cheesecoursemanual2012_0

5/21/2018 CheeseCourseManual2012_0

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Cheese Making Technology

Art Hill, 2011


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HOW TO USE THIS MANUAL

This manual, updated July 2011, is intended to be used as a supplement to the lectures andlaboratories of Cheese Technology,a short course by the Department of Food Science,University of Guelph. It is not designed to be a stand-alone reference, but many have found

previous editions useful as general reference to cheese science and technology. The onlineversion is posted for personal use only. If you refer to this document in any oral or writtenmaterials please provide the authors name (Art Hill) and enough information to easily locatethe online document.

A list of recommended references for further reading is included in Chapter 2. When thesereferences are cited else where in the manual, they are identified by author and year. Youcan then find the reference in the alphabetical list in Chapter 2. Other citations are noted inparentheses or foot notes as appropriate.

This manual is not written in a formal style. Some stylistic devices conform to convention. In

other cases I have created my own conventions. The following are some of my quirks. When referring to the milk of specific species, I have abandoned all attempts to

determine where to put the esses and apostrophes. So, sheeps, goats, cows,reindeers and yaks milk are simplified to sheep, goat, cow, reindeer and yak milk.

Some sections are written in point form rather than prose. The most important acronyms are defined on the following page. Others that are used

only in a particular section are defined where they first occur in the text. Percentages mean percent by weight unless otherwise noted.

Read on ---- I hope you enjoy it and that you find useful information.

Professor Art HillDepartment of Food Science,University of Guelph, Guelph, ON N1G 2W1Email: [email protected]: 519-824-4120 x53875Fax: 519-824-6631


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ABBREVIATIONS

Acronym Long Form Comment

CN Casein number Weight % of casein in the total

(crude) protein of milk.

FDM Fat in the dry matter 100 x Fat/(100 - moisture)

LAB Lactic acid bacteria

MNFS Moisture in the non-fat substance 100 x Moisture/(100 - fat)

NFS Non-fat solids The trade also uses the acronym SNF(Solids non-fat) .

NPN Non-protein nitrogen Nitrogenous materials present inmilk that are often included in

estimates of total milk protein. Crudeprotein means NPN is included in thetotal; true protein means NPN isexcluded. If protein is not specifiedas crude or true, assume it is crude.

NSLAB Non-starter lactic acid bacteria

PF Ratio of protein to fat in the milk PF is the principal determinant of theFDM in the cheese.

pH A measure of acidity Neutral = 7.0

Acid < 7.0Alkaline > 7.0

SM Salt as weight % of cheesemoisture

100 x Salt/moisture

TA Titratable acidity Expressed as weight % of lactic acid.


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TABLE OF CONTENTS

1. INTRODUCTION TO CHEESE MAKING ........................................................................ 1

2. RECOMMENDED REFERENCES ................................................................................... 11

3. PROCESS AND QUALITY CONTROL PROCEDURES ................................................ 133.1 Introduction ........................................................................................................... 133.2 Cheese Sampling .................................................................................................. 153.3 Total Solids .......................................................................................................... 163.4 Titratable Acidity ................................................................................................. 163.5 pH ........................................................................................................................ 183.6 Babcock Methods for Milk Fat ............................................................................ 20

3.7 Cheese Salt ........................................................................................................... 22

3.8 Culture Activity Test ........................................................................................... 223.9 Detection of Bacteriophage ................................................................................. 233.10 Inhibitory Substances ......................................................................................... 243.11 Rennet Activity .................................................................................................. 273.12 Yeasts and Moulds ............................................................................................. 273.13 Presumptive Coliforms ...................................................................................... 283.14 Staphylococci ................................................................................................... 28

4. RAW MILK QUALITY .................................................................................................... 314.1 The Principal Milk Components ......................................................................... 314.2 Factors affecting gross milk composition ............................................................. 32

4.3 Milk as a growth medium ..................................................................................... 334.4 Types of micro organisms and their activity in milk ............................................ 364.5 Pathogenic Bacteria ............................................................................................. 374.6 Antibiotics ............................................................................................................. 384.7 Mastitic Milk ......................................................................................................... 394.8 Raw Milk quality tests ......................................................................................... 40

5. TREATMENT OF MILK FOR CHEESE MAKING ......................................................... 455.1 Clarification .......................................................................................................... 455.2 Standardization of cheese milk composition ....................................................... 455.3 Heat treatments ..................................................................................................... 49

5.4 Homogenization .................................................................................................... 505.5 Additives to Cheese milk ...................................................................................... 50

6. STANDARDIZATION OF MILK FOR CHEESE MAKING ........................................... 536.1 PF, FDM and CN ................................................................................................ 536.2 Methods of Standardizing .................................................................................... 53


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6.3 Units ..................................................................................................................... 546.4 Calculations ........................................................................................................ 556.5 Addition Of Cream .............................................................................................. 596.6. General Guidelines for Standardization ............................................................... 59

7. CULTURES ........................................................................................................................ 647.1 General Functions of Cheese Cultures ................................................................. 647.2 General characteristics of lactic acid cultures ...................................................... 657.3 Classification of Lactic Acid Cultures .................................................................. 657. 4 Summary: technological properties of lactic acid cultures ................................. 677.5 Secondary Cultures .............................................................................................. 687.6 Culture Production, Distribution and Storage...................................................... 687.7 Bacteriophage (bacterial viruses).......................................................................... 69

8. COAGULATION ............................................................................................................... 738.1 Milk Structure ....................................................................................................... 73

8.2 Enzymatic Coagulation of Milk ........................................................................... 748.3 Acid coagulation .................................................................................................. 778.4 Heat-Acid coagulation .......................................................................................... 77

9. CHEESE MAKING STEP BY STEP ................................................................................. 809.1 Ripening the Milk ................................................................................................. 809.2 Setting the Vat ...................................................................................................... 809.3 Cutting The Curd .................................................................................................. 819.4 Cooking ................................................................................................................. 839.5 Draining ................................................................................................................ 839.6 Washing ................................................................................................................ 83

9.7 Curd Handling ....................................................................................................... 839.8 Pressing ................................................................................................................. 849.9 Salting ................................................................................................................... 84

10. RIPENING AND PACKAGING ...................................................................................... 8810.1 Ripening processes: chemical and physical changes .......................................... 8810.2 Principal Ripening Agents .................................................................................. 90

10.3 Cheese Composition for Optimal Curing ........................................................... 9210.4 Temperature of Curing ........................................................................................ 9310.5 Humidity of Curing ............................................................................................. 9310.6 Ripening Treatments ........................................................................................... 94

10.7 Packaging ............................................................................................................ 94

11. PROCESS CONTROL ..................................................................................................... 9611.1 The Objectives of Cheese Manufacturing .......................................................... 9611.2 Moisture Control ................................................................................................. 9611.3 pH Control ......................................................................................................... 97


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11.4 Mineral Control .................................................................................................. 9711.5 Texture Control ................................................................................................... 9811.6 Flavour Control .................................................................................................. 98

12. YIELD EFFICIENCY ................................................................................................... 101

12.1 Distribution of Components During Cheese Making ....................................... 10112.2 Factors Affecting Yield .................................................................................... 10112.3 Principles of Yield Optimization ..................................................................... 10212.4 Yield Control .................................................................................................... 10212.5 Recovery of Milk Components ......................................................................... 10212.6 Yield Expression ............................................................................................... 10312.7 Yield Prediction.... 10312.8 Composition Control..104

13. DEFECTS AND GRADING .......................................................................................... 10713.1 Defects .............................................................................................................. 107

13.2 Grading ............................................................................................................. 109

14. SANITATION .............................................................................................................. 113

15. SOFT-RIPENED CHEESE ........................................................................................... 114A. Feta Cheese ......................................................................................................... 114B. Camembert Cheese .............................................................................................. 116C. Blue Cheese ......................................................................................................... 118

16. SEMI-HARDCHEESE -- WASHED ............................................................................ 121A. Brine Brick .......................................................................................................... 121

B. Colby ................................................................................................................... 123C. Gouda .................................................................................................................. 125D. Montasio (Fruilano) ............................................................................................ 126

17. FIRM TO HARD CHEESE: LOW TEMPERATURE: PROVOLONE, CHEDDAR128A. Provolone ............................................................................................................ 128B. Cheddar ............................................................................................................... 130

C. Romano ............................................................................................................... 132D. Swiss Cheese ....................................................................................................... 133

18. HEAT-ACID PRECIPITATED CHEESE ..................................................................... 136

A. Ricotta Cheese .................................................................................................... 136B. Queso Blanco (Frying Cheese) ............................................................................ 137C. Paneer (contributed by Sunil Radhakrishnan) ..................................................... 138

19. FRESH CHEESE ........................................................................................................... 140A. Cottage Cheese - Short Set (Emmons & Tuckey, 1967)..................................... 140


