EXPERIMENTAL SESSION ORGANISATION · 2- Repeat with adding a drop of Sudan red III and compare both...

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EXPERIMENTAL SESSION ORGANISATION

3 groups :

GROUP 1 GROUP 2 GROUP 3

Carnicella Federica Caterina Corrieri Paola de Tullio Cristina, Lucrezia Introna Maddalena Baus Lukas Gadomski Bastian Förg Marie Sophie Nagel Kirsten Starostka Kinga Zamkotowicz Tomasz Woźniak Anna Handziuk Zuzanna

d'Ingeo Daniele, Domenico de Bari Fabio Sergio Mastrofilippo Cosimo Michele Scuccimarro Annamaria Rurainsky Ines Schmengler Heiko Schneider Mareike Schönershoven Lukas Männicke Lucie Bobrowska Anita Dyczek Katarzyna Kalarus Zuzanna Petka Rafał

la Grasta Giovanni Lucanie Marco Minervini Annamaria Porta Maria Gabriella Schüller Laura Stephan Eric Wieghardt Julia Lochmann Joel Eitelbach Florentine Grzywnowicz Agata Jakubowska Marta Kościelniak Agnieszka Wolny Monika

WINE ROOM BREAD ROOM CHEESE ROOM

Sonia, Malcolm, Inès, Nasma

Maya, Joanna, Fatou

Celia, Igor, Nancy, Anaïs

Saturday, April 21 8h30-11h30

Group 1 Group 2 Group 3

Sunday, April 22 14h-19h


Monday, April 23 8h-11h


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Equipment :

- Flour - Bread - Beaker - Copper sulphate - Sodium hydroxide

- 100°C Water bath - Distilled water - Slide cover slips - Fehling solution - Iodine water

- Pipettes , pipettes filters - Spatula - Proteins : Egg white - Glucose - 10 Test tubes

� Test the presence of starch, reducing sugars, prote ins, in flour and bread. 1- Carry out a test on each. 2- Carry out control tests

Reactant Detected compounds Protocol

Fehling-solution

Reducing sugars (glucose, maltose, lactose …)

1. In a test tube, pour 3mL of the product to be tested. 2. Add 2mL of Fehling-solution. 3. In the water bath, warm until boiling.

Biuret reaction Proteins

1. In a test tube, place a few drops of the product to be tested. 2. Pour a few drops of copper sulphate. 3. Cover with a sodium hydroxide solution 4. Wait for a few minutes.

Iodine solution Starch 1. Pour some product to be tested on a slide cover slips 2. Add a few drops of iodine solution.

Control test flour bread

Reducing sugars

Proteins

Starch

Comenius Project – Marseille

BREAD ROOM

Comparative study : flour – Bread.

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Equipment: - 10g/L Yeast Suspension, deprived

of nutrients and oxygen; - 10g/L Glucose solution; - 10g/L Starch solution;

- 10g/L lactose solution; - Little syringe with catheter; - 1 beaker ;

- Computer assisted experimental device (“ExAO” in French) with O2 sensor, éthanol sensor and CO2 sensor.

� Using the EXAO device, measure the O2/ ethanol/CO2 concentration evolution in a medium containing yeast suspension:

1- Follow the instructions of bioreactor and sensor installation; 2- Sample 30 mL of yeast suspension in the first two thirds (to avoid a big concentration of yeast) ;

3- Fill the bioreactor eliminate all air bubbles ; 4- Close it to avoid any exchange with outer air; 5- Start the agitation.

� Carry out measuring : 1- Open the software « atelier scientifique » and select the sensors to use ; 2- Enter parameter for a 5-minute measuring; 3- Click start ; 4- After 1 minute, put 1 mL of glucose solution in the bioreactor with a syringe.

