Cheese Ripening: Impact on Cheese as a Functional Food

Abstract

One of the most popular types of fermented dairy products is cheese. The process of cheese aging is essential for improving cheese quality, and health benefits. Ripened cheese at different times acquired wide diversity of characteristic aromas and textures due to establishing a cascade of intrinsic complex biochemical and metabolic outcomes, resulting in a dynamic shift in microbial flora. Various functional bioactive compounds could be released during the cheese ripening process. Many strategy approaches are employed to accelerate cheese ripening based on increasing lipolysis and proteolysis rate. During cheese aging, microbial spoilage as early and late blowing may occur so, designing smart ripening rooms are very essential equipped with computerized monitoring systems including sensors, software platforms, temperature, and humidity data loggers.

1. Introduction

Cheesemaking depends on the concentration of milk proteins with or without milk fat. After the salting and packing processes, cheese acquired a longer shelf life. Around the world, cheeses can be produced using a variety of milk sources, processing methods, starter culture, coagulants, and ripening conditions, giving rise to a large number of variants with a vast diversity in terms of texture, flavor, and shape [1]. Cheese ripening (aging or maturing) is the most crucial industrial stage in cheese technology, which establishes a cascade of complex biochemical steps, producing a wide variety of microbial flora and different volatile compounds. This is the critical stage during which the cheese’s firmness, aroma, flavor, and other specific cheese characteristics are acquired. Ripening occurs under temperature and humidity circumstances that differ depending on the type of cheese. The longer the cheese ripens, the less moisture it retains and the firmer and stronger-tasting it becomes. Cheese kinds are actually defined based on how they ripen; Brie, Camembert, and Roqueforti are examples of mold-ripened cheese, while Limburger and Tilsit are examples of surface-ripened cheese. Internally-ripened cheeses fall into six categories: semi-hard cheeses like Monterey Jack, hard cheeses like Cheddar and extra-hard cheeses like Parmesan and Asiago; pasta filata, which includes mozzarella and provolone; high-salt cheeses like Feta; cheeses with eyes like Dutch types (Gouda and Edam) and Swiss types (Emmental and Gruyere) [2]. The curd’s remaining citrate is metabolized by some citrate-positive lactic acid bacteria (LAB), like Lactococcus lactis subsp. Cremoris, and Lactococcus lactis subsp. lactis biovar. diacetylactis, producing a variety of taste compounds, including acetoin, acetate, diacetyl, 2-butanone, and 2,3-butanediol, which are linked to the development of Dutch cheeses [3].

The development of flavor and sensorial characteristics in cheese is significantly influenced by ripening conditions, especially the time factor. Each variety of cheese has a distinct volatile chemical at a variable concentration, and flavor which could be measured by a diverse series of methodologies, and computational and descriptive approaches [4, 5]. Several bioactives and healthy peptides are produced from milk components during ripening as a result, mainly, of its degradation by starter cultures endo and exo-enzymes [6].

2. Factors affecting the cheese ripening process

2.1 Cheese microbiota

In the cheese industry, characterization of the cheese microbiota is crucial, since some specific microorganisms improve cheese characteristics while others may decrease quality [7]. The cheese microbiota is a sophisticated ecosystem that can develop from raw milk, acidifying starters, and adjunct cultures, as well as a group of prokaryotic, eukaryotic, and viral populations that may arise from machinery and the environment of the cheese manufacturing plant.

LAB are the majority microbiota with a significant role during cheese production and ripening. The shape, numbers, and proportions of LAB are then shaped by cheese making and ripening which exert a selection pressure on microorganisms. Diverse microbiota members may interact cooperatively and competitively, which could have an impact on ripened cheese’s rheological, organoleptic, and safety properties [8, 9]. The progression of various microbial species and their interaction during the cheese-making and -ripening processes, in particular, are crucial for the formation of the distinctive sensory characteristics of each cheese variety [10, 11].

Notably, LAB are a kind of Gram-positive bacteria with the ability to withstand acidic pH, and they are primarily cocci or rods at the microscopical examination. The LAB can convert milk carbohydrates to lactic acid. There are more than sixty genera of LAB, the most common genera found in food for fermentation are Lactobacillus, Leuconostoc, Pediococcus, Enterococcus, Lactococcus, Streptococcus, and Weissella [12, 13]. The natural bacterial population of LAB in milk is the fundamental basis for the manufacture and technology of raw milk cheeses. A recent study conducted in Italy indicates that the microbial population of milk, particularly that used to produce Grana Padano PDO cheese, may be impacted by the multifaceted nature of farming practices. The implementation of stringent hygiene measures and management levels in the dairy farm affects negatively the presence of diverse milk microbiota and decreases bacterial load [14].