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B. Quark ................................................................................................................... 142C. Cream Cheese ..................................................................................................... 142

20. PROCESS CHEESE ....................................................................................................... 14520.1 Introduction ....................................................................................................... 145

20.2 Standards: Canadian Regulations ..................................................................... 14520.3 Ingredients ........................................................................................................ 14520.4 Process Systems ................................................................................................ 14720.5 Microbiology .................................................................................................... 14820.6 Calculations ...................................................................................................... 14820.7 Procedure .......................................................................................................... 149

21. LOW FAT CHEESE ....................................................................................................... 15221.1 Importance of Fat In Cheese ............................................................................. 15221.2 Current Status of Low-fat Cheese ..................................................................... 15221.3 Effects of Reduced-Fat On Cheese Composition ............................................. 152

21.4 Defects .............................................................................................................. 15221.5 Low-fat Cheddar Make Schedule ..................................................................... 153

22. CHEESE MAKING FROM ULTRAFILTERED MILK ............................................. 15522.1. Terms and Principles ..................................................................................... 15522.2 Benefits of UF in the Dairy Industry ............................................................... 15622.3 Properties of UF Milk Retentates .................................................................... 15622.4 Development of UF Applications in the Cheese Industry ............................... 157

23. CHEESE SUBSTITUTES ............................................................................................. 15923.1 Why: .................................................................................................................. 159

23.2 Threat or Opportunity ....................................................................................... 15923.3 Varieties currently available in US ................................................................... 15923.4 Types of Substitutes .......................................................................................... 15923.5 Cheddar Cheese Substitute ............................................................................... 160

24. WHEY PROCESSING ................................................................................................. 161


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LIST OF FIGURES

Figure 1.1 Flowchart of Cheese Making Process. ............................................................. 9

Figure 1.2. Effects of particular processing conditions on various cheese makingparameters. ......................................................................................................10

Figure 3.1. Culture Activity Test ...................................................................................... 30

Figure 4.1 Seasonal variation of fat, protein, lactose and protein:fat ratio in Ontarioproducer milk .................................................................................................. 43

Figure 4.2. The concept of pH............................................................................................44

Figure 5.1 Membrane concentration/fractionation ........................................................... 52

Figure 5.2 Microfiltration Flowchart ............................................................................... 52

Figure 7.1 Natural Fermentation of Raw Milk ................................................................. 72

Figure 8.1 Structural elements of milk. After Walstra and Jenness, 1984. DairyChemistry and Physics, Wiley & Sons, N.Y.................................................... 78

Figure 10.1 Cheddar cheese composition curing. (A) New Zealand standards forPremium and First Grade Cheddar cheese. (B) Typical ranges for highquality Canadian Cheddar. Note: pH measured between 3 and 14 days after

manufacture. .................................................................................................... 95

Figure 24.1 Whey processing & utilization ..................................................................... 162

Appendix 1. Some Common Unit Conversions

Appendix 2. Measurement of titratable acidity

Appendix 3. Measurement of pH


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LIST OF TABLES

Table 1.1 Some properties of cheese categorized according to type of coagulation and

procedures used for pH and moisture control. .................................................. 7

Table 1.2 Typical composition (% by weight) of some cheese varieties.......................... 8

Table 4.1 Typical gross composition (kg/100kg) of cow, dairy sheep and goat milk(Wong et al. 1988). ......................................................................................... 41

Table 4.2 The principal caseins and some properties of importance tocheese making ................................................................................................. 41

Table 4.3 The principal whey proteins and some properties of importance to

cheese making ................................................................................................. 42

Table 4.4 Typical fat and protein contents (kg/100 kg) for the milk of several breeds ofdairy cows (from various sources). ................................................................. 42

Table 6.1 Some cheese varieties with some characteristics, composition and suggestedratio of protein/fat in standardized milk. Fat and moisture levels for mostvarieties correspond to definitions given in the Canada Agricultural ProductsAct and Regulations, Section 28. .................................................................... 61

Table 7.1 Some lactic acid bacteria commonly used in cheese making. ....................... 71

Table 9.1 Record of Manufacture ................................................................................... 86

Table 9.2 Record of Quality Control .............................................................................. 87

Table 11.1 pH versus time profiles for several cheese varieties100

Table 12.1 Distribution of milk components during cheese making (% by weight) andpercent transfer from milk to cheese. ............................................................ 101

Table 12.2. Examples showing High, Medium and low levels of Cheddar moisture control

.105Table 12.3. Variation of composition in 290 kg blocks of stirred curd Cheddar cheese..105

Table 13.2 Cheese Judging Score Card...........................................................................112


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Table 20.1 Process cheese composition control: Example ............................................ 145

Table 20.2 Process cheese composition control ............................................................. 151

Table 22.1 Ultra filtration of whole milk: typical composition of concentrate and

permeate (polysulfone membrane, tubular configuration, operation at 50C(Glover, 1985). .............................................................................................. 158


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1.INTRODUCTION TO CHEESE MAKING

This section is both introduction and overview. As we study particular aspects of cheesemaking (the trees) you will find it useful to return here to refresh your view of the forest.

The Basic Process(Figure 1.1)

Cheese making can be described as the process of removing water, lactose and someminerals from milk to produce a concentrate of milk fat and protein. The essential ingredientsof cheese are milk, coagulating enzyme (rennet), bacterial cultures and salt. Rennet causesthe milk proteins to aggregate and ultimately transform fluid milk to a semi-firm gel. Whenthis gel is cut into small pieces (curds), the whey (mostly water and lactose) begins toseparate from the curds. Acid production by bacterial cultures is essential to aid expulsion ofwhey from the curd and largely determines the final cheese moisture, flavour and texture. Aflow chart showing the general operations of cheese making is in Figure 1.1. Figure 1.2illustrates, very diagrammatically and generally, some associations among some cheese

processing parameters and some cheese properties.

Cheese Families

The objectives of cheese making are: (1) To obtain the optimum cheese composition withrespect to moisture, acidity (pH), fat, protein and minerals (especially calcium); (2) Establishthe correct structure of the cheese at the microscopic level; and (3) Ripen to perfection.Objectives (1) and (2) are achieved by varying initial make procedures and it is then possibleto achieve objective (3). Most of these variations in initial make procedures are different

means to control the rate and extent of acid development, and the rate and extent of moisturerelease. Grouped according to texture and basic manufacturing procedures, seven cheese

families are described below and summarized in Table 1.1. This course is concerned mainlywith Families 4, 5, 6, and 7, but procedures for some acid coagulated fresh cheese (Family1) and heat-acid precipitated cheese (Family 3) are included for your information. Table 1.2contains composition data for some common cheese varieties.

Cheese families are described in this section under five headings, namely, varieties, type ofcoagulation, pH or acidity control, moisture control and curing. Other technologicalcharacteristics of cheese are listed at the end of this chapter.

Please note that while these categories are helpful to classify most cheese in technologicalgroups, the categories cannot be applied rigidly. For examples: (1) pasta filata varieties vary

widely in composition, manufacturing techniques and degree of ripening, so they dont fitany category well; and (2) The manufacture of Cheshire types is similar to Cheddar up to thepoint of draining, but after that their excessive acid development is similar to Feta.

Family 1. Predominantly Acid-coagulated Fresh Cheese

In North America, 'fresh cheese' normally refers to cheese produced by acid coagulation at

30 - 32C with little or no added rennet. Acid is normally produced via fermentation by lacticcultures but some fresh cheese may also be produced by direct acidification with glucono-


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delta-lactone. Cheese made for fresh consumption is also made via rennet coagulation(Family 2) and a procedure known as heat-acid precipitation (Family 3).

Varieties: Cottage, Quark and Cream

Coagulation: The distinguishing characteristic of these varieties is that coagulation isachieved by acidification to pH 4.6 - 4.8, with little or no coagulating enzyme. Acidificationis normally by lactic acid producing cultures. Most other American and European cheesevarieties also use lactic acid producing cultures, but gelation is induced by a coagulatingenzyme at pH 6.5 - 6.7, before much acid development has taken place.

pH Control: Depending on variety there may be little or no effort to control or adjust pH.

Cottage cheese, after cutting at pH 4.6 - 4.8, is cooked to 52C, which is sufficient toinactivate the culture and prevent further acid development. For some varieties, includingcottage, acidity is also reduced by washing the curd before salting.

Moisture Control: Curd moisture is reduced by syneresis during cooking but remains high,60 - 70%, in the finished cheese.

Curing: Fresh cheese as the name implies is consumed fresh and has a shelf life of only 2 - 3weeks.

Family 2. Predominantly Rennet-coagulated Fresh Cheese

In Latin American, Middle Eastern and some European countries, fresh rennet cheese isproduced with little or no culture. Without acid production by lactic acid bacteria, cheese pH

remains high and the resulting cheese does not melt when used in a stir fry or other cooked

recipes. For reasons of safety and quality, these varieties must be handled with extra attentionto sanitation and refrigeration.

