SCRIPT FOR THE PRACTICAL CLASS OF DAIRY INDUSTRY

1 - pH

The production of lactic acid and/or other acid components produced by

microorganisms, change the acidity and, consequently, the pH.

PRINCIPLE: verification of the milk pH using a pH meter.

Sample of milk; potentiometer; 50 mL beaker.

TECHNIQUE: read the pH of the sample, checking if the electrode is fully

immersed in the sample.
Normal pH result: 6.2-6.8

2- DETERMINATION OF STABILITY BY ALCOHOL TEST

PRINCIPLE: 76% alcohol is used in equal parts with the milk to be

analyzed and it is verified whether there is coagulation or not.

MATERIAL
5 pipettes of 2 mL; 4 test tubes; 76% alcohol.

TECHNIQUE: Place 2 mL of 76% alcohol in a clean, dry test tube, add 2

mL of the sample. Homogenize and check the coagulation.

INTERPRETATION:

WITHOUT COAGULATION: normal milk glides in a thin uniform layer along

of the tube - acidity below 19oD;

2. FINE COAGULATION: acidity from 19 to 20oD;

3. COAGULATED: acidity above 22oD.

3 - ALIZAROL TEST
PRINCIPLE: the alizarin solution in contact with the milk forms a brick-red color in the
normal milk, a violet color in alkaline milk and a yellow color in acidic milk.
MATERIAL: * Milk;
Alizarol solution;
Salut Acidimeter.
TECHNIQUE: place approximately 2 mL of milk in a test tube and,
at rest, add 2 mL of alizarin solution.
Brick-red - WITHOUT COAGULATION - milk
normal, freshness, acidity of 17 to 18°D;

Red-brown - WITH FINE COAGULATION - milk with acidity of 18 to 21°D;

Yellow - COAGULATED - milk with acidity above 21°D;
Violet - WITHOUT COAGULATION - alkalized milk or diluted with water.
4 - DENSITY
The density of a substance is obtained by relating its mass or weight of equal
volume of a substance taken as a standard. Water is the standard for solids and liquids.
while hydrogen is taken as the standard of gases.
The density of milk at room temperature varies between levels of 1.028 to 1.034;
averaging: 1.032.
PRINCIPLE: densimeters or hydrometers operate on the following principle: "if a body
It floats in a liquid, it is pushed upwards by a force equal to the weight of the liquid that it displaces.
dislocate.
Note: milk densitometers are called lactodensimeters or lactimeters.

METHOD: Density reading using the QUEVENE lactometer.

Material:
- Milk sample;
- QUEVENE lactodensimeter;
- Thermometer;
- 250 mL graduated cylinder.
Care with the sample:
- They must be homogeneous and representative;
- Should not contain greasy lumps or clots;
- The material used must be clean;
- The lactodensimeter must be calibrated.

Technique: (knowing the temperature of the milk before taking the reading).
Place 200 mL of milk in a 250 mL graduated cylinder. Introduce the densitometer without it
Touch the edges of the graduated cylinder. Make the first reading after allowing the densimeter to settle.
at the highest point of the lactodensimeter stem. Clean the stem with paper and return.
carefully up to very close to the point of the first reading before releasing it. Note only
the second reading. Read and record the temperature of the milk. Correct the influence of the temperature by
calculations, or through specific tables.

5 - ACIDITY OF MILK BY THE DORNIC METHOD

The acidity of milk is quite variable, being higher in milk with higher levels of
dry, degreased extract, reflecting the buffering power of the product, within the understood range.
between the pH of the sample and the phenolphthalein's transition pH.
The milk, when it comes out of the udder, is slightly acidic, partially due to some
components such as: casein, phosphates, citrates, CO2, etc., and partly to the internal reaction that occurs
during the titration with alkaline solution.
This acidity is normally understood to be between 14 and 18.oIt is called titratable acidity
initial or natural.

