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AgriQuora . 19th Jul, 2021

Quality Tests for Milk and Milk Products

Quality assurance for milk and milk products require a concerted effort from all the key stake holders along the entire dairy value chain. This is because dairy products are highly susceptible to bacterial contamination and/or spoilage.

To ascertain the quality of the raw material and monitor the quality throughout the value chain, the following tests will be very instrumental.

Preliminary/Platform Quality Tests for Milk

The reception platform is the gate where you determine the quality that is fit for production purposes. Here, the quality you assign to the milk will determine the subsequent quality parameters determined at the quality assurance laboratory. These platform tests majorly serve two purposes.

  • First, you can use the tests to identify if the milk is fit for production.
  • Secondly, you can use the tests to monitor the quality of the milk during processing.

These tests will help you to ensure that the product is safe and of the right quality for human consumption.

Reception of Milk at the Platform

At the reception platform, the staff offload the milk from the vehicles and onto the reception platform. The quality assurance officer in charge of reception must ensure that the milk is of right quality before receiving it for further processing.

Preliminary Platform Tests

The initial platform tests are majorly organoleptic (based on the senses) and simple chemical tests. The officer knows the product to be processed from the milk; therefore, he/she ensures adherence to strict quality parameters at the platform. He/she must reject any milk that does not meet the minimum quality standards. Accepting milk of substandard quality will ruin the quality of the finished product.

The officer looks at the color of the milk, feels the temperature of the milk, and smells for an offensive odour. Tasting of raw milk is generally discouraged as it may be a conduit for distribution of zoonotic/infectious diseases. The officer must reject any milk of questionable quality based on platform tests results.

Any mistake in accepting poor quality milk will compound the problem by spoiling the whole batch. Blending a small amount of bad milk with a large volume of good quality milk does not improve its quality; it spoils the milk instead. You can never improve the quality of milk by blending proper milk with a poor quality milk.

1. Organoleptic Tests for Milk

This involves all the preliminary quality tests conducted at the farm level, at the processing plant’s reception platform or at the collection center.

It is a simple, quick and cost effective method of checking quality of milk and allows for segregation of good quality and poor quality milk.

  • The grader must have good senses of sight, smell, and touch because the results of this test is obtained instantly.
  • Start testing by shaking the milk can a little bit then opening the lid and check for any aroma/flavor that emanates from the can by smelling it.
  • Check for visual appearance on the milk such as colour, any other foreign objects on the surface of the milk and the sanitary condition of the milk can. The sanitation of the can indicates how the milk has been handled.
  • Feel the milk can to check the milk temperature. The temperature of the milk can give more information about when the milking was done.

2. Clot on Boiling Test for Milk

This is a cheap, quick and easy to perform quality test that is mainly used to give information on the acidity of the milk. A small amount of milk is boiled in a test tube and checked for any signs of coagulation.

If the milk coagulates, it is an indication that the milk will not be able to withstand processing conditions due to high acidity.

This test is however not quantitative and does not detect any acidity level below 0.2% of lactic acid concentration. Any milk that fails this test is considered too sour for processing.

3. Alcohol Test for Milk

This is a simple and quick method used to test the acidity of milk. Mix equal amounts of alcohol (72% ethanol concentration) and milk sample and check for signs of coagulation.

The mechanism behind alcohol test.

Milk proteins remain stable due to two factors:

  1. Hydration forces
  2. Negative charges on the protein particles

Hydration forces keep the proteins suspended in milk owing to the hydrophilic property of ϰ-casein. Negative charges keep the proteins repellent from each other hence they do not agglomerate into lumps. Destabilizing these forces will lead to flocculation of the proteins.

Alcohol is a dehydrating agent which removes water from the system thereby affecting the hydration forces that are responsible for keeping the proteins suspended.

If such milk has elevated acidity due to microbial activity, the proteins will flocculate due to the combinative effect of hydration force destabilization and charge neutralization.

When combined with alizarin, it gives alcohol alizarin test, which is more informative than just the alcohol test. Alizarin is a dye that changes colour depending on the acidity level.

ColourRed-brownYellowish-brownYellowLilac/pale red
pH6.6 – 6.76.4 – 6.6≤6.3≥6.8
 Normal milkSlightly acidicAcidic (coagulates)Alkaline

Confirmatory Quality Tests for Milk

The officer must subject all the milk that pass platform tests (organoleptic) to additional confirmatory tests. These additional chemical tests further confirm that the milk is of exceptional quality.

Here, alcohol and specific gravity tests become very handy. You carry out alcohol tests by mixing equal amounts of 80% ethanol with milk using the alcohol gun. If the resultant mixture clots, the milk does not have stable proteins to withstand the heat treatment during pasteurization.

Such protein instability may be due to salt imbalance (colostrum in milk or late lactation) in the milk, or the milk is already aged (high acid). Accept only the milk that does not yield a positive alcohol test.

The lactometer test is necessary to determine milk adulteration by adding water (or any other foreign products). Adulteration alters the density of milk and you can detect the difference by the using a lactometer. Normal milk has a lactometer reading ranging from 1.028 to 1.032. Any reading below this range suggests less milk solids (more water than normal) and any reading above this range suggests higher amount of milk solids than normal.

All the milk that passes these tests indicate milk with proper chemical tests. Even though you can accept the milk at this stage, some processors or milk vendors prefer to conduct resazurin test at this point just to be sure that the milk is of high quality.