� Carry out the same experiment, using the other solu tion. � Rinse the bioreactor after each measurement ; � Superimpose the curves

� Describe the results and conclude

Computer assisted experimental device (“ExAO” in French)

Yeast ( electron microscope)


BREAD ROOM

Quantitative study of yeast metabolic activity

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Equipment :

- 6 50-mLvolumetric flasks - 12 test tubes + 2 racks - 50 g/L yeast suspension - 37°water bath - 100°C water bath - electric heating device

- clamp - 15 rubber balloons - wheat flour - distilled water - 1 packet of baking powder - 1 electronic balance

- Glucose - starch - Lime water - Graduated cylinder - potassium bichromate with acid

� Carry out the following montages in order to deter mine the nature of the gazes liberated by the yeast: 1- Take 5 volumetric flasks ; 2- Fill them as follows and then shake :

� volumetric flask 1: Flour: 2g + yeast suspension : 40 mL � volumetric flask 2: Flour : 2g + water : 40 mL � volumetric flask 3: yeast suspension : 40 mL � volumetric flask 4: Glucose : 2g + suspension of yeast : 40 mL � volumetric flask 5: starch : 2g + yeast suspension : 40 mL

3- Place a balloon over each flask in order to collect the gases. Place them in a 37°C water bath and wait � Test the kind of gases of the flasks : place the balloon on a tube previously filled with 2mL of lime water so

that the gas can react; � Test the alcohol presence in the test solution :

� pour 3 mL of each volumetric flask solution, add 3 mL of potassium bichromate and wait for a few minutes in a 100°C water bath

� carry out the same test with 3mL of alcohol instead in order to compare

� Compare baker yeast and chemical yeast 1- Take 4 test tubes 2- Fill them as follows and then shake:

� 2 Tubes 1 : 5g of chemical yeast � 2 Tubes 2 : 10 mL of yeast suspension + 2g of glucose

3- Cover one tube « 1 » and one tube « 2 » with a balloon. Place in a 37°C water bath and wait 4- Cover one tube « 1 » and one tube « 2 » with a balloon. Heat with electric heating device 5- Test the nature of the gases in the tubes : place the balloon on a tube previously filled with 2mL of lime water

so that the gas can react;



BREAD ROOM

Qualitative study of yeast metabolic activity

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Principle:

To make some cheese, the first step consists in making the milk curdle. For that purpose, several techniques are used. One of them consists in decreasing the pH ; we can wonder about the nature of the molecules which coagulate.

Milk is a complex food ; we suggest looking for the molecules which are present in curd and in whey. Equipment:

- Milk curd - Milk - Beaker - Spatula - Test tube - Glass rod

- Sudan red III - Drainage system :

funnel + filter + Erlenmeyer flask

- Copper sulphate - sodium hydroxide

- Water bath in 100°C - Distilled water - Acetic acid - Microscope - Slide cover slips - Fehling solution

- Ammonium oxalate - Iodine water - Pipettes , pipette filters - Calcium ions - Proteins : Egg white - Glucose

� Study the milk under the microscope. 1- Prepare a microscopic preparation of milk and observe it. 2- Repeat with adding a drop of Sudan red III and compare both preparations.

� Filter the curd milk in an Erlenmeyer flask, using a funnel and filter paper. Place the aggregate from the filter on a beaker cover .

� Test the presence of reducing sugars, proteins, lip ids and calcium in the whey and the aggregate.

1- For that purpose, divide the whey and aggregate into 4 parts. 2- Carry out a test on each part. 3- Carry out control tests

Reactant Detected compound Protocol

Fehling-solution

Reducing sugars (glucose, maltose, lactose …)

1. In a test tube, pour 3mL of the product to be tested. 2. Add 2mL of Fehling-solution. 3. In the water bath, warm until boiling.

Biuret reaction Proteins

1. In a test tube, place a few drops of the product to be tested. 2. Pour a few drops of Copper sulphate. 3. Cover with a sodium hydroxide solution 4. Wait for a few minutes.

