Z. Factors affecting the entrapment process (pH and concentration of nisin Z solution and cholesterol concentration in the lipid membranes) were optimized. The use of nisin Z monoclonal antibodies made it possible to quantify, using a competitive enzyme immunoassay, and visualize, using transmission electron microscopy, nisin Z. Nisin Z was entrapped in different liposomes with encapsulation efficiencies (EE) ranging from 9.5% to 47%. The pH of nisin Z aqueous solution and nisin Z concentration had a significant effect on the amount of encapsulated nisin. An increase in cholesterol content in lipid membranes up to (20%, w/w) resulted in a slight reduction in EE. Nisin-loaded vesicles did not severely disturb Cheddar cheese fermentation and showed stability to Cheddar cheese temperature cycle. Long term stability of liposome-encapsulated nisin Z was demonstrated for 27 days at 4°C in different media including milk with different fat levels (3.25%, 2.0% and 1.0%), skim milk, sweet whey and phosphate buffer saline (PBS). Liposome stability determined as the quantity of released nisin Z was highest in milk followed by PBS and whey, respectively.     

Proteolytic pattern and organic acid profiles of probiotic Cheddar cheese as influenced by probiotic strains of Lactobacillus acidophilus,Lb. paracasei,Lb. casei or Bifidobacterium sp.

Cheddar cheeses were produced with starter lactococci and Bifidobacterium longum 1941, B. lactis LAFTI^® B94, Lactobacillus casei 279, Lb. paracasei LAFTI^® L26, Lb. acidophilus 4962 or Lb. acidophilus LAFTI^® L10 to study the survival of the probiotic bacteria and the influence of these organisms on proteolytic patterns and production of organic acid during ripening period of 6 months at 4 °C. All probiotic adjuncts survived the manufacturing process of Cheddar cheese at high levels without alteration to the cheese-making process. After 6 months of ripening, cheeses maintained the level of probiotic organisms at >8.0 log₁₀ cfu g⁻¹ with minimal effect on moisture, fat, protein and salt content. Acetic acid concentration was higher in cheeses with B. longum 1941, B. lactis LAFTI^® B94, Lb. casei 279 and Lb. paracasei LAFTI^® L26. Each probiotic organism influenced the proteolytic pattern of Cheddar cheese in different ways. Lb. casei 279 and Lb. paracasei LAFTI^® L26 showed higher hydrolysis of casein. Higher concentrations of free amino acids (FAAs) were found in all probiotic cheeses. Although Bifidobacterium sp. was found to be weakly proteolytic, cheeses with the addition of those strains had highest concentration of FAAs. These data thus suggested that Lb. acidophilus 4962, Lb. casei 279, B. longum 1941, Lb. acidophilus LAFTI^® L10, Lb. paracasei LAFTI^® L26 and B. lactis LAFTI^® B94 can be applied successfully in Cheddar cheese.     

Impact of ropy and capsular exopolysaccharide-producing strains of Lactococcus lactis subsp. cremoris on reduced-fat Cheddar cheese production and whey composition

Cheddar cheese mixed starter cultures containing exopolysaccharide (EPS)-producing strains of Lactococcus lactis subsp. cremoris (Lac. cremoris) were characterized and used for the production of reduced-fat Cheddar cheese (15% fat). The effects of ropy and capsular strains and their combination on cheese production and physical characteristics as well as composition of the resultant whey samples were investigated and compared with the impact of adding 0.2% (w/v) of lecithin, as a thickening agent, to cheese milk. Control cheese was made using EPS-non-producing Lac. cremoris. Cheeses made with capsular or ropy strains or their combination retained 3.6–4.8% more moisture and resulted in 0.29–1.19 kg/100 kg higher yield than control cheese. Lecithin also increased the moisture retention and cheese yield by 1.4% and 0.37%, respectively, over the control cheese. Lecithin addition also substantially increased viscosity, total solid content and concentrating time by ultra-filtration (UF) of the whey produced. Compared with lecithin addition, the application of EPS-producing strains increased the viscosity of the resultant whey slightly, while decreasing whey total solids, and prolonging the time required to concentrate whey samples by UF. The amount of EPS expelled in whey ranged from 31 to 53 mg L⁻¹. Retention of EPS-producing strains in cheese curd was remarkably higher than that of non-producing strains. These results indicate the capacity of EPS-producing Lac. cremoris for enhanced moisture retention in reduced-fat Cheddar cheese; these strains would be a promising alternative to commercial stabilizers.     