PRINCIPLE: determination of the acidity of milk, titrating with NaOH N/9 solution (Dornic),
and using a phenolphthalein solution as an indicator.
METHOD: Dornic
MATERIAL:
Milk to be titled;
Phenolphthalein solution;
N/9 NaOH solution;
25 mL burette;
4 erlenmeyer flasks of 50 mL;
4 volumetric pipettes of 10 mL.
TECHNIQUE:

With the aid of a volumetric pipette, measure 10 mL of the milk sample;

Transfer to a 50 mL Erlenmeyer flask;
Add 1 mL of alcoholic phenolphthalein solution;
Take the burette or the apparatus containing Dornic solution (NaOH N/9) and titrate (Do the
titration drop by drop until a permanent pink color appears.

CALCULATIONS:
1) When the titration is carried out in a burette that measures directly in degrees Dornic (°D) not
there is no need to do any calculation. In this case, the measured value is already in degrees
Dornic.

2) When the titration is carried out with a common burette, the following calculation should be done:

Dornic GrayoD) = V x 10;

Where V = mL of N/9 NaOH solution needed to neutralize 10 mL of milk.

1/10 x V

the difference between the results of two parallel determinations should not exceed 0.3oD.

6 - FORMALDEHYDE RESEARCH
Test tubes;
Milk
50% sulfuric acid
For ferric chloride

TECHNIQUE:
1. Add 5 ml of milk to a test tube, then add 2 ml of
H2SO4Add 1 ml of ferric chloride, mix everything and heat over a Bunsen burner.
2. The result will be positive for blue-violet and negative for yellow.
greenish.

7 - FAT

The determination of the fat percentage in milk serves to establish a base for
collection of milk by commercial or industrial destination (consumption milk for cheese, etc.);
assistance in the selection of dairy herds; milk integrity in the investigation of frauds;
dairy contests at exhibitions and even forecasts of industrial yields.

PRINCIPLE: Under the action of sulfuric acid, the fat separates from the other components of the milk.
with the help of amyl alcohol and subsequent centrifugation.

METHOD: Originally envisioned by Dr. Gerber, now the most used method for
determination of the percentage of fat in milk.

MATERIAL: Milk
Gerber cream test for milk;
10 mL and 1 mL pipettes;
Sulfuric acid (d=1.820 - 1.825/15-20)o C) - concentrated;
Amyl alcohol (d=0.815);
 Double boiler 65-68oC;
Centrifuge;
Absorbent paper.

CAUTIONS: Sulfuric acid is highly corrosive. In case it comes into contact with clothing, place
immediately on site ammonia or diluted solutions of NaOH or carbonate of
sodium. If it comes into contact with skin, mouth, or eyes, wash immediately with plenty of water
apply a 2% boric acid solution. apply picrate ointment to the hands.
of the goal.
Amyl alcohol should not be pipetted, the odors are toxic, it should be stored.
in a dark, tightly closed bottle, as moisture absorption can dilute it.

TECHNIQUE: Place 10 mL of concentrated sulfuric acid in the butyrometers.

Standardize the milk sample.
Take 11 mL of milk and put it in the butyrometer, being careful to clean the
the tip of the pipette with a towel and let the milk flow slowly along the walls of the
butterometer with inclined pipette. Drain all the milk.
Add 1 mL of amyl alcohol.
Clean the neck of the butyrometer, cork with clean and dry corks, without
defects, associating with a slight twist to the right.
Shake the butyrometers until the clot is completely dissolved. Centrifuge the
1000-1200 rpm for 3-5 minutes.
Make the reading.

CAUSAS DE ERRO:
Very dark content: excess acid, denser acid, hot acid.
Clear content: slightly acidic, less dense acid, very cold acid.
Lack of uniformity: inadequate centrifugation, lack of amyl alcohol.

8 - TOTAL DRY EXTRACT (EST) AND DEGREASED DRY EXTRACT (ESD)

The procedure is carried out in this discipline through the determination of the EST with
with the help of the 'Ackermann Disk', which contains concentric scales for milk density,
fat theory and externally, the total dry residue. The reading is performed by aligning the
density values and fat content. Looking for the arrow of the disc. Where it points will be
indicating the total dry extract (in %).
The degreased dry extract ESD is the result of subtracting the fat content from
value of EST or, ESD = EST - % fat.