Varieties: Queso Blanco, Queso Fresco, Italian fresh cheese, Halloumi

Coagulation: The distinguishing characteristic of rennet coagulated fresh cheese is that littleor no culture is used. Coagulation is, therefore, entirely by rennet at the natural pH of milk.

pH Control: The pH is determined by the amount of culture. If no culture is used, the pHremains in the range of 6.5-6.7. In some Queso Blanco varieties a small amount of culture isused to reduce the pH to about 5.8 which reduces the growth of both spoilage (increases

shelf life) and pathogenic (increases food safety) micro organisms. Further acidification isinhibited by cooling and salting. Too much acidification below pH


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Family 3. Heat-Acid Precipitated Cheese

Varieties: Ricotta (Italy), Channa and Paneer (India)

Coagulation: Coagulation is accomplished by direct acidification of heated milk. High heat

treatment of milk (temperatures greater than 75C) causes denaturation of the whey proteins.Subsequent acidification of the hot milk coagulates both casein and whey proteins so most ofthe milk protein is recovered in the cheese.

pH Control: The final acidity (pH) is determined by the amount of acid added. Final pH isnormally in the range of 5.3 - 5.8. Any organic acid can be used, but acetic, lactic and citricacids are most common.

Moisture control: Moisture can be reduced by holding the curd in the hot curd-whey mixture

after coagulation, and by draining and pressing procedures. Moisture is generally high (55 -80%) due to the high water holding capacity of whey proteins. High concentrations of wheyproteins also decrease cheese meltability and account for the excellent cooking properties ofheat-acid precipitated cheese.

Curing: Heat-acid precipitated varieties are normally consumed fresh. Exceptions are someaged heat-acid varieties such as Mizithra (Greek) a type of ricotta cheese which is cured,dried, and consumed as a grating cheese. It is also possible in some cases to hot pack heat-acid varieties to obtain extended shelf life.

Family 4. Soft-Ripened Cheese

Varieties: Feta, Camembert, Brie, Blue

Coagulation: Coagulation is primarily rennet (enzymatic) with three important differences

relative to cooked and pressed varieties (Families 5-7).

(1) The amount of lactic acid bacteria inoculum is relatively large and the ripening periodbefore renneting is extended. The result is that acidification has considerableinfluence on the development of curd structure during setting and demineralization ofthe curd is increased.

(2) Cutting is delayed (i.e., setting time increased) to further encourage acidification anddemineralization before cutting.

(3) Cutting is accomplished with large knives or the curd just broken up with paddles tominimize moisture and fines losses before filling the forms.

pH Control: The curd is placed in the forms while still sweet and let stand in a warm roomfor several hours. Acidification (i.e. conversion of lactose to lactic acid) continues until theaccumulation of lactic acid inhibits culture growth. Acid development is also influenced by


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the time and amount of salting. The pH is normally about 4.3 - 4.6 on the day followingmanufacture and in the case of Feta remains low during curing. The pH of mould ripenedvarieties increases during curing (i.e., acidity decreases), especially Camembert and Brie.

Moisture Control: Syneresis is induced by acid development after forming and by brine

salting. Moisture content is typically 45 - 60%.

Curing Time: 2 - 8 weeks.

Family 5. Semi-hard Washed Cheese

Varieties: This is the largest and most diverse group of cheese including Gouda, Edam,Colby, Brick, Montasio, Oka, Muenster and many others.

pH Control: The distinguishing feature of these varieties is the practice of washing to

remove lactose. Part or all of the whey is removed and replaced with water to leach lactose

from the curd. The objective is to limit the amount of lactose to a level which permitssufficient lactic acid development to produce a minimum pH of 5.0 - 5.2, but not enough toferment and produce cheese pH less than 5.0. Achievement of this minimum pH target isassisted by the buffer capacity of the milk proteins, especially the caseins, which have a pHbuffer maximum near pH 5.2.

Moisture Control: The amount of syneresis is controlled mainly by the temperature and timeof cooking and by the temperature of the wash water. Higher temperatures during cooking orwashing cause the curd to contract and expel moisture. Also, important are the rate of aciddevelopment and salting treatments. Washed curd varieties typically have moisture contentsof 40 - 50%. With few exceptions, such as part skim Mozzarella, production of a rennet

coagulated cheese with a moisture content of 40% or greater requires a washing treatment toremove lactose and lactic acid.

Curing: 2 weeks - 9 months.

Family 6. Firm/hard Cheese: Low temperature

Firm/hard cheeses (Families 6 and 7) are characterized by lower moisture, with someexceptions such as some Pasta Filata types. Lower moisture, generally less than 40%, permitsremoval of sufficient lactose by syneresis to avoid the necessity of washing. Low moisture isachieved by high temperature cooking (Family 7) or by controlled fermentation and curd

handling (Family 6).

Varieties: Cheddar types and some Pasta Filata types. Cheddar and Pasta Filata manufactureare similar in the early stages. Pasta filata varieties are distinct in that they are worked andstretched in hot water and brine salted. Cheddar types are salted before hooping and pressing.

pH Control: The distinguishing feature of these cheese is that acid development is largelycontrolled by the amount of syneresis during curd ripening (Cheddaring). During curdripening the drained curd is allowed to mat forming slabs of curd, which are piled and turned


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while the culture continues to grow and produce acidity. During this process, lactose isreduced via conversion to lactic acid and by acid induced syneresis. The process ofCheddaring is also applied in the manufacture of many pasta filata (stretched cheese)varieties, so I prefer to describe it generally as curd ripening. In recent times, curdripening techniques have been modified in various ways to facilitate automation.

The objective is to obtain a minimum pH of 5.0 - 5.3 within 1 - 3 days after manufacture.This is similar to the minimum pH target for semi-hard washed cheese varieties. Lactosecontent is substantially reduced by fermentation with associated moisture loss during curdripening, salting, and pressing in the case of Cheddar.

Moisture Control: Moisture is controlled by cooking temperature and time, stirring out afterdraining, curd ripening, amount of culture, and salting treatments. Typical moisture contentis 35 - 39% for Cheddar types and up to 52% for Pasta Filata types.

Curing: 1 - 36 months.

Family 7. Hard Cheese: High Temperature

Varieties: Romano, Parmesan, Swiss

pH Control: Type of culture, time-temperature profile during pressing until cooling, lactoseremoved by syneresis. There is little acid development before draining.

Moisture Control: Rapid syneresis induced by high renneting temperature and high cookingtemperature.

Curing: 1 - 36 months.

Other Technological Criteria

The cheese families described above provide a useful coat rack to help organize cheeseaccording to the initial manufacturing procedures that determine cheese composition and itsprimary microstructure. The following is a more comprehensive summary of technologicalparameters that determine cheese characteristics. Species: cow, goat, sheep, buffalo, yak, other Milk standardization

Fat and protein contents

Whey and milk blends Coagulation

Predominantly rennet gelsPredominantly acid gelsHeat-acid precipitates

Moisture controlCooking temperature and timeMesophilic versus thermophilic culturesAmount and acidifying properties of the culture


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Curd ripeningHeat treatment of the milk

Type of pH controlDirect acidification versus fermentationAmount and type of culture

Lactose removal: Washing (American, Dutch) High temperature syneresis (Swiss, Hard Italian) High acid syneresis (Feta, Cheshire) Curd ripening (Cheddar, Pasta Filata)

Extent of acid developmentLow acid (minimum pH > 5.8), Latin American fresh cheeseMedium acid (minimum pH 4.9 - 5.5), most European varietiesHigh acid (minimum pH < 4.9), Fresh cheese, soft ripened cheese

Salting proceduresVat salting before forming

Surface salting after formingImmersion in salt brine

Type and duration of ripeningFresh versus ripenedInterior, including blue veined cheeseInterior and surface ripened Bacterial/yeast smears White surface mould Mixed rinds

Type of rindRindlesswaxed, film wrapped, painted

Dry rind (cured at 85% relative humidity)Surface ripened (cured at 90-95% relative humidity)

Texture and bodyOpenings: mechanical, small holes, large holesFirmness

Melting propertiesNo melt: softening without flow (frying cheese)Stretching: Low melt and stretchable (Mozzarella)Fondue: Medium melt, medium elasticity (Raclette)High melt: flows readily with little stretchabiltiy (aged Cheddar)


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Table 1.2. Typical composition (% by weight) of some cheese varieties. Adapted from Hill (2007).