Influence of Homofermentative Lactobacilli on Physicochemical and Sensory Properties of Cheddar Cheese

Cheddar cheeses were produced under pilot plant conditions using a commercial Streptococcus culture amended with one of 10 homofermentative Lactobacillus strains. During the ripening period, pH, acidity, salt, moisture, fat, texture, fissure formation, gas development and sensory status were evaluated. Lactobacillus treated cheese did not differ much from the control in pH and acidity but acidity increased substantially after draining and cheddaring. Lactobacillus numbers increased at all stages as compared with the uninoculated control. High quality Cheddar cheese was produced by L. casei-subsp-casei (119-10/62) and L. casei-subsp-pseudoplantarum (137-10/62) from 7 to 12 vats aged for 2 months at 15°C and for a further 10 months at 7°C or 15°C. Fissure formation was observed in cheese made with L. casei-subsp-rhamnosus, one of the four cultures of L. casei-subsp-casei (LH13) and two of the three strains of L. casei-subsp-pseudoplantarum (83-4-12/62 and L3E). Certain Lactobacillus strains produced cheese with slight flavor defects. Other strains, in particular L. casei-subsp-rhamnosus, contributed to high acidity (72 - 0.89° domic) and low pH (5.2) at salting.     

Isolation and characterisation of exopolysaccharide-producing Weissella and Lactobacillus and their application as adjunct cultures in Cheddar cheese

This study characterised exopolysaccharide-producing lactic acid bacteria and examined their potential for use in Cheddar cheese manufacture. Two strains were chosen for incorporation as adjunct cultures in Cheddar cheese manufacture: namely, the homopolysaccharide-producers Weissella cibaria MG1 and Lactobacillus reuteri cc2. These strains both produce dextrans with molecular masses ranging from 10⁵ to 10⁷ Da. Both strains were used in the production of miniature Cheddar cheeses that employed a conventional commercial cheese starter culture Lactococcus lactis R604. A cheese was also included that used purified dextran as an ingredient. The W. cibaria strain survived in cheese with levels increasing by 1.5 log cycles over the ripening period. All experimental cheeses (adjunct or exopolysaccharide ingredient) had higher moisture levels compared with the control cheese made using starter alone. Inclusion of the adjunct strains had no detectable negative effects on cheeses in terms of proteolysis.     

Comparison of growth and survival of single strains of Lactococcus lactis and Lactococcus cremoris during Cheddar cheese manufacture

Traditionally, starter cultures for Cheddar cheese are combinations of Lactococcus lactis and Lactococcus cremoris. Our goal was to compare growth and survival of individual strains during cheesemaking, and after salting and pressing. Cultures used were 2 strains of L. lactis (SSM 7605, SSM 7436) and 2 strains of L. cremoris (SSM 7136, SSM 7661). A standardized Cheddar cheese make procedure was used that included a 38°C cook temperature and salting levels of 2.0, 2.4, 2.8, 3.2, and 3.6% from which were selected cheeses with salt-in-moisture levels of 3.5, 4.5, and 5.5%. Vats of cheese were made using each strain on its own as biological duplicates on different days. Starter culture numbers were enumerated by plate counting during cheesemaking and after 6 d storage at 6°C. Flow cytometry with fluorescent staining by SYBR Green and propidium iodide was used to determine the number of live and dead cells in cheese at the different salt levels. Differences in cheese make times were strain dependent rather than species dependent. Even with correction for average culture chain length, cheeses made using L. lactis strains contained ～4 times (～0.6 log) more bacterial cells than those made using L. cremoris strains. Growth of the strains used in this study was not influenced by the amount of salt added to the curd. The higher pH of cheeses with higher salting levels was attributed to those cheeses having a lower moisture content. Based on flow cytometry, ～5% of the total starter culture cells in the cheese were dead after 6 d of storage. Another 3 to 19% of the cells were designated as being live, but semipermeable, with L. cremoris strains having the higher number of semipermeable cells.     

Accelerated Cheddar Cheese Ripening with an Aminopeptidase Fraction from Squid Hepatopancreas

ABSTRACT: An aminopeptidase (AP) fraction from squid (Illex illecebrosus) hepatopancreas was added to Cheddar cheese at 2 levels, and its influence on ripening indices was determined for up to 3-mo storage at 11 °C. Two commercial enzymes (Neutrase ® and Flavourzyme ®) were similarly tested. Cheese with the higher level of squid AP contained more soluble N, amino acids, and Cheddar flavor after 1 mo, but it developed defects in texture and bitterness as ripening progressed. Cheese with less squid AP did not differ from the control with respect to all ripening indices over 3-mo storage. Ripening Cheddar contains cysteine protease inhibitor(s) that inhibit low levels of squid AP but not Neutrase ® and Flavourzyme ®.     