At this stage, you can weigh the amount of milk received for pricing purposes. After weighing, release the milk into the dump tank.

Interestingly, current dump tanks come fitted with the weighing scales to ease the work at the reception platform. Again, the dump tanks have muslin clothes, which filter off foreign particles that may be present in the milk.

More confirmatory tests (in the lab)

Once you have received the milk into the dump tank, you should take a sample of the composite to the lab for further quality assurance. Using this sample, conduct chemical tests such as titratable acidity, butter fat content, COB, etc.

Please note that some milk processors will conduct resazurin tests on the platform before accepting the milk into the batch to reduce the risk of accepting bad quality milk into the batch.

Resazurin tests check the microbial load in milk; the lesser the load, the better the quality of milk. Poor resazurin results indicate the unsanitary storage conditions for the milk. It also suggests a very poor keeping quality of the milk.

Titratable acidity test determines the level of natural and developed acidity in the milk. Fresh milk should have near neutral pH with little native acidity attributed to the milk proteins.

Here's more in-depth analysis of the confirmatory tests for milk.

1. Acidity Test (Titration/pH method)

Acidity test is performed to determine the level of natural or developed acidity in milk. The level of acidity in milk can be expressed in two ways.

Titratable acidity: - shows the total acidity but does not show the strength of the acid

pH: = shows the strength of acid

Reasons for doing acidity test:

  • To find out the quality of milk
  • To monitor storage conditions because when dairy products are stored in favourable conditions for the growth of microorganisms, they produce acidity.
  • To control manufacturing e.g. for yoghurt, fermentation is done until a desired level of acidity is achieved and then stored in a cold room.
  • Cheese processing also depends on acidity

Natural acidity of freshly drawn milk is due to its constituents e.g. CO2, albumins and globulins, casein and phosphates. These substances react in the milk and produce acidity.

ConstituentLactic acid concentration (%)
Albumins and globulins0.01
Titratable acidity equivalent0.13

Titratable acidity is based on the principle that an acid of a known volume and strength will completely neutralize an alkali with equal volume and strength. The alkali that is commonly used in titration of acidity in milk is 0.1N NaOH.

Titratable acidity is the measure of the amount of alkali required to shift the pH of milk of 6.6 to a determined pH based on the indicator used.

When measuring the acidity of the milk, total acidity is usually used because it encompasses both natural and developed acidity.

Expression of titratable acidity:

The lactic acid concentration is measured in terms of:

Soxhlet Henkel degree (°SH): - the volume (ml) of 0.25N NaOH used per 100ml of sample

Thorner degree (°Th): - the volume (ml) of 0.1N NaOH used per 100ml of sample

Domic degree (°D): - the volume (ml) of 0.1N NaOH used per 100ml of the sample

Acid determination should not be used as the sole basis of accepting or rejecting milk because milk can have acceptable acidity but suffers from flavor defects and other bacterial problems.

Relationship of acidity in different products

The acidity of milk and milk products is usually on the plasma portion of the milk.

Therefore, when you want to compare two dairy products (milk and cream), which have different plasma and fat contents, the calculation of the acidity should be based on the plasma portion of the milk.


Milk with 4% fat content and 0.16% lactic acid and cream with 40% fat content, what will be the acidity of the cream?

Plasma: milk – 96%, cream – 60%.

Comparable acidity of the cream to milk



X=0.1% lactic acid

The same method can be used for cream and butter.

Factors affecting titratable acidity in milk:

  1. Changes from calcium di-calcium phosphate to tri-calcium phosphate and then to phosphoric acid.
  2. Action of calcium phosphate
  3. The stage of lactation e.g. colostrum milk has high proteins hence high acidity
  4. Mastitis or condition of the udder. Mastitis lowers the acidity because blood (which is alkaline) leaks into the milk.
  5. Enzyme activity e.g. lipolysis of fats into fatty acids
  6. The effect of the feeds

2. pH Determination

pH is the concentration of hydrogen ions in milk. It is determined using two methods

a. Calorimetric method

Different indicators that cover various pH ranges are used. The unknown substance with indicator added to it will match the colour of solutions with known pH.

The colour solutions should contain the same indicators as the unknown. The accuracy of this method can be increased by using a comparator which will compensate for the turbidity of the milk.

b. Electrometric method

Uses pH meter/ potentiometer. This method involves measuring the difference in two different voltages of two electrodes in a sample. It gives direct measure of pH with direct values on the display unit.

3. Inhibitory Tests

Performed to check for antibiotics and drug/antibiotic residue in milk. When the antibiotics are present in significant amounts, they inhibit fermentation process. This makes it an important test to consider when dealing with fermented milk products.

Drug residues and the antibiotics are also a health hazard as some people are sensitive to antibiotics. The antibiotic test can be done through various methods, the most common of which are:

Delvo test: - This is the easiest and the fastest method to use but it is very expensive to acquire and operate.

Fermentation test: - bacteria in milk produce acid as they metabolize the milk, antibiotics and drugs inhibit growth of micro-organisms. Milk with antibiotics will take too long to ferment or fail to ferment completely.

4. Specific Gravity (Lactometer Test)

Lactometer is a hydrometer used to measure specific gravity of milk and scale is limited to the specific gravity of milk.

Milk fat has a specific gravity of 0.93, milk has a specific gravity of 1.032 and the milk solids have a specific gravity of 1.036.

Lactometer works on the principle of fluid displacement. The displaced fluid has equal weight as the floating body. Liquids with lower specific gravity have larger displacements than liquids with higher specific gravity.