Type Cheese Moisture Pro-tein Fat

Total

CHO FDM Ash Ca

Acid

Coagulated

Cottage

Creamed cottage

QuarkCream

Neufchatel

79.8

79.0

72.053.7

62.2

17.3

12.5

18.07.5

10.0

0.42

4.5

8.034.9

23.4

1.8

2.7

3.02.7

2.9

2.1

21.4

28.575.4

62.0

0.7

1.4

1.2

1.5

0.03

0.06

0.300.08

0.07

Heat-Acid

Coagulated

Chhana

Frying cheese

Ricotta-3% fat milk

Ricotone-from whey &milk

53.0

55.0

72.2

82.5

17.0

19.7

11.2

11.3

25.0

20.4

12.7

0.5

2.0

3.0

3.0

1.5

53.2

44.8

45.7

2.9

Fresh

Rennet

Coagulated

Queso Blanco

Queso de Freir

Italian fresh cheese

52.0

52.4

49.0

23.0

23.0

28.0

20.0

19.5

16.0

42.0

41.0

31.4

Soft

Ripened

Camembert

FetaBlue

Gorgonzola

51.8

55.242.0

36.0

19.8

14.221.0

26.0

24.3

21.329.0

32.0

0.5

2.3

50.3

47.550.0

50.0

3.7

5.25.1

5.0

0.39

0.490.53

Semi-hard

Washed

Colby

Gouda

EdamFontina

Havarti-Danish

Munster

40.0

41.5

41.442.8

43.5

41.8

25.0

25.0

25.024.2

24.7

23.4

31.0

27.4

27.825.5

26.5

30.0

2.0

2.2

1.4

1.1

51.7

46.9

47.644.6

46.9

51.6

3.4

3.9

4.23.3

2.8

3.7

0.68

0.70

0.73

0.72

Hard

Cheese

Low-

Temp.

Cheddar

Manchego-Spain

Provolone

Mozzarella

36.7

37.9

40.9

54.1

24.9

28.1

25.6

19.4

33.1

26.9

26.6

21.6

1.3

2.1

2.2

52.4

45.2

45.1

47.1

3.9

3.6

4.7

2.6

0.72

0.76

0.52

HardCheese

High-

Temp.

ParmesanRomano

Swiss

Keflatyri-Greece

29.230.9

37.2

34.2

35.731.8

28.4

24.8

25.826.9

27.4

28.3

3.23.6

3.4

36.539.0

43.7

6.06.7

3.5

4.7

1.181.06

0.96


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Milk Analysis Fat and protein Inhibitors Somatic cells Total bacterial count Thermoduric bacteria Coliform bacteria

Figure 1.1. Flowchart of Cheese Making Processes. Adapted from Hill (2007).

Milk Preparation

Milk TreatmentsStandardize microflora

Cold storageMicrofiltration

BactofugationHeat treatment

Standardize compositionCream separationAdd skim milk,

condensed skim, milkprotein concentrates,caseins etc.

UltrafilterBlend with whey

(ricotta)Homogenize

Milk Additives

CaCl2 Nitrates Colors or de-colorants Ripening enzymes Lysozyme

Inoculation Primary lactic cultures Secondary cultures: Lacticculture adjuncts, moulds, smearcocktails etc.

Coagulation:

gelation or

precipitation

Chymosin (Calf rennet) Microbial enzymes Recombinant chymosin

Acidification of warm milk, director fermentation

Direct acidification of hot milk

Milk Receiving

Cutting or Breaking

Cooking and othercurd in w hey

treatments

Mesophilic cook up to 39Thermophilic cook up to 55C Wash with cold or warm water Press under the whey, Swiss

and Dutch types

Draining and postdraining curd

treatments

Dip curd and whey into formsCollect and form floating curd

(ricotta)

Curd ripening and milling(Cheddar, Pasta filata)

Vat salting (Cheddar, American)Hot water stretching and forming

(Pasta filata).Forming in hoops and pressingSurface or brine salting

Ripening

Dry rinds: oil, salt, brush, hang;8-15C, 85% RH

Surface ripened: inoculate, washand turn regularly; 12-15C, 95%RH

Rindless: wax, film wrap, 5-13C

Retail Packaging

Whey

Creaming bycentrifugation

Defatted Whey

Whey

Cream

Concentration and drying

Ultrafiltrationand drying

WheyPowder

Whey

ProteinConcentrate

Permeate


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Variable DrainingpH

MinimumpH

Ca Syneresis MNFS RennetRetention

Milk protein/fat UE UE UE

UE

Pasteurization intensity

CaCl2: amount added

Starter: amount added

Pre-ripening time1

Rennet: amount added UE UE

Renneting temperature NE UE UE UE

Curd size NE UE UE

Stirring intensity UE UE UE

Cooking intensity UE

Time until draining2 NE

Wash water amount NA

Wash water temp. NA UE

Time until salting3 NA NE UE

Salt: amount added NA UE

Pressure in the moulds NA UE UE

Figure 1.2. Effects of particular processing conditions, assuming other factors do notchange, on pH at draining, minimum pH occurring in the cheese during early stages of

curing, calcium retained in the cheese, the rate of syneresis, the moisture in the non-fatsubstance (MNFS) and the amount of calf rennet activity retained in the cheese. The figureis only intended to show trends that apply to most rennet coagulated cheese within normalranges of moisture content and percent fat in the dry matter. U = Unknown effect, but likelysmall. NE = no effect. NA = not applicable. Adapted from Hill (2007).1Time between adding culture and adding rennet2Total time between cutting and draining3Total time between draining and salting; applicable to vat salted varieties


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2. RECOMMENDED REFERENCES

Alfa-Laval. Dairy Handbook. Alfa-Laval, Food Engineering AB. P.O. Box 65, S-221 00Lund, Sweden. [Well illustrated text. Excellent introduction to dairy technology].

American Public Health Association, Standard Methods for the examination of dairy

products. 1015 Eighteenth St. NW, Washington, D.C.Battistotti, B., Bottazzi, V., Piccinardi, A. and Volpato, G. 1983. Cheese: A guide to the

world of cheese and Cheese making. Facts on File Publications, New York, NY.Berger, W., Klostermeyer, H., Merkenich, K. and Uhlmann, G. 2002. Processed Cheese

Manufacture, A JOHA guide. BK Ladenburg, Ladenburg..Carroll, R. and Carroll, R. 1982. Cheese making made easy. Storey Communications Inc.,

Ponnal, Vermont [Well illustrated manual for small and home cheese makingoperations].

Chandan, R. 1997. Dairy Based Ingredients. Amer. Assoc. Cereal Chemists,St. Paul, Minnesota.

Davis, J.G. 1965. Cheese. American Elsevier Publ. Co., New York.

Eck, A. and Gillis, J.-C., 2000. Cheesemaking from Science to Quality Assurance, LavoisierPublishing, Paris.

Fox, P.F., Guinee, T.P., Cogan, T.M., McSweeney, P.L.H. 2004. Cheese chemistry, physics,and microbiology. Third Edition. Volumes 1 and 2. Elsevier Academic Press, Sandiego,CA.

Hill, A.R. 2007. Physical factors affecting cheese flavour. In, Improving the flavour ofcheese. B. Weimer, Editor, Woodhead Publishing Ltd, Cambridge, England; pp. 252 278.

International Dairy Federation Special Issue No9301. Factors Affecting Yield of Cheese.

Kammerlehner, J. 2009. Cheese Technology.Publishing House Josef Kammerlehner D-85354 Freising, Germany.

Kosikowski, F.V. and Mistry, V.V. 1997. Cheese and Fermented Milk Foods, 3rd Edition,F.V. Kosikowski and Associates, Brooktondale, NY.

Law, B. 1999. Technology of cheese making. Sheffield Academic Press, Sheffield, UK.Lawrence, R.C., Heap, H.A. and Gilles, J. 1984. A controlled approach to cheese

technology. J. Dairy Sci. 67: 1632-1645.Lelivre, J., Freese, O.J. and Gilles, J. 1983. Prediction of Cheddar cheese yield. N.Z.J.

Dairy Sci. Technol. 18: 169-172.Masui, K.. and Yamada, T. 1966. French Cheeses: The Visual Guide to More than 350

Cheeses From Every Region of France. DK Publishing, New York.Morris, Margaret P., 2003. The Cheesemakers Manual. Glengarry Cheesemaking & Dairy

Supply, Winchester, ON, Canada [especially useful for small scale cheese makingt].


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Official Methods of Analysis of the Association of Official Agricultural Chemists, P.O. Box540, Benjamin Franklin Station, Washington, D.C.

Pfizer Cheese Monographs. 1963. C. Pfizer and Co., New York [These monographs are welldated, but are still useful, especially Volumes 3 and 5.1. Italian Cheese Varieties

2. American Cheese Varieties3. Cottage Cheese and Other Cultured Milk Products4. Ripened Semi-soft Cheeses5. Swiss Cheese Varieties6. Lactic Starter Culture Technology

Price, W.V. and Bush, M.G. 1974. The process cheese industry in the United States: Areview. I. Industrial growth and problems. J. Milk and Food Technology37: 135-152.II. Research and Development. Ibid 37: 179-198.

