Lesson 22. CHEMICAL, PHYSICAL, MICROBIOLOGICAL AND SENSORY CHANGES
Module 9. Changes during ripening of cheese
Lesson 22
CHEMICAL, PHYSICAL, MICROBIOLOGICAL AND SENSORY CHANGES
22.1 Cheese Ripening
Unique characteristics of each cheese variety are determined by the curd manufacturing operations but these characteristics are largely developed during ripening process. For example, the type of microflora established in the curd is determined by the manufacturing process but its effect on cheese characteristics develop largely during the ripening process.
Ripening involves microbial and chemical changes which are responsible for development of typical characteristics of varieties of cheeses. Microbial changes involve death and lysis of the starter cells, development of non-starter microflora and growth of secondary microflora. Ripening usually causes softening of the cheese texture due to hydrolysis of the casein matrix, change in pH and change in water binding ability of the curd. Flavor production is largely described by a series of biochemical events taking place during ripening.
The primary events occuring during cheese ripening are metabolism of residual lactose, lactate metabolism, proteolysis and lipolysis. These reactions are mainly responsible for textural changes and development of flavor in cheese. However, many secondary changes occur simultaneously and modify cheese texture and flavor. Since the biochemistry of cheese ripening is complex, the objective of this chapter is to present an overview of the principal biochemical pathways contributing towards cheese ripening.
Ripening involves microbial and chemical changes which are responsible for development of typical characteristics of varieties of cheeses. Microbial changes involve death and lysis of the starter cells, development of non-starter microflora and growth of secondary microflora. Ripening usually causes softening of the cheese texture due to hydrolysis of the casein matrix, change in pH and change in water binding ability of the curd. Flavor production is largely described by a series of biochemical events taking place during ripening.
The primary events occuring during cheese ripening are metabolism of residual lactose, lactate metabolism, proteolysis and lipolysis. These reactions are mainly responsible for textural changes and development of flavor in cheese. However, many secondary changes occur simultaneously and modify cheese texture and flavor. Since the biochemistry of cheese ripening is complex, the objective of this chapter is to present an overview of the principal biochemical pathways contributing towards cheese ripening.
22.2 Metabolism of Residual Lactose
Lactic acid bacteria (LAB) is added in the form of starter culture to cheese metabolize lactose to lactate. The rate and extent of acidification influence texture of the curd by controlling the rate of demineralization. The pH of the cheese curd is largely determined by the extent of acidification during manufacturing process. This influences the solubility of the casein, which in turn affects the texture of curd. pH also affects the activity of enzymes involved in ripening, thereby having an indirect effect on cheese texture and flavor development.
Most of the lactose is lost in whey during cheese manufacturing. However, low levels of lactose remains in the curd. This residual lactose is converted to L-lactate during early stages of ripening by the action of starter bacteria. The rate of conversion is dependent on temperature and salt-in-moisture levels of the curd. Starter activity is stopped very quickly at the end of manufacturing operations due to low pH, salt addition and lesser amount of fermentable lactose. Lactose that remains unfermented by the starter is probably metabolized by non-starter lactic acid bacteria (NSLAB) flora present in curd and they convert the residual lactose to D-lactate. D-lactate can also be formed by the racemisation of L-lactate.
Most of the lactose is lost in whey during cheese manufacturing. However, low levels of lactose remains in the curd. This residual lactose is converted to L-lactate during early stages of ripening by the action of starter bacteria. The rate of conversion is dependent on temperature and salt-in-moisture levels of the curd. Starter activity is stopped very quickly at the end of manufacturing operations due to low pH, salt addition and lesser amount of fermentable lactose. Lactose that remains unfermented by the starter is probably metabolized by non-starter lactic acid bacteria (NSLAB) flora present in curd and they convert the residual lactose to D-lactate. D-lactate can also be formed by the racemisation of L-lactate.
22.3 Lactate Metabolism
Lactate produced by fermentation of residual lactose serves as an important substrate for a range of reactions occuring during cheese ripening. L-lactate can be racemised to D-lactate by NSLAB flora. D-lactate is less soluble than L-lactate which results in the formation of Ca-D-lactate crystals. These crystals are not harmful but they appear as white specks on the surface of the mature cheese.