Ammonium oxalate Calcium Ions 1. In a test tube, pour 3mL of the product to be tested. 2. Add a few drops of Ammonium oxalate.


CHEESE ROOM

Constituents of milk and coagulation

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Board of results

Control tests Whey Aggregate

Reducing sugars

Proteins

Calcium

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Principle:

Sugars are separated by an absorption chromatography The stationary phase is made up of silica gel; the mobile phase is a single phase mixture of 3 solvents with different polarities: methyl ethyl ketone, acetic and methanol acid. They make up spots, revealed by the potassium permanganate associated to soda ash.

Each sugar is characterized by its migration distance within the mobile phase. Equipment

- 5g/L Glucose control solution - 5g/L Lactose control solution - 5g/L Galactose control solution - 5g/L Fructose control solution - Toothpicks - Hair-dryer

- Solvent : 3 volumes of methyl-ethyl-ketone, 1 volume of acetic acid and 1 volume of methanol ;

- 10 x 5 cm silica gel chromatographic plates

- a chromatography bath holder

- Developer : 50 mL of 2% potassium permanganate solution and 50 mL of 4% soda ash solution

- Ventilated hood - Big claw - Bawl for developer

� Preparation of the bath holder

1- Work under the ventilated hood 2- In the bath holder, pour the solvent to 1 cm height, so that the level remains below the line drawn on the

support plate and wait for saturation (about 10min).

� Assessment of control solution deposits, milk and y oghurt 1- With a pencil draw a line 2cm from the lower edge of the support plate 2- On this line slightly mark the deposit locations every 0,8cm, starting from 0,4cm from the edge. 3- Write a number under each point 4- Place drops of each solution over each point (3-step operation):

• Sample some solution with the toothpick ; The deposits should not exceed a 2 to 3mm diameter;

• Place it in 3 steps on the support plate, drying the deposit with the hairdryer between each step; • Write down the number of the deposit corresponding the solution which is being studied ; • You should use each toothpick only one time.

� Migration and visualization:

1- Put the support plate in the bath holder in a diagonal position; 2- Cover it immediately with the lid 3- Let it migrate until the solvent front reaches the upper edge of the support plate; 4- Take the support plate and dry it, plunge it for 10sec in the developer, then dry it.

� Describe and conclude


CHEESE ROOM

Sugar thin-layer liquid chromatography

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Equipment :- Yogurt - Roquefort - Milk - Spatula - Slides and cover slips

- Needle - Microscope + camera - Methylene blue : dye marking the

cellular components - Pipettes

- Water - Beaker cover - Absorbent paper

1- Yogurt microscopic observation. • Pour 1mL of yogurt supernatant in a test tube • Add a few drops of methylene blue • Sample a drop and prepare a microscopic slide • Observe and scan

We can observe two forms of microorganisms :

- Streptococcus with spherical form or in colony - Lactobacillus with stick form

2- Milk microscopic observation

• Proceed the same way 3- Roquefort microscopic observation.

• Sample some blue part with a niddle. • Prepare a microscopic slide • Observe and scan

We can observe myceliums and spores of Penicillium roqueforti 4- write down the caption over the corresponding p ictures

(optical microscope):

(optical microscope):


CHEESE

Microorganism observation

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Milk contains sugars. The presence of bacteria in milk provokes the conversion of part of these sugars in lactic

acid. The freshness of milk is assessed by titrating its acidity with a sodium hydroxide solution.The titration result is

expressed in Dornic degrees (°D).

1 Dornic degree corresponds to the presence of 0,1g of lactic acid equivalent in 1L of milk Fresh milk usually titrates from 15 to 20°D

We wish to compare the acidity of various milks: fresh cow milk, curdy milk and ewe or goat milk. To get a rigorous comparison, we need to perform a precise titration in two steps. → 1st step – approximate titration : pour the sodium hydroxide solution in the milk, mL by mL, until the indicator colour changes. Note the value of the added volume (Vb). → 2nd step – accurate titration : after preparing a new solution of milk, directly pour in it a volume of the sodium hydroxide solution equal to (Vb - 2mL) then continue pouring drops of sodium hydroxide, one after the other, until the indicator colour changes. The Ve volume of sodium hydroxide poured to get the equivalence is a more accurate.