Lactometers have temperature sensitive and are designed to be used at specific temperatures. It is used to check for adulteration of milk, either by skimming or addition of water or both. When water is added, the specific gravity is lowered towards one.

The design of the lactometer allows for direct reading on the stem whose calibration ranges between 26 to 38. This is because the figure obtained from the reading are on the second and third decimal points of the specific gravity.

The first figures of the specific gravity are 1.0, which are then succeeded by the figures obtained from the lactometer reading.

Any deviation in temperature from the recommended reading temperature is adjusted by either adding or subtracting accordingly.

Common methods of milk adulteration:

1. Skimming

Skimming is the removal of fat from the milk. Given that fat is the lightest portion of the milk, its removal will increase the lactometer reading.

A sample of milk that has low lactometer reading has a high fat content compared to the one with a low fat content.

Fat contentLactometer readingSNF content
lowhighHigh (skimmed)
Calculate % fat content removed Calculate % lowering SNF

2. Watering

When adulteration is watering lactometer reading will go low and SNF will reduce as well as the fat content.

3. Skimming and watering

The percentage of fat removed and the percentage reduction of SNF are not the same when watering and skimming are done. Such samples have low fat content, low lactometer reading and low SNF than normal milk.

5. Freezing Point Test for Milk

Milk freezes at -0.55°C while pure water freezes at 0°C. A small adulteration with water will cause detectable change in freezing point of milk. This makes freezing point test the most accurate test for milk adulteration by water addition.

Freezing point depends on the soluble constituents of milk, especially lactose and chlorides. Freezing point remains the same even though these are differences in the composition of the two components.

The freezing point does not depend on composition of the sample but on the concentration of dissolved salts (lactose and chloride).

As milk freezes, its composition gets unbalanced; solids get concentrated on the remaining liquid part of the milk.

How to determine the amount of water in adulterated milk

Amount of added water = 100(Freezing temp of milk – Freezing temp of suspect milk)

6. Dye Reduction Test for Milk

These quality tests show comparative activity of the micro-organism in milk hence used as a rough indication of the microbial load in milk. It is based on the observation of the blue colour imparted in the milk, which will disappear with time.

The length of time depends on the number of microorganisms present in the milk holding all other factors (such as nutrient content, moisture content, and temperature) constant.

The colour change is assumed to be due to two reasons:

  1. Consumption of oxygen in milk by the microorganism
  2. Enzyme reductase produced by the microorganisms

The time reduction indicates the possible number of microorganism. This test has low correlation with other tests especially those that show presence of bacteria.

To perform this test, take a definite amount of methylene blue and mix with 10 ml of milk. Mix uniformly and incubate at 37°C in a water bath and wait for colour change.

The longer it will take for the colour to change, the lower the microbial load. All the glassware used must be sterile. The interpretation of the results is as follows:

Table 1: Dye reduction test result intepretation of pasteurized milk

Length of time (hours)Quality of milkGrade of milk
5 or moreExcellent1
3 – 5Good2
2 – 3Fair3
Less than 2 hoursPoor4

Raw milk test has a much rapid decolouration.

7. Resazurin Test for Milk

Resazurin test can take 10 minutes for fresh milk and up to one hour for pasteurized milk. This test works on the principle of a dye indicator, which changes colour when oxidized.

Using sterile equipment; add 1 ml of the dye into 9 ml of milk sample, mix uniformly and incubate at 37°C in a water bath for 10 minutes for fresh milk and up to one hour for pasteurized milk.

After the time has elapsed, observe the colour change using a lovibond comparator. The results are interpreted according to the table below

ReadingColourQuality of milkVerdict
5Light blueVery goodAccept
4PurpleGoodAccept (conditionally)
3Purple pinkSuspectSeparate
2Light pinkSuspectSeparate
0WhiteVery badReject

The 10 minutes resazurin test is used at the reception to accept or reject milk due to its rapidity.

8. Butterfat Test for Milk

The most common and accurate method used for butterfat testing is the Gerber method. The method is used for determining the butterfat content in milk and milk products.

Butterfat determination is important because it can be used to value the milk for payment purposes and to determine how to standardize the milk.

You will require the following chemicals and ingredients

  • Amyl alcohol
  • Sulphuric acid (S.G 1.820 – 1.825 or 90 -91% concentration)
  • Milk

Sulphuric acid dissolves all the non-fat content of the milk. The fat content remains in a liquid form due to high heat produced by the acid-protein reaction.

Amyl alcohol provides distinct layer between the two phases, which are separated due to their density differences.

Stronger/weaker concentration should be available. Stronger concentration attacks the amyl alcohol while weaker concentration reduces oxidative effect of the acid.

Amyl alcohol used should be colourless – iso-amyl alcohol – an isomer of two alcohols; methyl and butanol.

Different Gerber butyrometers are used for different products depending on their different fat contents.

Problems encountered during butterfat quality tests:

i) Curd test

There is a formation of a slightly coloured solution after addition of the acid or a curdy fat column. This usually results from using a weak acid, which fails to break the fat globule membrane. It may also result from putting insufficient acid or insufficient mixing of milk and acid.

ii) Charred test

This happens when there is a darkened fat column with dark specs at the base of the fat column. This may be because the acid is too strong, or the milk and acid were mixed slowly, or the milk was added/dropped into the acid. There might be too much acid causing the milk to burn.