LAB play a significant role in raw milk cheeses like Parmigiano Reggiano (the long-ripened hard cheeses) both as starter cultures (SLAB) during curd acidification and as non-starter cultures (NSLAB) during cheese-ripening [15, 16, 17].

The majority of facultative heterofermentative lactobacilli, or other terms called NSLABs, are frequently isolated from cheese [18]. For instance, one common NSLAB isolated from long-ripened, hard-cooked cheeses is the Lactobacillus caseigroup, which includes Lacticaseibacillus rhamnosus, Lacticaseibacillus paracasei, and Lacticaseibacillus casei species. These species contribute to the formulation of the distinctive cheese flavor during the ripening process [19].

During cheese ripening, a complicated sequence of activities happens, resulting in a series of biological reactions. Proteolysis is one of the most critical processes, which is launched by the starter, followed by non-starters, and completed by proteolytic enzymes secreted by the bacterial population. These mechanisms cause conformational changes in the particular peptides and amino acid composition, which continually alters during the aging process. The author Sforza et al. [20] discovered a high association between peptide evolution and enzyme activity, allowing for the classification of cheeses based on their ripening circumstances.

Another earlier study established a relation between the hard cheese Parmigiano Reggiano microbiota with the proteolysis rate that takes place during ripening, which is the cause for the valuable and high-quality characteristics of long-ripened cheeses. The study found the raw milk microbiota for different samples from various dairies were quite similar on the curd level. Once ripening time progressed the microbial composition changed, revealing significant differences between cheese samples at early ripening storage time (1 and 2 months) and at late ripening storage (time 7 and 9 months old). These were attributed to NSLAB species, which are more correlated with different peptide profiles and the significant difference in kinetics and activities of the proteolytic enzymes. This study underlines the critical role of NSLAB in ripened cheese proteolysis. Additionally, the potential use of several peptides as indicators of a specific microbial composition allows for the preservation and valorization of cheese’s specificity and connection to its production location [21].

The reaction between propionic acid bacteria and LAB influences the organoleptic properties of the produced final cheese product as texture, flavor, and ripening stability. Specific inoculation doses from Propionibacterium freudenreichii strains, one of the propionic acid bacteria spp., usually chosen, are utilized for the manufacture of Emmental cheeses and other Swiss-type cheeses which under propionic acid fermentation produce characteristic taste with distinctive eye formation [22].

A study investigated that the amount of free lysine, glycine, and methionine that is available in cheeses is influenced by propionibacteria and LAB. The highest content of free amino acids was presented in the mature cheeses that were produced by combining mesophilic LAB Propionibacterium and L. casei strains. Compared to the mesophilic LAB starter culture or the addition of L. casei, it was clear that the addition of Propionibacteria had a greater impact on the free amino acid content in matured cheese samples. Notably, this study demonstrates that the growth of Propionibacteria in milk is exaggerated by mesophilic LAB, resulting in the production of a significant amount of free amino acids [23]. Not only changing the cheese’s body and texture and forming the flavors that give it its distinct character, chemical reactions that take place on the surface of the cheese during ripening also provide an extra protection layer against dry storage conditions [24]. Furthermore, during maturation, the surface microbiota grows quickly, and the residing microorganisms have inhibitory characteristics that hinder pathogenic foodborne bacteria or molds that produce mycotoxin from colonizing the cheese [25, 26]. Numerous enzymes can initiate chemical reactions which may originate from natural milk enzymes, rennet extract utilized in manufacture, or produced by bacteria that survive pasteurization or are added during the process of manufacture or ripening. For instance, three distinct strains of Brevibacterium linens were used as surface inoculants to initiate the formation of red smears during the fermentation of Munster cheese [27]. Environmental pathogens pose a significant risk to the ripening and manufacturing processes of bloomy rind cheeses, such as Camembert, Brie, and related varieties. Due to the product’s exposure to the open air during ripening, this risk is especially elevated [28]. The most growing fungi include Penicillium candidum, Kluyveromyces marxianus, and Geotrichum candidum [29]. All predominantly used fungi have a great role in ripening and characteristic organoleptic properties [30].