Robinson, R.K., Editor. 1990. Dairy Microbiology, Volumes 1 and 2. Elsevier AppliedScience, NY.

Scott, R., Robinson, R.K. and Wilbey, R.A. 1998. Cheese making Practice. 3rd Edition.

Applied Science. Publ. Ltd., London.Gunasekaran, S., and Ak. M.M. 2003. Cheese rheology and texture. CRC Press, Boca Raton,

FL.Troller, J.A. 1993. Sanitation in Food Processing. 2nd Edition. Academic Press. New

York.Walstra, P., Geurts, T.J., Noomen, A., Jellema, A. and van Boekel, M.A.. 1999. Dairy

Technology. Marcel Dekker Inc. New York, NY.Weimer, B.C. Editor. 2007. Improving the flavour of cheese. CRC Press, New York.Wong, N.P., Jenness, R., Keeney, M. and Marth, E.H. 1988. Fundamentals of Dairy

Chemistry. Van Nostrand Reinhold Company, New York, NY.

Web Sites

Agriculture Canada, http://res.agr.ca/CDRN/home.htmlAmerican Cheese Society http://www.cheesesociety.org/index.cfmCentre For Dairy Research, Madison, WI. http://www.cdr.wisc.edu/Cheese basics, http://www.efr.hw.ac.uk/SDA/cheese2.htmCanadian Dairy Commission, http://www.milkingredients.ca/dcp/index_e.aspDanlac Forum, http://www.danlac.com/forum/Food Science University of Guelph: http://www.foodsci.uoguelph.ca/cheese/welcom.htmGlengarry Cheese Supplies, http://glengarrycheesemaking.on.ca/Ontario Cheese Society http://www.ontariocheese.org/index.php

Specialist Cheese Makers Association http://www.specialistcheesemakers.co.uk/Vermont Institute for Artisan Cheese http://www.uvm.edu/~viac/


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3. PROCESS AND QUALITY CONTROL PROCEDURES

3.1. Introduction

Chemical and microbiological analyses of cheese milk, finished cheese and cheese whey are

required to maintain efficient operations and to ensure food safety and quality. This chapterdescribes some analytical procedures relevant to cheese making operations, but it is notintended to be a comprehensive process and quality control manual. The following generalcomments are intended to orient the reader to the general types of analyses required in cheeseoperations. Subsequent chapters will identify process and quality control requirements in thecontext of each step in the cheese making process.

Milk Analysis

Milk composition analyses should include both fat and protein, determined by infrared milkanalysers. Note that casein content rather than total protein content is the critical parameter

with respect to cheese yield. Cheese makers are, therefore, advised to regularly monitor therelative amounts of casein, whey proteins and non-protein nitrogen in their milk. Monthly orbimonthly analysis of protein distribution by Rowland fractionation is sufficient to monitorseasonal trends. Alternatively, an indication of casein and whey protein distribution can beobtained by comparing protein concentration in cheese whey to the protein concentration inthe initial milk. This has the advantage that infra red milk analysers can be calibrated tomeasure protein in cheese whey. See Chapters 6 and 12 for details on standardization of milkcomposition and the importance of casein to cheese yield.

Quality measurements of cheese milk should include total counts (and/or psychrotrophiccounts), tests for inhibitors and somatic cell counts. Depending on the types of controls in

place at the producer level, cheese makers may need to monitor bacteria counts, inhibitorsand somatic cell counts of individual producer milks.

Cheese Analysis

Cheese composition analyses should include fat (by Babcock, Mojonnier, or near infra redprocedures), moisture, salt and pH. Cheese pH should be measured at the time ofmanufacture, 4 - 7 days after manufacture and periodically during curing. Other compositionparameters should be determined several days after manufacture to permit time forequilibration of soluble components. Salt in particular, requires time to become evenlydistributed throughout the cheese and in the case of brine or surface ripened cheese, uniform

salt distribution may never be achieved. For Cheddar cheese and other vat salted cheese,representative samples for accurate determination of salt content can be usually be obtainedas early as seven days after manufacture.

With respect to process and quality control, the pH profile during manufacture and curingis vital. pH profile is a term I use to describe the set of pH values at critical process controlpoints in the cheese making process. Other critical process control parameters are the ratio ofsalt to moisture (SM), the moisture in the non-fat substance (MNFS), and the fat in the drymatter (FDM). These ratios are normally reported as percentages and calculated as follows.


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Note that percent total solids is 100 minus percent cheese moisture

Routine cheese microbial analyses should include yeasts and moulds, total coliforms andstaphylococci. For raw milk cheese, all vats must be tested for the presence of Salmonella,Staphylococci, Listeriaand enteropathogenicE. coli. Cheese made from heat treated but notpasteurized milk must also be considered higher risk and should be monitored on a regularbasis for the presence of common pathogens. Microbial analyses should be performed at the

time of manufacture and after curing. Cheese whey should be monitored for the presence ofbacteriophage specific for the culture currently in use.

Analytical Quality Control

A simple but vital truism is that inaccurate analytical results are of less value than noanalytical results. Important causes of low yield efficiency and poor process control areinsufficient and inaccurate chemical and microbial analyses. Effective control of quality andplant efficiency requires effective quality control of analytical procedures. Smaller cheesemanufacturers generally find its more economical and reliable to have most analysesperformed by an outside laboratory. But, whether the analyses are performed in house or by

an outside laboratory, be certain that your laboratory services are accurate and reliable. InCanada, dairy laboratory reliability can be assured by certification with the CanadianLaboratory Accreditation Programme (LAP), Ottawa, (613) 247-1395. The LAP is able toprovide ongoing certification for both milk analysis (composition and quality) and cheese

composition analysis.I strongly recommend that cheese makers use LAP certified

testing, whether lab services are provided from inside or outside the company(yes, Iknow the manager of the LAP program, and no, he doesnt pay me to recommend it).

Some analytical procedures are detailed in subsequent sections. The reader is also referredto:1. Standard Methods for the examination of dairy products. American Public Health

Association, 1015 Eighteenth St. NW, Washington, D.C.2. Official Methods of Analysis of the Association of Official Agricultural Chemists,

P.O. Box 540, Benjamin Franklin Station, Washington, D.C.

3.2. Cheese Sampling

solidstotalcheese%

100xfatcheese%=FDM

fatcheese%-100

100xmoisturecheese%

=MNFS

moisturecheese%

100xsaltcheese%=S/M


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Chemical Analysis

Depending on the size and shape, firm to hard cheese should be sampled using a cheese trier(at least 100 g sample) or by taking a sector sample. Soft cheese can be blended for samplingor sector sampled depending on its texture. Cheese samples are stored in opaque air tight

containers and fragmented using a grater or other device before analysis. It is important togrind and mix the sample well before sub-sampling for analysis.

If the analytical procedure requires less than a 1 gm sample it is desirable to prepare a liquidcheese homogenate and a sub-sample from the homogenate. An homogenate suitable formost purposes can be prepared as follows. Weigh 40 g cheese into a blender container Add about 100 g of 7% sodium citrate solution Blend until homogenous using a high speed blender. Rinse blender shaft into container and make up to final weight of about 200g.

Note that cheese is notorious for inhomogeneous composition. Brine salted cheese havepronounced salt and moisture gradients, namely, higher salt and lower moisture near thesurface. Large blocks or wheels of pressed cheese will have moisture and pH gradients,namely, increasing moisture and decreasing pH towards the interior. In addition to moistureand salt gradients, surface ripened cheese also has pH gradients, namely, pH increases at thesurface during curing. These difficulties greatly complicate the matter of obtaining accuratecomposition and mass balance (yield) data. A useful approach to improve yield control oflarge blocks is to set aside small blocks (eg., 20 kg blocks of Cheddar) for early compositionand quality testing, and subsequently, conduct representative sampling of the large blocks(eg., 240 kg blocks of Cheddar) during the cut/wrap process.

Microbial Analysis

Obtain samples as described above for chemical analysis. Triers or knives used for samplingmust be flame or alcohol sterilized. Samples should be stored in sterile bags such as Whirl

Pack bags, stored at 0-4C and analysed within 24 hours.

Equipment Balance, 1,000 g capacity Blender Blender container autoclaved or sanitized with 200 ppm chlorine solution for 5 min.

Procedure1. Break the cheese into small pieces while still in the bag. Use a pestle or similar

device if necessary.

2. Heat dilution blanks of sterile aqueous 2% sodium citrate to 40C. Transfer 30 g ofcheese to sterile blender container, add 270 ml diluent and mix for 2 min. at speed

sufficient to emulsify the cheese properly. If temperature exceeds 40C duringblending, use a shorter mixing time or decrease initial temperature of citrate solution.This 1:10 dilution should be plated or further diluted immediately.