The isolation and identification of yeasts obtained during the manufacture and ripening of Cheddar cheese

Sources of yeast, which may contaminate the curd during the manufacture of Cheddar cheese, were examined in a single cheese factory. A total of 77 yeast species present in the factory environment, manufacturing and ripening of Cheddar cheese were identified according to cellular long-chain fatty acid analysis and verified with conventional identification techniques. Product line samples were taken at critical control points in the manufacturing process and analysed after incubation at 25°C for 96 h. The progression of yeast species during cheese-making and ripening was monitored after renneting and at subsequent 48-h intervals. Dominant species wereDebaryomyces hanseniiandCryptococcus albidus, whileYarrowia lipolytica, Rhodotorula minuta, Torulaspora delbrueckii, Rhodotorula glutinisandKluyveromyces marxianuswere present at low numbers. The results obtained showed that yeasts were present in all cheese samples examined, at quantities ranging from 9×10²to 1·4×10⁷cfu g⁻¹.     

Isomaltooligosaccharide increases the Lactobacillus rhamnosus viable count in Cheddar cheese

Five batches of Cheddar cheese were manufactured containing different levels of isomaltooligosaccharide (IMO) and a probiotic strain of Lactobacillus rhamnosus to study the effect of IMO on the survival of starter lactococci and probiotic micro‐organisms, on proteolytic patterns, cheese composition and sensory properties. The cheese was exposed to conditions simulating those found in the gastrointestinal tract to evaluate the survival of Lb. rhamnosus. Results demonstrated that the addition of Lb. rhamnosus and IMO did not affect the main compositional variables of Cheddar cheese. The counts of starter culture and probiotic organisms increased in cheese which contained Isomaltooligosaccharide (Batches 3, 4 and 5) more than in the control (Batches 1 and 2) during the fermentation. The probiotic counts in fresh cheese (B‐4) was 9.23 log₁₀ cfu/g which was more than one log cycle greater than in the control (B‐2). The probiotic counts remained above 8 log₁₀ cfu/g at the end of the manufacturing process. Primary proteolysis was not affected by the addition of probiotic bacteria and IMO, but the level of secondary proteolysis was slightly higher compared with the control group. The addition of IMO improved the texture and sensory quality of the cheese and the probiotic bacterium had the same effect. Under conditions that simulated the gastrointestinal tract, the probiotic bacteria in cheese (B‐4) exhibited good survival and remained above the recommended 6–7 log₁₀ cfu/g.     

Use of high pressure treatment to attenuate starter bacteria for use as adjuncts for Cheddar cheese manufacture

Attenuated starter bacteria cannot produce acid during cheese manufacture, but contain enzymes that contribute to cheese ripening. The aim of this study was to investigate attenuation of starter bacteria using high pressure treatment, for use in combination with a primary starter for Cheddar cheese manufacture, and to determine the effect of such adjunct cultures on secondary proteolysis during ripening. Lactococcus lactis ssp. cremoris HP and L. lactis ssp. cremoris 303 were attenuated by pressure treatment at 200 MPa for 20 min at 20 °C. Cheddar cheese was manufactured using untreated cultures of both these starter strains, either alone or in combination with their high pressure-treated equivalents. High pressure-treated starters did not produce acid during cheese manufacture and starter counts in cheeses manufactured using high pressure-treated starter did not differ from those of the controls. Higher levels of cell lysis were apparent in cheese manufactured using high pressure-treated strains than in the controls after 26 d of ripening. Small differences were observed in the peptide profiles of cheeses, analysed by reversed-phase HPLC; cheeses manufactured using high pressure-treated starters also had slightly higher levels of amino acids than the relevant controls. Overall, addition of high pressure-treated starter bacteria as a secondary starter culture accelerated secondary proteolysis in Cheddar cheese.

Industrial relevance

Attenuated starters provide extra pool of enzymes, which can influence cheese ripening, without affecting the cheese making schedule. This paper presents an alternative method for attenuation of starter bacteria using high pressure treatment and their subsequent use to accelerate secondary proteolysis in Cheddar cheese during ripening.