Cheeses that have been ripened by mold may be distinguished into two main types: those with blue veins and the other with surface mold-ripened cheeses or bloomy rinds [31]. The development of a particular mold called Penicillium roqueforti gives Roqueforti, also referred to as blue-veined cheeses, characterized by unique flavor and appearance. Roqueforti cheese is produced in several cities across the globe. Each of these countries’ distinct blue cheese varieties has its own characteristics and methods of manufacture [32].

The microbiota of cheese has been shown to have anti-cancer and cholesterol-lowering capabilities in addition to participating in the enhancement of cheese flavor through the synthesis of volatile molecules [33, 34, 35]. The cheese microbial dynamic is affected by the interactions among some factors as LAB used as SLAB or NSLAB, cheese-making processes, and some storage conditions [36, 37, 38]. Owing to the cheese microbiota being correlated to the quality and physicochemical properties of cheese, it became critical to understand the cheese microbial properties. In a study reported by Choi et al. [39], the post-inoculation cheese microbiota was found to be dominated by SLAB, and the authors observed that the addition of SLAB resulted in modifications to the microbial community structure, microbial diversity, biomarkers, and predicted functional qualities. Additionally, 105 and 119 days after age, undefinable Lactobacillus, or NSLAB, were found.

2.2 Enzymes

The primary factor in turning milk into cheese is the presence of enzymes, which can be found in the milk itself or introduced as rennet or microbial enzymes. The disintegration of caseins is by far the most significant of the enzymatic processes. Rennet, native milk proteinase, and peptidases generated by starter cultures, enzymes of secondary starters, and enzymes of non-starter cultures are the five primary systems that aid in the hydrolysis of casein [40].

Depending on the cheese type, the ripening time for cheeses made with rennet might range from a few weeks to several years. Microbiological and biochemical alterations take place during ripening, giving the variety’s distinctive flavor and texture their development. Primary (lipolysis, proteolysis, and metabolism of residual lactose, lactate, and citrate) or secondary (metabolism of fatty acids and amino acids) processes can be used to categorize biochemical changes in cheese during ripening. Early in the ripening process, lactate is quickly produced from residual lactose. An essential precursor for several processes, such as racemization, oxidation, and microbial metabolism, is lactate. In certain types, the metabolism of citrate is quite important. Cheese’s lipolysis is aided by lipases derived from several sources, especially milk [5].

Cheese ripening can be accelerated by increasing the concentration of important enzymes used in cheese production. Plasmin, chymosin, and intracellular and/or cell wall proteinases and peptidases of the LAB and NSLAB are among the proteinases and peptidases found in cheese. Proteases and lipases from animals or fungi are frequently utilized in the production of enzyme-modified cheese. It is uncommon, though, for these enzymes to be used directly to enhance the flavor and ripening of cheese. The primary disadvantages of this approach are the limited availability of commercial enzymes that have been approved for use in cheese ripening and the incapacity to blend the enzymes into the cheese matrix. Exogenous enzymes can be either single enzymes or commercial enzyme combinations. Exogenous enzymes can be added directly into cheese blocks, in combination with starter cultures or coagulants, with cheese-milk, or at the stage of dry salting. When making cheddar cheese, the latter process is employed [41]. Exogenous enzymes were supposed to accelerate cheese ripening and aid in the production of unique tastes in specific cheese varieties. In that regard, a few examples of distinct enzymes or their mixes were reviewed and documented [5].

2.3 Dairy animal feeding

It was worth noting that dietary supplementation of lactating dairy cows may change the quality of dairy products, especially cheese products. The characteristic volatile substances that contribute to cheese flavor were attributed to the aromatic qualities of milk obtained from lactating dairy ruminants. Extensive research has been conducted concerning lactating dairy ruminants fed specific experimental diets, such as those distinguished, for example, by the addition of trace elements, natural supplements, or agricultural byproducts rich in bioactive compounds. Cheese contains a variety of volatile substances, such as carboxylic acids, lactones, ketones, alcohols, and aldehydes. The relative amounts of each substance are determined by the biochemical processes that take place during ripening. These processes are primarily mediated by endogenous enzymes and elements of bacterial origin whose function can be greatly influenced by the bioactive substances consumed by animals in their diet and released in milk through the mammary gland. According to Ianni et al. [42] there was a significant correlation between the quality of the biochemical changes in cheese products throughout ripening and the various dairy animal feeding practices.