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3. Further dilutions can be prepared as required. Pipette 11 ml of the 10-1

dilution of thehomogenate, avoiding foam, into 99 ml dilution blank (0.1% peptone) or 10 ml into90 ml dilution blank. Shake this and all subsequent dilutions vigorously 25 times in aone foot arc. Prepare 10

-1, 10

-2, and 10

-3dilutions.

3.3. Total Solids

Oven Method

1. Pre-dry aluminum dishes (105C, 1 h) and weigh to the nearest 0.1 mg on ananalytical balance.

2. Weigh quickly 3-5 g of fragmented cheese into the aluminum dish. The weight ofsample is the total weight minus the weight of the dish from Step 1.

3. Dry to constant weight (about 16 h) at 105C. To check for constant weight: weigh atleast two samples, return both samples to the oven for an additional 20 minutes, andre-weigh. The difference between the weights before and after the additional drying

period should be less than 1 mg.4. Cool in desiccator and determine total dry weight. Sample dry weight is the total dry

weight less the weight of the dish determined in Step 1.5. Report total solids and moisture contents on weight percent basis as follows:

Note:Several rapid moisture tests based on infrared or microwave drying are available.Check with your laboratory equipment supplier.

Application notes

Accurate cheese moisture analysis is critical to composition and yield control. Rapidmoisture tests (e.g., microwave moisture oven) can be used to obtain early feed back (e.g.,cheese moisture immediately after pressing) information to help with process control.

3.4. Titratable Acidity

Principle

See discussion of pH and acidity in Section 3.5.

Apparatus and Reagents

1. An acidimeter equipped with a burette graduated in units of 0.1 ml up to 10 ml, andsome means of filling the same without undue exposure of the solution to the carbondioxide of the atmosphere.

SolidsTotal%-100=Moisture%(b)

wetweight

dryweight=SolidsTotal%(a)


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2. N/10 sodium hydroxide solution.3. A dropping bottle containing a 1% alcoholic phenolphthalein solution.4. White cup, glass stirring rod, 17.6 ml pipette (or 8.8 or 9.0 ml pipette)5. For cream, Torsion balance and 9 g weight.

Method

1. Mix sample thoroughly by pouring it from one container to another. The temperature

of the sample should be near 20C.2. Pipette 17.6 ml of milk or cream into a white cup. Note: 8.8 ml pipettes may also be

used but are no longer as readily available as 17.6 ml pipettes. Readily available 9 mlpipettes may also be used.

3. Add six drops of phenolphthalein indicator solution to milk, or 10 drops if theproduct is cream.

4. Titrate the sample with the N/10 sodium hydroxide solution (0.1 Normal NaOH)while stirring the sample with the glass rod. Look for the appearance of a faint pink

colour, which signals the endpoint. Add another drop or half a drop of NaOH if thepink colour does not persist for 30 s.

5. Record the number of ml of NaOH used to reach the endpoint. This value is calledthe 'titre'. Titratable acidity reported as percent lactic acid is dependent on the volumeof sample.

For the 8.8 ml pipette, % Lactic acid = titreFor the 17.6 ml pipette, % Lactic acid = 0.5 x titreFor the 9.0 ml pipette, % Lactic acid = 0.98 x titre.

Note that there is practically no lactic acid in fresh milk, but it is a North Americanconvention to report TA in terms of % lactic acid.

Application notes

As described in the next section, both titratable acidity (TA) and pH are measures of acidity.For most process control purposes, pH is a more useful measurement. Many cheese makers,however, still use TA to monitor initial acid development (that is to check for cultureactivity) during the first hour after adding the culture. For this purpose, TA is a more reliableindicator because relative to pH measurement, it is more sensitive to small changes in milkacidity.

When using TA to monitor initial culture activity, note that:

You are looking for a measurable increase in TA to confirm that the culture is active.For example, if the initial TA taken immediately after the culture was added is0.180% lactic acid, and the TA after one hour of ripening is 0.190 % lactic acid, thechange in TA is 0.010%.

Different people will interpret the coloured endpoint differently, so it is importantthat the same person takes both the initial and final TA measurements.

The principal divisions on most acidimeters are units of 0.1% lactic acid withsubdivisions corresponding to 0.01% lactic acid. It is possible to interpolate betweenthe subdivisions to obtain readings to the third decimal place. In practice, for most


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analysts, the sensitivity is about 0.005% lactic acid. That means it is possible toreliably measure a change in TA of 0.005% lactic acid. So if the TA increase isgreater than 0.005% you can conclude that the culture is active. In most cases TAincreases in the range of 0.005% to 0.010% are obtained after 30 - 60 minutes ofripening (that is, 30 - 60 minutes after adding the lactic cultures).

If you are using a bulk culture, it is critical to take the initial TA reading after theculture is added, because the culture is acidic and will increase the initial reading.

3.5. pH

Concepts of Acidity and pH

All aqueous systems (including the water in you and in cheese) obey the followingrelationship between the concentration of hydrogen ions (H+) and hydroxyl ions (OH-). Note,the square brackets indicate concentration in moles per litre. A mole is 6 x 1023molecules,

that is, the numeral six with 23 zeros after it.

[H+] x [OH

-] = 10

-14

Because the actual concentrations in moles per litre are small, it is customary to express thevalues as exponents. For example, if we know that the concentration of hydrogen ions [H

+] in

a sample of milk is 0.000001 moles/L which is equivalent to 10-6

moles/L, we can calculatethe concentration of hydroxyl ions as 10

-14/10

-6= 10

-8moles/L which is the same as

0.00000001 moles/L.

If [H+] = [OH

-] the solution is neutral with respect to acidity.

If [H+] > [OH

-] the solution is acidic.

If [H+] < [OH

-] the solution is basic or alkaline.

Chemicals which contribute H+

or absorb OH-

are acids, while bases contribute OH-

or absorb H+.

The concept of pH evolved as a short hand method to express acidity. We have already seenthat a hydrogen ion concentration of 0.000001 moles/L can be expressed as [1 x10-6], anexpression which defines both the unit of measurement (the square bracket meansconcentration in moles/L) and the numerical value. The concept of pH is a furtherabbreviation which expresses the concentration of hydrogen ions as the negative log of thehydrogen ion concentration in units of moles/L. This sounds complex but is quite easy toapply. For example, the log10of hydrogen ion concentration of [10

-6] is equal to -6. The finalstep is to take the negative of the log (-1 x -6 = 6). So, 0.0000001 moles/L = [10-6] moles/L

= pH 6. From the relationship expressed in Equation 3, if the concentration of either OH-orH+is known, it is always possible to calculate the concentration of the other. So, if the pH ofa solution is 6, the pOH is 14 - 6 = 8. Because this relationship is understood (well, OK, at

least understood by the chemists!), the convention is to only report pH. Note, that because

the negative sign was dropped for convenience, decreasing pH values mean increasing

acidity, or increasing concentration of H+ions. So, although both TA and pH are

measures of acidity, pH decreases with increasing acidity.

All of this can be summarized by a description of the pH scale. The pH scale for most


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practical purposes is from 1 to 14, although pH less than one is theoretically and practicallypossible.

pH 7.0 is neutral acidity [H+] = [OH

-]

pH < 7.0 = acid condition [H+] > [OH

-]

pH > 7.0 = alkaline or basic condition [H+] < [OH

-]

pH Versus Titratable Acidity

TA and pH are both measures of acidity but, for most purposes, pH is a better process controltool because the pH probe measures only those H+ that are free in solution and not associatedwith salts or proteins. This is important because it is the free H+that modify proteinfunctionality and contribute sour taste. The pH, rather than titratable acidity, is also the best

indicator of the preservation and safety effects of acidity. It must be emphasized, that the

most important factor available to the cheese maker to control spoilage and pathogenic

organisms is pH. Also, the pH history of the milk and cheese or whey is important troubleshooting information. Cheese moisture, mineral content, texture and flavour development

are all influenced directly by the activity of free hydrogen ions (i.e. pH).

Titratable acidity (TA) measures all titratable H+up to the phenolphthalein end point (pH8.5) and, therefore, it varies with changes in milk composition and properties. During cheesemanufacture, the pH gives a true indication of acid development during the entire process, sothe optimum pH at each step is independent of other variables such as milk protein content.However, the optimum TA at each step in cheese making will vary with initial milkcomposition, milk heat treatments, and the procedure used to standardize milk composition.

An illustration of the difference between TA and pH is the effect of cutting. Up to the time ofcutting, TA of the milk increases with the development of acidity by the culture. After

cutting, the TA of the whey is much lower. This does not mean that acid developmentstopped. It simply means that titratable H+associated with the milk proteins are no longerpresent in the whey. This leads to the concept of buffer capacity, which is an important

principle in cheese making. The effect of protein removal on the TA of whey is related to theability of protein to buffer the milk against changes in pH. That same buffer property is thereason it helps to take acidic medication, like aspirin, with milk.