When testing low fat content sample, centrifugation should be done more times (three times) because the milk is homogenized.

iii) Babcock method of testing butterfat

This method uses the same principle as the Gerber method, the only difference is the design of the butyrometers used. Babcock method uses butyrometers with cylindrical stems.

Chemical Test Procedures for Quality Assurance in Dairy Products

Listed here are the conventional chemical test procedures that are carried out on dairy products to assure quality of the products. You will realize that I have not indicated the lactoscan equipment in any of these tests.

The machine can execute some of the tests indicated here automatically and give you a printout of the results or display the results on its screen.

Since it is not possible for everybody to access the lactoscan, these methods remain very instrumental and in some cases, they have to be done to confirm the lactoscan results.

These tests remain to be the most reliable quality assurance measures in the industry going by the local industry standards.

1. Chemical test procedures for determining the C.I.P detergent strength

The purpose of this method is to ensure accurate determination of concentration of detergents used for Cleaning In Place (C.I.P).

This work instruction covers the procedure for determination of strength of lye (NaOH) and Nitric acid (NHO3)

Necessary Apparatus and Reagents

Preparation Instructions

  1. Calculate the acid concentration by multiplying the titer value by the factor 0.233
  2. To determine the acid strength, titrate the acid sample against 0.1 N NaOH solution until the colour changes from clear to purple
  3. Calculate the % concentration by multiplying the titre by the factor 0.133
  4. Record the titer value obtained
  5. Add 2-3 drops of phenolphthalein indicator and titrate against 0.1 N Hcl solution until the pink colour disappears.
  6. Draw 3 ml of lye solution using the 10 ml pipette and transfer into a beaker cup
  7. Fill the 50 ml burette with 0.1N Hcl solution and adjusted to the 0.0ml mark
  8. To determine lye concentration, titrate the lye sample against 0.1N Hcl

2. Chemical test procedures for determining density

The purpose of this method is to ensure uniform practice in determination of density of both raw and pasteurized milk.


  • Lactometer jar
  • Milk lactometer (with a valid Calibration certificate).
  • Thermometer range 10 -110°C (calibrated)

Preparation Instructions

  • Raise the temperature of the milk sample to 40°C; keep the sample at 40°C for 5 minutes. During this time, mix the sample by rotating and inverting. Cool to 20°C, mix again.
  • Fill the lactometer jar with sample milk and gently insert the lactometer into the milk.
  • Wait for at least 2 minutes (until the setup settles) and do not disturb the apparatus.
  • Read the density at the level of meniscus formed between the milk and the lactometer stem. At the same time record the milk temperature.
  • If the time of taking the reading the temperature was not equal to the calibration temperature, correct the reading by adding the value specified in the table below, which corresponds to hydrometers calibrated at a temperature of 20°C

22.0+ 0.0014
21.5+ 0.0013
21.0+ 0.0012
20.5+ 0.0011
20.0  0
19.5- 0.0009
19.0- 0.0008
18.5- 0.0007
18.0- 0.0006
  • Remove the lactometer gently and drain off the milk into a sample bottle.
  • Carry out butter fat test on milk.
  • Calculation of Total Solids (T.S).

T.S = {(Corrected Lactometer Reading)/4} + (1.22 x B. Fat) + 0.14

S.N.F = T.S. – B.F

T.S. = CLR – 1 (1000 ÷ 4) + (1.2 x B.F) + 0.14

S.N.F. = T.S. – B.F (Richmond formula for finding S.N.F)

3. Resazurin Test

Resazurin test is conducted to ensure milk (pasteurized and raw) has the microbial load within specifications.

Required Equipment and Reagents

  • Water bath at 37°C ±0.5°C
  • Sterile test tubes.
  • Sterilized rubber stoppers
  • Resazurin solution -0.005% made after every 8 hrs.
  • Sterile 1ml pipettes
  • Comparator with standard resazurin disc

Preparation Instructions

  • Prepare the water bath and set the temperature at 37°C
  • Pour out 10mls of milk sample into the sterile test tube (most of them come already calibrated to the 10ml mark)
  • Add 1ml of resazurin solution into the contents of the test tube.
  • Stopper the test tube with clean, sterilized to avoid any possible contamination.
  • Mix the sample gently until the dye is uniformly distributed
  • Place the test tube in the water bath at 37°C and read after 10 min, 30 minutes then 1st, 2nd and 3rd

Acceptable Standards

Only resazurin 5 and 6 after 10 min is acceptable for raw milk, for pasteurized milk only the milk that finishes 3 hrs with the resazurin 6 is acceptable.

4. Chemical test procedures for determining the acid content in dairy products

Acidity test is carried out to determine the amount of lactic acid in raw and pasteurized milks, fermented milks, and cream.

Equipments and reagents


  • Fill the burette to the mark. Remove with tissue any NaOH at the tip of the burette.
  • Using a blowout pipette take 10 ml of the milk sample and transfer into the beaker
  • Add 2-3 drops of phenolphthalein indicator into the sample
  • Mix the indicator into the sample gently
  • Titrate the sample quickly and continuously until the first permanent faint pink colour appears and persist for at least 1 sec.
  • Read off the volume of sodium hydroxide used and divide by 10 to get the percentage acidity, which is expressed as percentage lactic acid.

N/B: Natural acidity in fresh milk is due to presence of phosphates, calcium, and carbon dioxide. Developed acidity is a result of microbial activity.