2.4 Level of sodium chloride

It is well-recognized that consuming too much sodium raises the risk of hypertension, cardiovascular disease, and even stomach cancer [43, 44]. The issue for the food industry is reformulating food products that contain less sodium aiming to offer low-salt food in the human diet is currently one of the top priorities for public health organizations [45]. To comply with the World Health Organization’s (WHO) recommendation of 2 g/day, it has been suggested by the World Health Organization that sodium intake be reduced by 30% [46].

Food businesses must carefully re-evaluate the composition and processing of high-sodium foods considering public awareness of excessive sodium intake and nutrition claims connected to salt content. Although it is usual practice to replace some ingredients in products through reformulation, it is still difficult to reduce salt in cheese due to sodium chloride’s various functions and essential activities in cheese making. Salt improves the taste and fragrance profile, controls the texture, final pH, and water activity, and influences microbiological growth. It also favors the drainage of remaining whey. In the end, salt concentration affects the shelf-life of cheese by regulating the activity of starter and non-starter LAB during cheese production and ripening. Any adjustment to the salting process, such as lowering the sodium chloride (NaCl) amount or substituting alternative salting agents, could upset the delicate balance within the parameters, changing the cheese’s quality. Depending on the kind of cheese and manufacturing method (for example, soft, semi-hard, hard, and mold-ripened cheeses), the decrease of NaCl content may be treated differently. As a result, specific tactics could be implemented to preserve the general quality and safety of various cheese types [47].

As a result, making cheeses with less NaCl content is becoming increasingly popular in the dairy business. Since NaCl is essential for textural qualities, microbial development, autolysis, enzyme activity during ripening, and ultimately cheese flavor, hence, reducing NaCl in cheese-making poses a number of technological, sensory, and microbiological issues [48]. Reduced NaCl semi-hard cheeses typically have enhanced cohesiveness, adhesiveness, acidity, bitterness, and an unpleasant aftertaste along with a decrease in salt and hardness as their sensory defaults [49, 50]. Additionally, reducing the NaCl concentration of cheese affects product safety due to the possibility of germs like Listeria monocytogenes growing in the product [51].

According to popular perception, cheese contains varying quantities of salt, depending on the type of cheese. NaCl content for soft, semi-hard, and hard cheeses ranges from 0.5% to 2.5%, whereas it ranges from 3% to 5% for blue-type cheeses. In cheese, NaCl performs a variety of crucial tasks, including modifying the curd and rind’s physical characteristics, regulating the microbiota’s growth as the cheese ripens, preventing the formation of infections or spoilers, and enhancing the flavor [52, 53].

In a study followed the strategy of reduction of NaCl or its partial substitution with other salts such as potassium chloride in a semihard cheese (Reblochon), it was reported that lowering the salt level in the semi-hard cheese samples caused spoiler growth to accelerate, as seen by increased Pseudomonas species formation and increased cheese proteolysis and lipolysis [45].

3. Bioactive components produced during cheese ripening

Bioactive peptides are released during the ripening process of cheese and are protein fragments in the form of short amino acid sequences. During cheese maturation, various functional bioactive compounds could be generated such as volatile fatty acids, exopolysaccharides, vitamins, organic acids, peptides, and amino acids (γ-aminobutyric acid and conjugated linolenic acid). Many laboratory and animal research studies demonstrated that most of these generated bioactive compounds exhibited different biological activities including antihypertensive, antioxidant, anticancer, and antimicrobial activities [54, 55, 56]. The above bioactivities lead to health-protective effects associated with a reduced incidence of cardiovascular disease risk factors, such as obesity, dyslipidemia, and type 2 diabetes [57], as well as reduced incidence of metabolic syndrome [58, 59]. The artisanal methods employed for the manufacturing of Mexican cheeses, depending on natural milk indigenous microflora, may offer many potential health benefits. A previous study [60] demonstrated that peptides derived from both fresh cheese and a model cheese (laboratory scale) had an antihypertensive peptide through suppressing ACE-inhibitory effect. Although artisanal cheese is under storage conditions, producing different product varieties such as (Crema de Chiapas, Cocido, and Fresco of Sonora). Few limited studies are available for artisanal cheeses in Mexico and the various derived bioactive compounds in artisanal cheeses, and those available focus primarily on the antioxidant and ACE activity of water-soluble extracts (WSE) obtained from different types of artisanal cheese varieties such as (Crema de Chiapas, Cocido, and Fresco of Sonora) from different storage conditions [61, 62].