Buffer capacity can be described as the ability of an aqueous system, such as milk, to resistchanges in pH with addition of acids (added H

+) or bases (added OH

-). Specifically, buffer

capacity is the amount of acid or base required to induce a unit change in pH. For example, asmall addition of acid to distilled water will cause a large reduction in pH. The same amount

of acid would have a small effect on the pH of milk, because milk proteins and saltsneutralize the acidity.

The two most important buffer components of milk are caseins (buffer maximum near pH 4.6- 5.2) and phosphate (buffer maximum near pH 7.0). The casein buffer maximum near pH 5.0is extremely important to cheese manufacture because the optimum pH for most cheese is inthe range of 5.0 - 5.2. As the pH of cheese is reduced towards pH 5.0 by lactic acidfermentation, the buffer capacity is increasing (i.e., each incremental decrease in pH requiresmore lactic acid). The effect is to give the cheese maker considerable room for variation in


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the rate and amount of acid production. Without milk's built in buffers it would be difficultto produce cheese in the optimum pH range.

Another way to illustrate the difference between TA and pH is to consider typical ranges ofpH and TA for normal milk. TA is a measure of the total buffer capacity of milk for the pH

range between the pH of milk and the phenolphthalein end point (about pH 8.5). The pH ofmilk at 25C normally varies within a relatively narrow range of 6.6 to 6.8. The normalrange for titratable acidity of herd milks is 0.12 - 0.20% lactic acid.

pH Measurement

The pH of cheese milk, whey, and soft cheese can be measured directly. Firm and hardcheese must be fragmented before analysis. Always measure cheese pH in duplicate and usecare in handling the electrode. Place the fragmented cheese in a 30 ml vial or small beakerand gently push the electrode into the cheese....too much haste is likely to break the electrodeon the bottom of the beaker. To ensure good contact, press the cheese around the electrode

with your fingers. There is no need to rinse the electrode between cheese samples.However, if the electrode is stored in buffer it should be rinsed with distilled water beforemeasuring cheese pH. Always store the electrode in pH 4 buffer or as directed by themanufacturer. Do not rub the electrode. The electrode should be washed with detergent andrinsed with acetone occasionally to remove fat and protein deposits.

3.6. Babcock Methods for Milk Fat

Apparatus and Materials

1. Babcock centrifuge.

2. Water bath at 55C.3. Torsion balance, 9 and 18 g weights.4. Babcock shaker.5. Glassware: 8% milk bottles, 50% cream bottles, 50% Paley bottles, 17.5 ml cylinders,

17.6 ml pipette.6. Reagents: - Babcock sulphuric acid (Sp. Gr. 1.82-1.83), N-butyl alcohol, glymol

Milk

1. Temper sample to 20C and mix by pouring gently from original container to abeaker of similar capacity 4-5 times.

2. Transfer 17.6 ml (18.0g) of milk to 8% bottle with 17.6 ml pipette. Allow pipette todrain then blow out the remaining drop into the bottle.

3. Add 17.5 ml sulphuric acid (Sp. Gr. 1.82-1.83) in at least three increments usingspecial cylinder. Rotate bottle between thumb and fingers while adding acid to washmilk from neck. Mix thoroughly 2 min. after each addition of acid by moving thebulb of the bottle in rapid circular motion. Final colour of mixture should bechocolate brown.

4. Centrifuge 5 min.


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5. Add distilled water at 60C to bring contents to within one-quarter inch of base of

neck. Do not mix.6. Centrifuge 2 min.

7. Add water at 60C to float fat into neck of bottle. Top meniscus should be abouteven with the top of the graduated portion. Do not mix.

8. Centrifuge 1 min.9. Temper bottles in water bath at 55C for 5 min.10. Measure length of fat column with dividers from top of upper meniscus to bottom of

lower meniscus. Place one divider point at zero mark and read percentage fat byweight directly where other point touches the scale.

Cream and Cheese

1. Temper cream sample to 20C and mix. Grind cheese to small particles.2. Weigh 9 g of cream into 50% cream bottle and add 9 ml of distilled water at 20C.

Weigh 9 g of cheese into a 50% Paley bottle and add 10 ml of distilled water at 60C.

3. Add 17.5 ml sulphuric acid in at least three increments. Mix until colour is uniformchocolate brown and all cheese particles are dissolved.


5. Add distilled water at 60C to bring contents to within one-quarter inch of base ofneck. Do not mix.


7. Add water at 60C to float fat into neck of bottle. Do not mix.8. Centrifuge 1 min.

9. Temper bottles in water bat at 55C, for 5 min.10. Place 4-5 drops glymol on the fat column letting these run down the side of the neck.

Measure the length of the fat column from the demarcation between fat and glymolto the bottom of the lower meniscus.

11. Report fat in percent by weight.

Skim milk, Buttermilk, Whey

1. Temper sample to 20C and mix gently.2. Transfer 2 ml N-butyl alcohol and then a 9 ml sample to an 18 g double neck bottle.

Mix thoroughly with a circular motion.

3. Add 9 ml of Babcock sulphuric acid for skim milk or buttermilk, 7 ml for whey.4. Centrifuge 6 min. Place bottles in the centrifuge cup with the small neck facing the

outside.5. Add water at 60C to bring contents 1 cm from the base of the neck. Do not mix.Centrifuge 2 min.

6. Temper bottles in water bath at 55C for 5 min.7. Place a finger over the large neck and press down until the lower meniscus of fat in

the small neck corresponds to a major division.


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3.7 Cheese Salt

Cheese salt determination is traditionally done using the Volhard procedure (OfficialMethods of Analysis of the Association of Official Agricultural Chemists, P.O. Box 540,Benjamin Franklin Station, Washington, D.C.). Other methods which have proven to give

accurate results are: Automatic Chloride Titraters operate on the principle of coulometric silver ion

generation to titrate chloride ions in the sample. When all chloride ions are titratedfree silver ions cause a conductivity change which signals the end of titration.

Quantab Chloride Titrater depends on the reaction of chloride ions with silverdichromate, which is brown, to form silver chloride chromate ion and silver chloridewhich is white. The reaction takes place on a calibrated strip which permits directestimation of chloride content.

3.8. Culture Activity Test (see also Figure 1.1 at the end of Section 3)

Purpose

This simple test is useful to ensure that cheese cultures have adequate activity beforeinoculating the cheese vat. For most cheese a general rule of thumb is that the activity andamount of inoculum should be sufficient to produce a titratable acidity of about 0.34% lactic

acid, in 10% reconstituted skim milk, after 4 h of incubation at 37C. The test is also usefulto compare types of cultures prepared under different conditions. For these purposes a pHversus time chart is quite useful (See Figure 3.1). A further application is to check sensitivityof the culture to bacteriophage in the plant (See Section 3.9 and Figure 3.1).

Procedure

1. Mix 10 g of low-heat, antibiotic-free skim-milk powder in 90 ml of distilled water ina 100 ml Erlenmeyer flask.

2. Sterilize at 15 lb pressure (1.05 kPa.) for 10 min.

3. Cool to 37C.4. Inoculate with 3.0 ml starter or other amount as appropriate. Rinse pipette twice by

drawing the sterile milk into it.

5. Incubate at 37C for at least 4 h. Longer if desired for pH versus time profile.6. Check pH at 30 min. intervals.7. Titrate 17.6 ml with N/10 sodium hydroxide (NaOH) using 1 ml phenolphthalein.

Divide the required ml of NaOH by 2 to obtain titratable acidity in units of percent

lactic acid.8. Record starter activity as follows:

Active, over 0.34%Slow 0.26 to 0.30%


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3.9. Detection of Bacteriophage (see also Figure 1.1 at the end of Section 3)

The following tests are based on the principle that bacteriophage specific to the culture in usewill be present in high numbers in the cheese whey. Therefore, by monitoring whey for thepresence of phage "a dead vat" on subsequent days can be avoided.

Culture Activity Test

The culture activity test described above can be used to detect the presence of phage incheese whey. Prepare 300 ml of reconstituted skim milk and place 99 ml in each of threebeakers. Add 1 ml of whey to Beaker 1 (100 x dilution), then transfer 1 ml from Beaker 1 toBeaker 2 (10,000 x dilution) and finally, transfer 1 ml from Beaker 3 to Beaker 4 to make a 1million times dilution. Add culture and monitor pH as described in section 3.8.