Lactose can be metabolized to acetate, ethanol, formate and CO2 depending on the population of NSLAB and availability of O2. In cheese wrapped with film, oxidation of lactate occurs to a lesser extent due to low level of O2 available.
Late gas blowing is a defect which is caused by anaerobic metabolism of lactate by Clostridium tyrobutyricum to butyrate and H2. The release of H2 causes cracks in cheese during ripening.
The above mentioned metabolisms contribute negatively towards cheese ripening. There are some positive contributions also of lactate metabolism. This is essential for cheese varieties characterized by the development of large eyes during ripening such as Emmental cheese. Propionibacterium freudenrichii metabolise lactate to propionate, acetate, CO2 and H2O. Propionate and acetate contribute to the flavor of cheese while CO2 is mainly responsible for eye formation.
Lactose can be metabolized to acetate, ethanol, formate and CO2 depending on the population of NSLAB and availability of O2. In cheese wrapped with film, oxidation of lactate occurs to a lesser extent due to low level of O2 available.
Late gas blowing is a defect which is caused by anaerobic metabolism of lactate by Clostridium tyrobutyricum to butyrate and H2. The release of H2 causes cracks in cheese during ripening.
The above mentioned metabolisms contribute negatively towards cheese ripening. There are some positive contributions also of lactate metabolism. This is essential for cheese varieties characterized by the development of large eyes during ripening such as Emmental cheese. Propionibacterium freudenrichii metabolise lactate to propionate, acetate, CO2 and H2O. Propionate and acetate contribute to the flavor of cheese while CO2 is mainly responsible for eye formation.
22.4 Lipolysis
Milk fat is essential for the development of the correct flavor in cheese during ripening. Cheddar and other cheeses normally made from whole milk do not develop correct flavor when made from skim milk or milks in which milk fat has been replaced with other lipids. Lipids may undergo oxidative or hydrolytic degradation in foods but the redox potential of cheese is very low, so mainly hydrolytic degradation of lipids takes place in cheese. The triglycerides present in cheese are hydrolyzed by lipases which result in the formation of fatty acids.
Sources of lipases in cheese are:-
• Milk (particularly unpasteurized)
• Rennet
• Starter culture
• Starter adjuncts
• Non starter bacteria (may come through ingredients or contamination)
• Exogenous lipase (if added deliberately)
• Rennet
• Starter culture
• Starter adjuncts
• Non starter bacteria (may come through ingredients or contamination)
• Exogenous lipase (if added deliberately)
Low level of lipolysis is required for the development of flavor of cheese but excessive lipolysis causes rancidity. Lipolysis of milk fat results in production of free fatty acids which contribute to the flavor of cheese and also act as precursors for development of other flavor compounds in cheese like esters, lactones, ketones and aldehydes. These secondary fat-derived compounds can be very potent flavor compounds.
Fatty acid esters are produced by reaction of fatty acids with an alcohol; ethyl esters are most common in cheese. Thioesters are formed by reaction of a fatty acid with a thiol compound formed via the catabolism of sulphur-containing amino acids. Fatty acid lactones are formed by the intramolecular esterification of hydroxyacids; γ- and δ-lactones contribute to the flavor of a number of cheese varieties. n-methyl ketones are formed by the partial β-oxidation of fatty acids.
Liberation of short and medium chain fatty acids from milk fat by lipolysis contribute directly to cheese flavor. This degree of flavor development depends on the variety of cheese. For example, it is very extensive in some hard Italian varieties, smear cheeses and blue mold cheeses. Excessive lipolysis causes rancidity in cheese varieties like Cheddar and Gouda.
Fatty acid esters are produced by reaction of fatty acids with an alcohol; ethyl esters are most common in cheese. Thioesters are formed by reaction of a fatty acid with a thiol compound formed via the catabolism of sulphur-containing amino acids. Fatty acid lactones are formed by the intramolecular esterification of hydroxyacids; γ- and δ-lactones contribute to the flavor of a number of cheese varieties. n-methyl ketones are formed by the partial β-oxidation of fatty acids.
Liberation of short and medium chain fatty acids from milk fat by lipolysis contribute directly to cheese flavor. This degree of flavor development depends on the variety of cheese. For example, it is very extensive in some hard Italian varieties, smear cheeses and blue mold cheeses. Excessive lipolysis causes rancidity in cheese varieties like Cheddar and Gouda.