5. Chemical test procedures for determining sediment content

Sediment test is conducted to check the amount of solid dirt in raw and pasteurized milk

Equipments and Reagents

  • sediment tester
  • thermometer (range 10 -1100C)
  • sediment test discs
  • distilled water
  • standard sediment grading chart

Preparation Instructions

  • Take a ½ litre of milk sample and adjust the temperature to 400c and then cool to 200c
  • Clean the sediment tester thoroughly with filtered water
  • Place a clean filter pad, with the name of the supplier into position.
  • Put the ½ litre milk sample into the tester and let all the milk pass through the sediment disc/ pad.
  • Remove the filter pad and place it on a clean surface (parchment paper) to dry in a dust free environment for at least 10 minutes.
  • Grade the milk using the standard grading card

6. Hydrogen Peroxide Test

Conducted to check presence of hydrogen peroxide in Milk


Hydrogen Peroxide Test Strips/ Sticks

Preparation Procedure

Dip the mashed part of the strip in a sample of milk


Colour change to blue indicates presence of hydrogen peroxide. If colour does not change then hydrogen peroxide is not present in the milk sample

7. Clot on Boiling (CoB) Test

This test is conducted to check the stability of milk proteins, whether it can withstand heat treatment


Test Procedure

  • Put a sample of milk in an aluminum container
  • Heat the milk sample over the flame of the Bunsen burner


  • If the milk clots, it is COB positive (cannot withstand heat treatment).
  • If it does not clot, then it is said to be COB negative

8. Butterfat Content Testing (Gerber Method)

Gerber butterfat test is conducted on milk to determine the amount of butterfat it contains

Apparatus/Equipment and Reagents

  • Butyrometer (Dr. N. Gerber 0 – 8%)
  • 10.94 ml bulb pipette
  • Self-heating centrifuge at 650C (1100 – 1200rpm)
  • Automatic dispensers for 1ml – amyl alcohol
  • Automatic dispensers for 10ml – sulphuric acid
  • Sulphuric Acid (1.815 - 1.820 g/ml at 200C) - Corrosive!
  • Amyl Alcohol (0.810 – 0.815 g/ml at 200C)

Preparation Procedure

  • Dispense 10 ml of sulphuric acid via an automatic dispenser in to the butyrometer. (Avoid wetting the neck of the butyrometer)
  • Pipette 10.94 ml of well mixed milk sample at room temperature and put it into the butyrometer gently along the side of the butyrometer so as to avoid mixing the milk with acid  (Avoid wetting the neck of the butyrometer)
  • Add 1 ml of Amyl Alcohol. (Avoid wetting the neck of the butyrometer)
  • Cork the butyrometer, shake and keep inverting the butyrometer until a deep chocolate color is observed.
  • Put the butyrometer in the centrifuge at 650C and spin for 5 min
  • Read the results and record.

9. Chemical test procedures for determining butter moisture content

This test is used to determine the moisture content in butter by evaporation method


Preparation Procedure

  • Weigh the Kohman cup and record the weight - A
  • Weigh 10 g of butter (from the centre of the sample) in to the Kohman cup - B
  • Heat the sample on a Bunsen burner swirling to avoid charring until all the moisture escapes. When the bubbling stops and colour changes to golden brown, stop heating - C
  • Allow the cup to cool to the original temp
  • After cooling the sample is reweighed – D
  • The difference in weight is the moisture content

For instance; say, Weight of Kohman cup – A = 40g

Weight of sample  = 10g

Weight of cup + butter sample – B = 50g

Weight of cup + sample after heating and cooling –D = 48.4g

Moisture content = B – D ÷ 10 x 100 = 50 – 48.4 ÷ 10 x 100 = [1.6 x 100]/10

Moisture content = 16%

10. Chemical test procedures for determining salt content in butter

This test is conducted in salted dairy products to determine the salt concentration in these products

Apparatus and Reagents

  • A pair of tongs
  • Kohman cup
  • Bunsen burner
  • Beaker
  • 250 ml volumetric flask
  • 6 ml pipette
  • Silver nitrate
  • Potassium chromate
  • Distilled water
  • Thermometer

Preparation Procedure

  • Warm the distilled water to a temperature of between 40 – 500C
  • Using the heated butter sample whose moisture content has been determined, rinse the butter contents from the Kohman cup with the warm distilled water as you put it put into the volumetric flask until the 250 ml mark.
  • Leave it to settle so that the fat floats and the curd sediments
  • 6 ml of the solution is pipetted from the middle and put in a beaker
  • Add one drop of potassium chromate indicator
  • Titrate it against Silver nitrate until endpoint.

11. Chemical test procedures for determining free fatty acids in butter

This test is conducted to determine the amount of fatty acids present in milk.

Equipment/Apparatus and Reagents

  • Conical flask, 250ml
  • Measuring cylinder 50ml
  • Pipette 10 ml
  • Weighing balance of 0.0gm precision
  • Ethanol-Ether mixture ( prepared by mixing equal volumes of 95% ethyl alcohol and diethyl ether, and neutral to phenolphthalein.
  • Sodium Hydroxide 0.1N.
  • Phenolphthalein indicator 2.5% in 95% ethyl alcohol.

Preparation Procedure

  • Weigh a sufficient amount of butter in a beaker or a flask
  • Melt indirectly using a Bunsen burner.
  • Weigh 10 grams of the melted butter, carefully avoiding the aqueous phase in to a beaker
  • Add 50ml of FFA (Diethyl ether/ Ethanol) reagent to the melted butter
  • Add 5 – 6 drops of phenolphthalein indicator
  • Titrate against N/9 NaOH up to endpoint
  • Read off the titre


Multiply the titre value by 0.282 to get FFA (0.282 is a constant factor)

12. Peroxidase Test

This test is conducted to verify the effectiveness of high temperature (>800 c) pasteurization of cream and milk used to make fermented milk products.