The following calculation can still be used for the measure of defatted dry extract (DDE):
ESD=8,652- (0.084xG)
Expected result: Min 8.4 g/100g

9 - PEROXIDASE RESEARCH

In the verification of the temperature at which the milk was heated, the peroxidase research is
of absolute importance. The presence of peroxidase is closely related to the time-binomial
temperature, a basic factor in the efficiency of pasteurization.

The peroxidase test must be positive for pasteurized milk. The negative result
indicates that the milk has been overheated or boiled.
PRINCIPLE:

Peroxidase is an enzyme that catalyzes the reaction:

peroxidase
H 2O2 + HA →2H 2O + A

Where HA is an oxidizable or hydrogen-donor substance. In this case, we used

a saturated guaiacol solution, which is colorless and, in contact with milk and hydrogen peroxide
it will form a salmon color, detecting the presence of the enzyme from raw milk or properly
pasteurized. In contrast, if the milk has been overheated or boiled, the reaction will give
white coloration, showing that the enzyme was destroyed.

METHOD: Dupouy test

MATERIAL:
milk sample
saturated solution of guaiacol;
10 volume hydrogen peroxide;
test tubes;
5 mL pipettes.

TECHNIQUE: Place 2mL of milk in a test tube at 25-30. oC, 2mL of solution
saturation of guaiacol and 1 to 2 drops of 10-volume hydrogen peroxide.

INTERPRETATION:

Salmon heart: positive test, properly pasteurized raw milk;

White color: negative test, overheated or boiled milk.

10 - REDUCTASE
Methylene blue or resazurin is used by indirect method to determine the
bacteriological quality of milk in relation to the metabolic rate of various microorganisms
that produce reducing substances from milk.

PRINCIPLE: The production of oxidoreductases by bacteria allows the transfer of

hydrogen in the medium, with methylene blue being one of the receptors.

CARE:
• Reduction tests must be conducted under light due to their
direct action on the dye, regardless of the microorganisms
presents.
• Concentrated solutions of methylene blue increase the reduction period.
• The incubation temperature influences the test result due to the temperature
great for the growth of various microorganisms.
• The redox potential of aseptically obtained milk is much lower than
that the frothed milk, being capable of reducing almost
instantaneously the methylene blue.
MATERIAL:
 Sterilized test tubes;
Rubber stoppers;
Sterile pipettes of 10 and 1 ml;
Cold water bath;
Steam bath at 40oC;
Bain-marie or oven at 37oC.
TECHNIQUE:
• Add 1 mL of methylene blue to each tube to be analyzed;
• Add 10 ml of milk and close the tube with a rubber stopper, labeling it.
with respect to the sample.
• Keep the tubes in an ice water bath until the operation is complete, which does not
It should exceed two hours, due to the tendency for fat to increase;
• Transfer the tube holder with the samples to a water bath at 40.oC for what
the temperature reaches 37oIn 5 minutes;
• Invert the tubes several times for a complete mixing of dye with
milk and transfer to a water bath or incubator at 37oC;
• Time the start of the testing period and invert the tubes every 30 minutes.
for the redistribution of bacteria and fat;
• Observe and note the tubes where there is discoloration of the methylene blue.
corresponding to 80% of its volume;
• Classify the milk according to the bleaching period.

CLASSIFICATION:
Time in hours Class

5 Great
4 Good
3 Regular
2 Bad
1 Very bad
Expected result for good quality milk: No discoloration of the methylene blue.
until the first 90 minutes.

NOTES:

High quality milk can be tested once a month, while lower quality milk can be tested more frequently.
Quality requires more frequent testing due to its greater variation in quality.
microbiological. The production of unhygienic milk, the use of dirty utensils, lack of
refrigeration and elevated temperature during storage, handling and transportation.

Table - Relationship of the number of bacteria present in milk x time of reduction of blue
methylene

Time of reduction Approximate number of bacteria per ml

Less than 20 minutes Above 20,000,000
20 to 120 minutes 4,000,000 – 20,000,000
2 to 5 ½ hours 500,000 – 4,000,000
5 ½ to 8 hours 100,000 – 500,000
Above 8 hours Less than 100,000
11 - ADDITION OF HYDROGEN PEROXIDE

Place in a test tube, 2 mL of sample, 2 mL of concentrated hydrochloric acid (HCl) and

a drop of diluted formalin solution 10% w/v.