Distinct peptide sets found in different cheese varieties contribute to their distinctive flavors and possible health benefits. These bioactive molecules support the body’s defense against potentially harmful substances by acting as antioxidants, anti-inflammatory agents, and even antimicrobial activities. Angiotensin-converting enzyme (ACE) inhibitory characteristics have been demonstrated by some cheese peptides, indicating that they may lower blood pressure by preventing the angiotensin-converting enzyme from acting [63].

Protein breakdown is typically linked to biological functional qualities and is aided by the action of milk-specific enzymes. Furthermore, the microorganisms used or added during cheese production may produce bioactive peptides [64]. Milk proteases and peptidases produce large and intermediate-sized peptides, which are then hydrolyzed by enzymes from the cheese SLAB and NSLAB strains. These are known as primary proteolysis reactions [65].

Both the body’s digestive processes and the fermentation procedures used to produce fermented dairy products produce functional peptides. They are produced by the proteins; casein and whey. When liberated from their original proteins, these peptides take on an active role. When they enter the circulatory system, they may have a systemic effect, or they can be absorbed in intact form and have different physiological effects locally in the gut. It is well known that lactoferroxins and casooxins function as opioid antagonists, while casomorphins and lactophorins generated from milk proteins are opioid agonists. The opioids’ analgesic qualities are comparable to aspirin. The actions of casokinins (which lower blood pressure), casoplatelins (which reduce blood clotting), immunopeptides (which boost immune function), and phosphopeptides (which carry minerals) are all examples of casokinin action. Casein phosphopeptides may improve calcium, phosphorus, and magnesium bioavailability for better bone health. They may also aid in the prevention of dental cavities and play a function in the secretion of enterohormones and immunological boosting. The involvement of casein peptides in blood pressure regulation looks promising. Certain casein and whey protein hydrolyzates prevent the conversion of angiotensin I to angiotensin II. Because angiotensin II elevates blood pressure by constricting blood vessels, inhibiting it causes blood pressure to fall. Dairy foods would thus be a natural functional food for managing hypertension due to their ACE inhibitory action. There are several commercially marketed whey products that include discrete bioactive peptides. The glycomacropeptide (GMP) is produced by proteolysis from kappa-casein [5, 63].

Proteins and lipids are the sources of the most common and extensively researched bioactive molecules. Cheese proteins are broken down to form bioactive peptides and amino acids such as gamma-aminobutyric acid and ornithine, while fats are hydrolyzed to produce and be an origin of bioactives like conjugated linoleic acid, carotenoids, fat-soluble vitamins, and sphingolipids, among other things. Depending on their nature, these chemicals exhibit various bioactivities [6, 35].

High quantities of lactic acid and other organic acids are produced by LAB during the fermentation of lactose [66]. The ripening process is a complicated process in which several milk enzymes participate, including rennet and enzymes from LAB, which leads to successive transformations that aim to affect the various curd ingredients [40, 67].

Proteinases found in LAB contribute to the proteolysis of cheese proteins, converting them into oligopeptides that can either be further degraded into shorter peptides and amino acids through the synergistic action of various intracellular peptidases produced by specific LAB. These peptides, amino acids, and their derivatives help the final cheese to develop its texture, flavor, and health benefits. The generated peptides were subjected to both in vitro laboratory experiments and in vivo animal models, which confirmed that they possessed a variety of biological activities, including the ability to scavenge free radicals (antioxidants), inhibit microbial growth (antimicrobial), fight inflammation (anti-inflammatory), immunity enhancer (immunomodulatory), opioid blocker receptors activity (analgesic), and lower blood pressure by suppressing the angiotensin-converting enzyme producing antihypertensive effect. Furthermore, some LAB produce vitamins, some antimicrobials, conjugated linoleic acid, and other functional lipids with anti-inflammatory and anticarcinogenic properties, as well as bioactives that contribute to physiological processes like neurotransmission and hypotension induction with diuretic effects [63].

During the early weeks of cheese ripening, NSLAB proliferate at a very slow rate, but eventually take control of the cheese microbiota following the death phase of the starter culture [68, 69]. The process of cheese ripening is sophisticated and a dynamic process. The variety of proteolytic enzymes naturally found in milk and the remaining coagulants, as well as the enzymatic metabolism of LAB, are crucial to this process [70]. Peptides are continuously released during ripening by the action of plasmin and LAB’s enzymes; some of these peptides are then digested, while others accumulate throughout storage [71, 72].