Apparatus/Equipment and Reagents

  • Red and blue litmus paper,
  • Test tubes (medium size),
  • Stop watch,
  • Water bath at 800C
  • 1n Hcl in pipette or dropping bottle,
  • 1% NaOH in pipette or dropping bottle,
  • 2% H2O2 in pipette or dropping bottle,
  • 2% paraphenylenediamine in pipette or dropping bottle.
  • Keep reagents in cold storage

Preparation Procedure

0.2% H2O2 for Storch’s test: Dilute 7 ml of 3% hydrogen peroxide solution (Ph.Nord III, 1963,p.302) with 98 ml of distilled water. Stabilize with 0.1 ml concentrated sulphuric acid. Store in amber bottle. Will keep several months if stored in a cool place.

2% Paraphenylenediamine: Dissolve 2g paraphenylenediamine (for analysis) in 100 ml of cold distilled water by shaking filter. Store in amber bottle. Will keep one week. A precipitate will occur in course of one to two days, which should be filtered off.

The principle involved here:

Raw milk contains an enzyme, milk peroxidase, which can accelerate the oxidation process of the hydrogen peroxide. The peroxidase enzyme is destroyed by heating. At 800c the enzyme will become inactive in 21/2 seconds. At higher temperatures the inactivation will take shorter time, at lower temperatures it will take longer.

Test procedure

Whole milk. Approximately 5 ml of milk is tested with litmus paper in a test tube. If the reaction of the milk is not about neutral, pH is adjusted with 0.1N Hcl or 1%NaOH. Add one drop of 0.2% H2O2 and 2 drops of paraphenylenediamine. Shake, check the time it takes for the blue colour to appear using a stopwatch.

Testing Cream. The procedure remains the same as for whole milk.


  1. 1 second = Instantaneous intense blue colour which turns dark blue in 30 seconds indicates 100% raw milk or inadequately pasteurized milk to which more than 20% of raw milk has been added, or milk which has only been heated to approximately 73 – 740c in the pasteurizer for 21/2 seconds.
  2. 2.5 – 10 seconds = Lighter blue colour appearing in the course of 5 – 10 seconds, assuming a dark blue colour in 30 seconds, but lighter than described under 1, indicates inadequately pasteurized milk to which 4 – 20% of raw milk has been added or milk which has only been heated to 740c – 750c in the pasteurizer for 21/2
  3. 15 – 20 seconds = Faint grayish colour appearing in 15 – 20 seconds, assuming a faint grey-blue tint in 30 seconds, lighter than described under 2, indicates adequately pasteurized milk/ cream to which 2 – 3% raw milk/ cream has been added, or milk/ cream which has only been heated to 75 – 770c in the pasteurizer for 21/2 seconds.
  4. 20 – 25 seconds = Slight greyish tint appearing in 20 – 25 seconds, assuming a light grey colour in 30 seconds, indicates adequately pasteurized milk to which 1 – 2% raw milk/ cream has been added or milk/ cream which has been heated to 770 – 780c in the pasteurizer for 21/2 seconds.
  5. No colour developing within 30 seconds indicates properly pasteurized milk/ cream (addition of less than 0.5% raw milk/cream cannot be demonstrated with Storch’s test).

Any colouring after 30 seconds is of no significance. Adequately pasteurized milk/ cream will always colour after prolonged standing owing to spontaneous oxidation of the paraphenylenediamine.

13. Viscosity Test Procedure

Conducted on milk and milk products to check consistency/viscosity

Required Apparatus

A rheometer would be ideal. In its absence, however, use the following:

  • Viscometer stand
  • Viscometer
  • Timing device (stop watch)
  • Sample (100ml)
  • Apertures (3mm and 5.5mm)
  • Beaker

Test Procedure

  • Clamp the viscometer to the stand at a fixed height.
  • Select the aperture that is appropriate for the sample to be tested and screw it to the viscometer.
  • Adjust, as necessary, the temperature of the sample. (200c - 250c)
  • Place a beaker beneath the viscometer.
  • Hold the aperture with the index finger.
  • Fill sample into the viscometer cup.
  • Remove the finger allowing the sample to flow, simultaneously switch on the stop watch.
  • Record the time taken for the sample to cross the bottom mark.
  • For improved assurance and precision, perform three tests, record the results from all three, and use the average as a best estimate of actual viscosity.
  • Clean the viscometer immediately after each use.

14. Brix Determination

Measures the refractive index of milk and milk products


  • Plastic pipette
  • Refractometer
  • Soft tissue
  • Sample (0.1ml)
  • Distilled water


  • Press the ON/ OFF key.
  • Using a plastic pipette, fill the sample well with distilled water.
  • Press the ZERO key.
  • Gently absorb the water with a soft tissue.
  • Using a plastic pipette, drip sample (0.1ml) onto the prism surface.
  • Press the READ key. Measurement is displayed in units of percentage BRIX.
  • Remove sample from the sample well by absorbing on a soft tissue.
  • Using a plastic pipette, rinse prism and sample well with distilled water. Wipe dry.