Bromocresol Purple (BCP) Phage Inhibition Test

This test is quite simple to perform, and produces more accurate results than the cultureactivity test.1. Prepare Materials

- BCP stock solution (1 g/100 ml water)

- Test tubes containing 9.9 ml sterile BCP-milk (5 ml BCP stock solution/litremilk)

- 30-320 water bath or heating block- 1 ml graduated pipettes- Membrane filter (0.45 u) -- optional- Disposable syringe -- optional- Clinical centrifuge -- optional

- Whey sample for phage testing- Freshly grown culture, frozen syringe, or frozen can of each strain

2. Add Whey to BCP Milk and Make DilutionsTransfer 0.1 ml of fresh (or filter-sterilized) whey to the first dilution tube (10

-2) and

mix well. Transfer 0.1 ml from the first to the second dilution tube and mix well.Repeat process for the third dilution tube. (If unfiltered whey is used, a control tubecontaining BCP milk and whey only, must be prepared. This control tube tests for thepresence of active culture in the whey that could mask phage inhibition of a strain.)Whey samples should be refrigerated immediately after collection and held cold untiltested for phage.

3. Add Culture to Control and Whey Dilution Tubes

Cheese culture (0.2 ml) is added to whey dilution tubes and to a control tube for eachstrain. If you are using direct-to-the-vat culture, dilute 1 ml of culture in 9 ml of milkand then add 0.2 ml of the mixture to the dilution tubes. The control tube containsonly BCP milk and culture---NO whey. The control tube serves to show starter straininhibition by colour comparison with the other tubes.

4. Incubate Tubes and Interpret Results. Incubate both control and dilution tubes for 6

hours at 30-32C. Compare the colour of the whey dilution tubes to that of thecontrol tube. Ignore coagulation. An uninhibited culture will produce sufficient acidto turn the BCP dye from blue to yellow. Strains should be removed from the culture


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blend when full inhibition persists at the 10-6

dilution level. The following systemshould be used to record phage inhibition:0 = No inhibition at any dilution1 = Partial inhibition at 10

-2dilution

2 = Full inhibition at 10-2

dilution

3 = Partial inhibition at 10-4

dilution4 = Full inhibition at 10

-4dilution

5 = Partial inhibition at 10-6

dilution6 = Full inhibition at 10-6dilution

3.10. Inhibitory Substances1

Regulations

Most jurisdictions have regulations concerning the testing methods and limits of certain

antibiotics in raw milk. The Milk Act of Ontario, Regulation 761, Section 52, Subsection,

states:"The milk of every producer shall be tested at least once a month for the presenceof an inhibitor by an official method."

An official method is described in a separate inhibitor policy document which states:The minimum sensitivity of an official method to test for the presence of an inhibitorunder section 52 of Regulation 761 shall be:a) 0.01 international units of penicillin per millilitre of milk by the Standard Disc

Assay (Bacillus stearothermophilus) procedure.b) 10 parts per billion sulfamethazine by the High Performance Liquid

Chromatography (modified Smedley and Weber) procedure.

A concentration of 0.01 international units of penicillin per millilitre of milk is equivalent to6 parts per billion (ppb). Note: 1 ppb is equivalent to a single penny in $10 million or onesecond in 32 years.

Detection Methods

It is beyond the scope of this manual to discuss any specific methods in detail. What followsare brief descriptions of five types of inhibitor tests that are currently used in the dairyindustry. For each category one or more brand name tests are listed to indicate possiblechoices. For cheese manufactures seeking assistance with inhibitor testing, many private labs

provide suitable services. Note, (1) none of the methods, especially rapid methods are able todetect all of the antibiotics that are used in dairy production and (2) Both false negative andfalse positive results occur. So, if you are using rapid tests in your operation it is importantthat the results are confirmed. In Ontario, a wide range of expertise and methodologies are

1This section is adapted from two reports prepared by: Mark Mitchell (1995), Ontario Ministry of

Agriculture, Food and Rural Affairs, Guelph, Ontario


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available from Laboratory Services Division, University of Guelph.

Growth Inhibition Assays

Examples: Delvotest P, Delvotest SP, BR test, BR-AS test, Charm Farm, and the Disk

Assay

This test format involves a standard culture of a test organism in an agar growth media,usuallyBacillus stearothermophilus, which is inoculated with a milk sample and incubatedfor periods of up to several hours. If the milk contains one or more inhibitory substances, thegrowth of the organism will be reduced or eliminated. The presence of an inhibitorysubstance is indicated by zones of inhibition or a change in colour of the media (pH andredox indicators).

The major disadvantages of these tests are that they are not very specific for identificationpurposes, have limited sensitivities to many antibiotics and take a long time before results are

available. Growth inhibition tests are only able to classify residues into either the -lactam(penicillin like antibiotics) or other than -lactam antibiotic families. A further concern isthat growth inhibition tests are subject to the effects of natural inhibitors such as lysozyme,lactoferrin, and defensins, which can be found in high levels in mastitic milk and may givefalse positive test results, particularly when used at the cow level. These effects can be

minimized by heating individual cow samples at 82C for 2-3 minutes in a microwave ovenor water bath before testing to destroy natural inhibitors and allow antibiotics, which aremore heat stable, to remain.

The advantages of these tests are that they are cheap, easy to perform and have a very broaddetection range.

Enzymatic Colorimetric Assays

Example: Penzyme Test for -lactams

The Penzyme test is based on the inactivation of an enzyme by -lactam antibiotics. Theenzyme (DD-carboxypeptidase or penicillin binding protein) is present in all bacteria and isinvolved in the synthesis of the bacterial cell wall. -lactam antibiotics will bind specificallywith this enzyme and block its activity, thus preventing the formation of the bacterial cellwall. This enzyme has been freeze dried and placed in sealed vials to which the milk sampleis added. After addition of 0.2 ml (200 L) of milk sample to the vial the sample is incubated

for 5 minutes at 47C. During this time any -lactams present in the milk bind to the enzymeand inactivate a certain amount depending on the concentration present.

Reagent tablets specific for the enzyme (D-alanine peptide and D-amino acid oxidase) are

then added to the milk sample and the sample is incubated at 47C for 15 minutes. Duringincubation any remaining active enzyme will react with the reagent added. The end productof the substrate and enzyme reaction (pyruvic acid and hydrogen peroxide) is measured by aredox colour indicator and the final colour is compared to a colour chart provided with the


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kit.

An orange colour (reduced) indicates a negative test result.

A yellow colour (oxidized) indicates a positive test result.

Microbial Receptor Assays:

Example: Charm II

This test uses bacterial cells (Bacillus stearothermophilus), which contain natural receptorsites on or within the cells for antibiotics, and radio labelled (C

14or H

3) antibiotics. Milk

sample is added to a freeze dried pellet of bacterial cells (binding reagent) in a test tube andthe sample is mixed and incubated. During incubation any antibiotic present in the milk willbind to its specific receptor site. Radio labelled antibiotic (tracer reagent) is then added andthe sample is mixed and incubated. Unbound receptor sites on the bacterial cell will be boundby the radio labelled antibiotic. The sample is then centrifuged to collect the bacterial cellsin the bottom of the test tube and the supernatant and butterfat is discarded. The bacterial

cells are then re-suspended and mixed in scintillation fluid. Binding is measured with ascintillation counter and compared to a positive and negative control. The more antibioticpresent in the sample the lower the scintillation counts determined by the equipment. Charmcurrently has test kits in this format for -lactams, macrolides, aminoglycosides andsulfonamides.

Immunoassay

Unlike other residue testing methods immunoassays are fast, sensitive, inexpensive,reproducible, reliable and simple to perform. The technique depends upon the measurementof the highly specific binding between antibodies (Ab) and antigens (Ag). Antigens are

substances which are foreign to the body (e.g. bacteria, viruses, toxins, pollens, drugs,hormones and pesticides) and that, when introduced into the body, give rise to the productionof antibodies. Antibodies are proteins produced in the body by white blood cells(lymphocytes) as a result of exposure to antigens (destroy invading pathogens). The extremesensitivity of the immunoassay is due to the development of certain labelling techniques formolecules (conjugates), enabling the measurement of very small masses (picogram or partsper trillion) of substances.

Immunoassays are classified according to the label which is attached to either the antigen(the analyte being measured) or the antibody. The label may be a radioactive atom as in radioimmunoassays (RIA), or an enzyme as in enzyme immunoassays (EIA or ELISA (Enzyme-

linked immunosorbant assay)) or a fluorescent substance as in fluorescence immunoassays(FIA).

There are 3 major types of immunoassays used commonly for the detection of antibiotics inmilk:1) Enzyme-Linked Immunoassay (e.g. LacTek tests, SNAP for Tetracyclines, Single

Step Block for SMZ)2) Enzyme-Linked Receptor Binding Assay (e.g. SNAP for -lactams, Delvo-X-Press)3) Radio immunoassay (CHARM II for tetracyclines and chloramphenicol)


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3.11. Rennet Activity

Coagulation Time versus Setting Time

Rennet is generally d

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