Fig. 22.1 Production of flavor compounds from fatty acids during cheese ripening
(Source: Fox et al., 2000)
22.5 Proteolysis
Proteolysis is the most important and complex of all the events during ripening of cheese. The extent and pattern of proteolysis is also used as an index of cheese ripening and quality of cheese. Proteolysis contributes significantly towards development of texture and flavor in cheese. Textural changes (softening of cheese curd) occur due to breakdown of protein network and release of carboxyl and amino groups resulting in the binding of more water and thus decrease water activity (aw). Proteolysis leads to the formation of peptides and free amino acids which contribute to cheese flavor. These amino acids also act as precursors for many reactions like transamination, deamination, decarboxylation, desulphuration, catabolism of aromatic compounds such as tyrosine, phenylalanine, tryptophan, etc. and generate many important flavor compounds. Proteolysis in cheese is catalysed by proteinases and peptidases and they originate from the following sources:
• Coagulant
• Milk
• Starter LAB
• Non starter LAB
• Secondary starters (e.g. P. camemberti in Camembert cheese and P. roqueforti in Blue cheese)
• Exogenous proteinases or peptidases, if added for accelerated ripening of cheese
• Milk
• Starter LAB
• Non starter LAB
• Secondary starters (e.g. P. camemberti in Camembert cheese and P. roqueforti in Blue cheese)
• Exogenous proteinases or peptidases, if added for accelerated ripening of cheese
In majority of cheese varieties, casein is initially hydrolyzed by the residual coagulant, often chymosin, which results in formation of large and intermediate-sized peptides. These peptides are then hydrolyzed by enzymes derived from starter and non-starter microflora of the cheese. The production of small peptides and amino acids is caused by the action of microbial proteinases and peptidases, respectively.
The final products of proteolysis are amino acids, the concentration of which depends on the cheese variety. The concentration of amino acids in cheese at a given stage of ripening is the net result of the liberation of amino acids from the caseins by proteolysis and their catabolism or transformation into other amino acids by the cheese microflora. Medium and small peptides contribute to a brothy background flavor in many cheese varieties; short, hydrophobic peptides are bitter. Amino acids contribute directly to cheese flavor as some amino acids taste sweet (e.g. Gly, Ser, Thr, Ala, Pro), sour (e.g. His, Glu, Asp) or bitter (e.g. Arg, Met, Val, Leu, Phe, Tyr, Ile, Trp).
The final products of proteolysis are amino acids, the concentration of which depends on the cheese variety. The concentration of amino acids in cheese at a given stage of ripening is the net result of the liberation of amino acids from the caseins by proteolysis and their catabolism or transformation into other amino acids by the cheese microflora. Medium and small peptides contribute to a brothy background flavor in many cheese varieties; short, hydrophobic peptides are bitter. Amino acids contribute directly to cheese flavor as some amino acids taste sweet (e.g. Gly, Ser, Thr, Ala, Pro), sour (e.g. His, Glu, Asp) or bitter (e.g. Arg, Met, Val, Leu, Phe, Tyr, Ile, Trp).
Fig. 22.2 Proteolysis in cheese during ripening
(Source: Fox et al., 2000)
(Source: Fox et al., 2000)
22.6 Microbiology of Cheese Ripening
Microorganisms including bacteria, yeasts and molds are present in cheese and contribute to the ripening process through their metabolic activity. The enzymes released by these microorganisms also add to the various metabolic activities like proteolysis, glycolysis and lipolysis.
Microorganisms in cheese may gain entry through their intended addition in the form of starter culture and they may be associated with the ingredients used in cheese making. The microflora associated with cheese ripening may be divided into two groups - the starter bacteria and non-starter bacteria. Starter bacteria are primarily responsible for acid production during manufacture to reduce the pH of milk to the desired level. The secondary microflora do not play any active role during cheese manufacture but contribute to the ripening process.
The factors controlling the growth of microorganisms in cheese include water activity, concentration of salt, oxidation-reduction potential, pH, ripening temperature, and the presence or absence of bacteriocins (produced by some starters).