15. Homo-Efficiency (H.E) test procedures

To determine the measure of creaming of fresh milk after pasteurization


  1. Refrigerator
  2. Measuring cylinder (200 – 250ml)
  3. Apparatus for butter fat determination
  4. Suitable pipette

Test Procedure

  • Place 200 – 250mls of pasteurized milk in measuring cylinder for a minimum of 12 hours in a refrigerator.
  • Without disturbing the sample, draw 10% milk from the top of the cylinder.
  • Mix thoroughly and determine the butter fat content of the above sample and call it, A.
  • Mix thoroughly the remaining milk sample and determine its butterfat content, B.
  • H.E = 100 – {[(Butterfat in A) – (Butterfat in B)]/Butterfat in B} x 100

 Interpretation of results


 96 – 100%Very good quality milk
90 – 95%Good quality milk
< 90%Poor quality milk

pH Determination: Monitoring The Acidity Of Dairy Products

You may wonder whether pH determination and monitoring is of any significance in dairy products processing. As we are about to discover, pH is very important to the living cells. This is because the living organisms need a proper balance of hydrogen and hydroxide ions to maintain the essential physiological processes.

Consequently, scientists have used pH to express the hydrogen ion (proton) concentration in a medium. It is essential to ensure that the concentration remains within a narrow range to continue supporting the tightly regulated body systems.

Microorganisms use milk as a substrate for their metabolic processes. They use enzymes, whose activity depend on the pH of the medium, to catalyze their metabolic reactions. Their metabolic byproducts such as lactic acid affect the quality of milk.

It is necessary to determine the pH of milk so as to check the strength of lactic acid in a given sample of milk to help gauge if the lactic acid concentration in the sample is within the acceptable range.

You use a pH meter to determine the pH of the sample. The meter uses an electrode system to measure the potential difference between its electrodes.

The pH meter displays the results numerically on the screen on a pH scale that ranges from 1-14. Normal fresh milk has a pH range of between 6.3 – 6.5.

pH determination experiment

For this exercise, we carried out an experiment with a fresh milk sample. We placed the sample in a beaker and dipped the electrodes of the pH meter into the sample.

We took the readings and repeated the process two more times to ensure that the result was consistent. Before every replicate of the test, we dipped the electrodes in distilled water to avoid ‘confusing’ the meter.

On every replicate of the test, the meter showed 6.44, which is within the acceptable range for fresh milk.


Lactic acid is undesirable in fresh milk since it deteriorates the quality of the product. Excessive production of lactic acid will lead to coagulation of the milk proteins and render the milk unfit for its intended use as a fresh product.

Even though lactic acid fermentation has been manipulated to produce amazing dairy products such as yoghurt and cheese, it may not be desirable when you aim to produce ice cream or fresh milk.

Furthermore, excessive acidity in fermented products such as yoghurt will impart excessive tart to the product and make it less palatable.

pH determination and monitoring in dairy products is important to ensure that the acidity remains within the prescribed range.

Protein Content Determination: Kjeldahl Process

Kjeldahl process of protein content determination involves digestion of the sample in a Kjeldahl digester in the presence of concentrated sulfuric acid and selenium tablet, which acts as a catalyst for the process.

It is a simple, precise, and a straight forward method, especially for learners, to get acquainted with.

You can apply this process on a large variety of foods to determine the nitrogen content, which you then translate as a protein content using a conversion factor.

Kjeldahl protein content determination procedure (cheese sample)

  1. Obtain a duplicate 0.2 g of the cheese sample to be tested and put in round bottomed flasks
  2. Add 10 ml of concentrated sulfuric acid followed by selenium tablet into each flask
  3. A control flask should contain no cheese/food sample to be tested
  4. Set the three bottomed flask into a Kjeldahl digester in a fume chamber to suck the fumes during the digestion process
  5. Heat the samples until the contents of the flasks remain clear (without any colour).
  6. After all the colour has disappeared, continue heating for one hour (time it) and then turn off the heat
  7. Transfer the contents of each conical flask into a separate clean distillation tube and mount onto a distiller.
  8. Into separate clean conical flasks, add 10 ml of HCl followed by a few drops of mixed indicator (methyl red and bromocressol green). Mount each onto the exhaust end of the distiller to collect ammonia gas escaping from each distillation tube during the distillation process
  9. Set the distiller to run for five minutes
  10. Set 50 ml of 40% NaOH in the burette for titration

After collecting the ammonia gas in the conical flask, titrate it using the 0.1N NaOH solution.

The titration process intends to determine the amount of unused Hcl.

This is the value that will be instrumental in the determination of the protein content using the illustrated formula below.

Nt = {(V1-V2)N*1.4007}/Weight of sample

Where; Nt – Nitrogen content

V1 – Titre value of the blank

V2 – Titre value of the sample

N – Normality of the acid used


After doing the experiment, the values I obtained were as follows:

Blank titre – 9.9 ml

Cheese sample titres (A – 5.7; B – 5.6)


To obtain the protein content of the sample, first determine the nitrogen content of each then calculate their average. From there, use the protein content conversion factor to get the protein content of the sample under analysis.

Sample A: Nt = {(9.9-5.7)*0.1*1.4007}/0.2 = 2.94

Sample B: Nt = {(9.9-5.6)*0.1*1.4007}/0.2 = 3.01

Therefore; the protein content of the cheese sample = 6.38*{(3.01+2.94)/2} = 18.98%

Note: 6.38 is the protein content conversion factor for cheese. Different foods have different conversion factors.