One of the major milk groups that can enhance the sensory qualities, shelf life, and microbial safety of cheese as well as the nutritional content and functional features of the finished products is NSLAB. It includes; Pediococcus, Enterococcus, Lactobacillus, and Leuconostoc genera. Due to the creation of certain metabolites throughout the ripening process, ripened cheese develops a diversity of flavors, nutritional qualities, and rheological characteristics [73, 74, 75]. Peptides, amino acids, biogenic amines, nucleic acids, carbohydrates, organic acids, vitamins, polyphenols, alkaloids, minerals, and any other molecules that can alter the sensory and rheological properties, as well as the nutritional value and health benefits of the finished products, are examples of metabolites [73].

The aforementioned functional biological activities have been linked to a decreased incidence of most common coronary artery disease risk factors, such as dyslipidemia, diabetes type 2, and overweight [57], as well as different metabolic syndromes [59].

As shown in Table 1 various bioactive components in different ripened cheese.

Type of bioactive component	Type of ripened cheese	Reference
1. Antioxidant peptides	Cheddar	[76, 77]
	Coalho	[78]
	Fresco	[79]
	Parmigiano-Reggiano	[80]
	Cottage cheese	[81]
2. ACE-inhibitory activity	Mozzarella, Italico	[82]
	Red cheddar, Camembert	[83]
	Emmental	[84]
	Manchego, Ronca, and goat cheeses	[85]
	Gouda	[33]
3. Gamma amino butyric acid (GABA)	Artisanal Spanish cheese	[86]
	Italian Pecorino Marchigiano and Pecorino Filiano cheeses	[87]
	Lighvan cheese (traditional semi hard Iranian cheese)	[88]
4. Organic acids	Ossalano cheese	[89]
	Cheddar cheese	[90]

Table 1.

List of some published bioactive compounds found in ripened cheese varieties.

4. Ripened cheese additives

One of the simplest and most traditional methods for preserving food to increase its shelf life is the direct addition of additives which may be chemical or naturally derived preservatives. These ingredients are added to cheese in order to prevent spoilage and pathogenic microbial growth, increase shelf life, enhance organoleptic and sensory characteristics, and maintain nutritional value [91, 92].

Food preservatives permitted for use in matured cheeses in the EU are classified into three functional groups: antimicrobials, antioxidants, and antibrowning chemicals. During cheesemaking, these compounds are added to the milk vat as antimicrobials and antioxidants, or to the cheese as surface defenders against unwanted agents. Lysozyme, sorbic acid/sorbates, nisin, natamycin, hexamethylene tetramine (HTM), nitrates/nitrites, and propionic acid/propionates are authorized additions for matured cheeses. As illustrated in Table 2 different cheese additives are added to cheese milk for enhancement of the cheese ripening process.

Cheese additive	Character	Function	Reference
Nisin	A bacteriocin with the designation E234 has been accepted as a food additive by the European Food Safety Authority (EFSA) and the Food and Drug Administration (FDA). Nisin is a 34-residue protein that is mostly generated by Lactococcus lactis strains	Nisin has an antibacterial action against a variety of Gram-positive foodborne and spoilage bacteria	[93, 94, 95, 96]
Natamycin	A bacteriocin that is produced by aerobic fermentation of Streptomyces natalensis and its related species; More than 40 nations have approved it as a food additive, and the FDA regards it as a GRAS (generally recognized as safe) substance	commonly employed antifungal to prevent yeasts and molds; contamination in dairy products (hard and semi-hard cheese)	[97]
Lysozyme	A naturally occurring enzyme that is prevalent in egg whites, from which it is typically separated for industrial preservation	It has some bactericidal properties against Gram positive bacteria like LAB and clostridia but less effective against Gram negative bacteria. Lysozyme (E-1105) has been employed to prevent the “late blowing” problem in hard and semi-hard cheeses	[98, 99, 100]
Ripening enzymes	Like proteinases, peptidases, and lipases produced by; Penicillium candidum	Strategies for accelerating the cheese ripening process by addition of liposome-encapsulated enzymes to cheese milk provide some definite advantages	[101, 102]

Table 2.

Additives to cheese milk and its role in cheese ripening.