22.6.1 Water activity
Water activity (aw) is defined as the ratio of the vapor pressure of water in a material (p) to the vapor pressure of pure water (po) at the same temperature. Its value ranges from 0 to 1.0. It expresses the water availability rather than total water present in the system. Water activity of cheese reduces during ripening process. This may be due to several reasons like:
• Evaporation of moisture if the cheese is not vacuum packed or paraffin coated
• Hydration of proteins bound water rendering it unavailable for bacterial growth
• Hydrolysis of proteins to peptides and amino acids and of lipids to glycerol and fatty acids
• The salt and organic acids (lactate, acetate, and propionate) dissolved in the moisture of the cheese reduce the vapor pressure
• Hydration of proteins bound water rendering it unavailable for bacterial growth
• Hydrolysis of proteins to peptides and amino acids and of lipids to glycerol and fatty acids
• The salt and organic acids (lactate, acetate, and propionate) dissolved in the moisture of the cheese reduce the vapor pressure
Growth of microorganisms at low aw is characterized by a long lag phase, a slow rate of growth, and a reduction in the maximum number of cells produced. Each of these factors helps to limit the number of cells produced. LAB generally have higher minimum aw values than other bacteria. The amount of salt in moisture in cheese also affects the growth of microorganisms in cheese.
The salt normally used in cheese making is about 2% of the weight of the curd. Salt is added to cheese mainly to suppress growth of unwanted microorganism and to assist the physico-chemical changes in the curd. The growth of unwanted microorganisms is essentially curbed by reduction in aw as salt act as a humectant.
The salt normally used in cheese making is about 2% of the weight of the curd. Salt is added to cheese mainly to suppress growth of unwanted microorganism and to assist the physico-chemical changes in the curd. The growth of unwanted microorganisms is essentially curbed by reduction in aw as salt act as a humectant.
22.6.2 Oxidation-Reduction potential
The oxidation-reduction potential (Eh) is a measure of the tendency of the solution to either gain or lose electrons when it is subject to change by introduction of a new species. The Eh of milk is about +150 mV whereas that of cheese is about -250 mV. As cheese ages, the products of proteolysis and lipolysis may reduce the Eh of cheese. This reduction of Eh makes cheese an anaerobic system, in which only facultatively or obligately anaerobic microorganisms can grow. Anaerobic sporeforming organisms present in the cheese may germinate and grow, causing defects like bitter and putrid flavor. Obligate aerobes, like Pseudomonas spp., Brevibacterium spp., and Micrococcus spp., will not grow within the cheese, even when other conditions for growth are favorable. Eh is therefore important in determining the types of microorganisms that grow in cheese.
22.6.3 pHMost bacteria require a neutral pH value for optimum growth and grow poorly at pH values below 5.0. The pH of cheese curd after manufacture generally lies within the range 4.5-5.3, so pH is also a significant factor in controlling bacterial growth in cheese. LAB, especially lactobacilli, generally has pH optima below 7, and Lactobacillus spp. can grow at pH 4.0. Most yeasts and molds can grow at pH values below 3.0, although their optimal range is from 5 to 7. B. linens, an important organism in smear-ripened cheese, cannot grow below pH 6.0. Micrococcus spp., which are commonly found on the surface of soft cheeses, cannot grow at pH 5 and only slowly at pH 5.5.
22.6.4 Temperature
The ripening temperature of cheese mainly depends on two considerations: the temperature should be such that the growth of undesirable spoilage causing and pathogenic bacteria can be checked and at the same time, this temperature should be conducive for various ripening reactions essential for development of typical flavor, body and texture of cheese. Ripening temperature for Cheddar cheese is 6-8°C while Camembert and other mold-ripened cheeses are ripened at 10-15°C. Emmental cheese is ripened initially for 2-3 weeks at a low temperature (~ 12°C), after which the temperature is increased to 20-24°C for 2-4 weeks to promote the growth of propionic acid bacteria and the fermentation of lactate to propionate, acetate, and CO2. The temperature is then reduced again to around 4°C. Use of higher ripening temperature is one of the techniques used for accelerated ripening of cheese but it also stimulates the growth of other microorganisms present in cheese.
Last modified: Wednesday, 3 October 2012, 10:16 AM