Freezing Point is More Dependable Than Lactometric Method in Determining Milk Adulteration

The freezing point of a pure water (0°C) depends on the structure and chemical properties of its molecules. Adding solutes to water will depress the freezing point of the solution. The freezing point of milk is lower than that of pure water due to the presence of milk solids.

Milk has a constant proportion of milk solutes hence will have a definite freezing point. Research has shown that milk freezes at temperatures between −0.53 and −0.56 °C. A deviation from this range indicates adulteration and explain why freezing point of milk gives accurate results for adulteration.

This is because the physiological conditions of a lactating mammal works to keep the osmotic pressure of blood and milk equal.

Milk adulteration plagues the dairy industry. It is the devaluation of milk quality by either addition of inferior/unacceptable product or removal of a valuable ingredient.

Is milk adulteration intentional or incidental?

Adulteration can be either intentional or incidental. Intentional adulteration happens when an individual decides to either remove content or add content to milk to achieve a certain goal. Incidental adulteration can happen at any point in the dairy value chain and may go unnoticed.

An adulterant is any extraneous matter that will make the food product unsafe or sub-standard.

Adulteration of milk by water addition will raise the temperature of milk towards the freezing point of water. Addition of solutes in milk will depress the freezing point of milk further.

Why would someone add water to milk?

Simple. Many shady producers/traders boost their milk volumes by dosing with water. This is partly due to the fact that they get paid by the mass of milk delivered.

Factors affecting the freezing point of milk

It is important to determine the freezing point of milk since it is easy to detect adulteration my water addition.

Freezing occurs when water and ice are in an equilibrium. This is due to constant vapor pressure between the two forms of water.

Here are the factors affecting the freezing point of milk:

1. Genetic predisposition

As we have already noted, the freezing point of milk vary due to some inherent factors. You will notice that milk from high producers tend to have a higher freezing point than that from low producers. This difference is attributed to the difference in solute concentrations between the two milk samples.

Low producers tend to produce milk with high concentrations of sugars, minerals, butter fat, and proteins. You can read the research findings here.

2. Stage of lactation

Milk composition varies at different stages of lactation. During early lactation, milk sugars are high in the colostrum milk while salts dominate in the late lactation.

Solutes present in the milk will disrupt the equilibrium and delay vapor escape from the surface of a freezing body. This will lower the freezing point of the milk sample.

3. Freshness of the milk sample

The osmotic pressure difference of the medium will also affect the freezing point of milk. Higher solute concentration will lead to a higher osmotic pressure, which will lower the freezing point of the sample.

Souring of milk leads to breakdown of lactose, which increases the number of solutes in the milk hence further depression of the freezing point.

4. Health of the lactating cow

As we have already noted, the osmotic pressure of a healthy cow is constant between the blood and the milk. Any deviation in the health of the animal will lead to disruption of the osmotic pressure equilibrium.

As a result, solutes will leak into the milk leading to higher solute concentration, which increases freezing point depression (lowers the freezing point further).

5. Mastitis

Mastitis plagues a large majority of dairy farms. Research has shown that mastitis will increase the freezing point depression of milk due to destabilization of milk solutes.

Watch the video explaining how the OptiFZP freezing point analyzer works. You can use this digital analyzer to determine the freezing point of milk.

Video by PAC LP

Skimming of milk destabilizes solute balance hence will affect the freezing point.

Methods for determining the freezing point of milk.

1. Hortvet Cryoscope method

Uses the Hortvet Cryoscope to determine the freezing point of milk. One of the earliest methods developed in the early 920s.

Please note that the machine is quite taxing to use efficiently and it does not guarantee reproducibility of results.

The Horvet cryoscope uses an ordinary freezing point thermometer.

2. Thermistor Cryoscope method

This cryoscope has special probes that measures the changes in temperature as well as electrical resistance.

The Association of Official Analytical Chemists (AOAC) recommends the Thermistor cryoscope method for determination of milk freezing point depression.

It is also called Fiske Cryoscope or Advanced Milk Cryoscope.

You can get a complete Thermistor Cryoscope manual here.

How to calculate the percentage of added water by using the cryoscope.

By determining the freezing point of milk, it is possible to find out the amount of water of adulteration. This relies on the convention that pure unadulterated milk has a freezing point depression of 0.54°C.

The term depression is used to denote the negative sign (-). That is why we indicate the value in absolute figures.

Use the formula below to get the percentage of added water

Added water =((0.54 − ΔT)/(0.54)) × (100 − SNF)

ΔT is the deviation in temperature

SNF = % Solids-not-fat of milk sample

Further Reading

  • Henno, M., Ots, M., Jo˜udu, I., Kaart, T., & Ka¨rt, O. (2008). Factors affecting the freezing point stability of milk from individual cows. International Dairy Journal, 18(2), 210-215. doi:10.1016/j.idairyj.2007.08.006
  • J.O.L.King. (1969). The Effect of Mastitis on the Freezing-Point of Cows' Milk. British Veterinary Journal, 125(1), 25-30. doi:https://doi.org/10.1016/S0007-1935(17)49160-3
  • Radewonuk, E., Strolle, E., & Jr., J. C. (1983). Freezing Points of Skim Milk Concentrates. 66(10), 2061-69. doi:https://dx.doi.org/10.3168/jds.S0022-0302(83)82051-7
  • Zagorska, J., & Ciprovica, I. (2013). Evaluation of Factors Affecting Freezing Point of Milk. International Journal of Nutrition and Food Engineering, 7(2), 106-111.

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