Heat to 60oIn a thermostatic bath (water bath).

result The presence of hydrogen peroxide is confirmed by the development of

blue-violet coloration.

12 - ADDITION OF NEUTRALIZER
Place in a test tube, 5 mL of sample, 5 mL of 65% ethyl alcohol, and 5 drops of acid.
rosolic (or aurin) at 1%.

RESULT: The addition of neutralizing substances is confirmed by the

development of rosacea coloration.

13 - DENSITY RECONSTITUENTS RESEARCH

Reconstituting substances of milk are added with the purpose of covering up fraud.
by dilution. As water decreases the density of milk, certain substances are added to it
milk to restore its normal density.
NOTE: make blank tests for comparison.

TECHNIQUE

1. Starches:
Transfer 10 mL of the sample to a test tube. Boil in a Bunsen flame.
from the Bunsen or alcohol lamp and cool in running water (to avoid the
iodine evaporation). Add 5 (five) drops of Lugol.
The appearance of blue or greenish color indicates the presence of starch.
Lugol - 5 g of iodine and 10 g of potassium iodide are dissolved in 100 mL of
water.

2. Chlorides:
In the test tube: 2 mL of sample, 2 mL of reagent "A", 2 mL of reagent
B
Yellow coloration: positive for chlorides.
Brown coloration: negative for chlorides (this coloration is observed after +/-
30 minutes
Reagent "A": 10 g of calcium carbonate dissolved in 100 mL of solution
aqueous 0.5% potassium chromate.
reagent 'B': 0.74% aqueous solution of silver nitrate.

3. Sugars (sucrose):
In a test tube add 2 mL of milk, 2 mL of HCl (muriatic acid).
Stir until dissolved and leave in a water bath (80oC) for 2-3 minutes.
 Observe if darkening occurs during this time. Brown coloration:
positive for sugars.

Table - Milk density correction according to temperature

Grouse
lactodensimetric Milk TemperatureoC)
(reading)
10 11 12 13 14 15 16 17 18 19 20
195 189 190 191 192 193 195 196 198 200 202 204
200 193 194 195 196 198 200 201 203 205 207 209
205 198 199 200 201 203 205 207 209 211 213 215
210 203 204 205 206 208 210 212 214 216 218 220
215 208 209 210 211 213 215 217 219 221 223 225
220 213 214 215 216 218 220 222 224 226 228 230
225 218 219 220 221 223 225 227 229 231 233 235
230 223 224 225 226 228 230 232 234 236 238 240
235 228 229 230 231 233 235 237 239 241 243 245
240 233 234 235 236 238 240 242 244 246 248 250
245 238 239 240 241 243 245 247 249 251 253 255
250 242 243 245 246 248 250 252 254 256 258 260
255 247 248 250 251 253 255 257 259 261 264 266
260 252 253 255 256 258 260 262 264 266 269 271
265 257 258 260 261 263 265 267 269 271 274 277
270 262 263 265 266 268 270 272 274 276 279 282
275 267 268 270 271 273 275 277 279 281 284 287
280 271 272 274 276 278 280 282 284 286 289 292
285 276 277 279 281 283 285 287 289 291 294 297
290 281 282 284 286 288 290 292 294 296 299 302
295 286 287 289 291 293 295 297 299 301 304 307
300 290 292 294 296 298 300 302 304 306 309 312
305 295 297 299 301 303 305 307 309 312 315 318
310 300 302 304 306 308 310 312 314 317 320 323
315 305 307 309 311 313 315 317 319 322 325 328
320 310 312 314 316 318 320 322 324 327 330 333
325 315 317 319 321 323 325 327 329 332 335 338
330 320 322 324 326 328 330 332 334 337 340 343
335 325 327 329 331 333 335 337 339 342 345 348
340 329 331 333 335 338 340 342 344 347 350 353
Source: Analytical Standards of the Adolf Lutz Institute Volume 1.
Example: If the read value is 300, it means that the density result is 1.030 g/cm3you
1030 kg/m3.