� EXPERIMENTAL PROTOCOL

Equipment and products: - 1 goblet - 4 beakers - 1 25mL graduated burette - 1 magnetic stirrer - 1 20mL calibrated pipette

- 1 Erlenmeyer flask - a definite Cb concentration NaOH

solution - 1 propipette - Phenolphtaleine

- distilled water - fresh cow milk or curdy milk or

ewe/goat milk

For each titration, we will use a reference control equal to the test solution to better assess the change of the indicator color. The indicator we'll use is phenolphthalein: which is colourless in an acid solution and turns fuchsia in a basic solution.

1- Pour the milk you intend to proportion in a beaker 2- Then measure out a Va=20mL volume of this milk by using

a graduated pipette equipped with a propipette. Place it in an Erlenmeyer flask.

3- Add a few drops of phenolphthalein 4- Fill the burette with NaOH solution 5- Perform the approximate and accurate titrations of the milk

which is on your lab table The titration result is reported in the following table (II,1)


CHEESE

Acidity titration in milk

NaOH solution, Cb =

Va = 20mL milk + few drops of phenolphthalein

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� Analysing the results

1- Record in the table below � Vb volumes poured to equivalence for the different milks

� The lactic acid concentration computed with :

� The titer , t, obtained with the relation , t = M.Ca with M=86g .mol-1

� The titer value in Dornic degree

Cow milk 1 Cow milk 2 Goat or ewe milk

Ve (mL) Equivalent volume

Ca (mol.L -1) Lactic acid

concentration

t (g.L -1) Titer

Titer in Dornic degree ( °D)

2- Are all titrated milks all edible? 3- Classify the different milks by increasing acidity

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The ferric casse can happen in wines when there is an excess of iron. In white wines, it's the white casse, due to

insoluble combinations between phosphoric acid and the iron in excess. In the red wine, it's the blue casse, which leads a turbidity due to of insoluble combinations between phenolic compounds (tannin, anthocyanins…) and the iron in excess. The risk is important when the iron concentration exceed 10 or 15mg/L. The iron present in wines can be of biological origin (brought in by the vine that took it from the earth), but it is mainly of technological origin ( (concrete vat wine making, equipment in oxided metal ...)

Principle of dosage

We perform a colorimetric dosage as the color of a solution depends of the concentration of the coloring species. The species to be titrated is the Fe³⁺ ion which creates a complex ion with the SCN⁻ thiocyanate ion while giving

a red blood color mixture. To have a total quantity of iron ion, we must oxide the iron II ions of the solution into iron III ions with using perhydrol .

We carry out a scale of tints , then we determine the concentration of the solution to be titrated first by comparing, then by carrying out a spectrophotometric titrate Equipment

- 2 25mL burettes - 10 test tubes - 6 plugs - 2 5mL graduated pipettes - 1dropping pipette - 2 20mLand 10mL calibrated pipettes - 4 beakers

Product - discolored white wine - S0 stock solution : t0=0,02g.L-1 - 2 mol.L sulfuric acid - 2mol.L solution of potasium thiocyanate - 20-volume perhydrol - distilled water

� PREPARATION OF THE SCALE OF TINTS 1- Fill in one burette with So solution and the other with distilled water. 2- Fill in each test tube using indications of the following chart. Number them. The total volume of solution in each test tube must be 10mL. 3- Complete the chart.

Tube 1 2 3 4 5

V₀ of S₀ (mL)

0.5 1.0 2.0 3.0 4.0

V₁ of distilled water (mL)

t(Fe³⁺) (mg.L ⁻¹)

The mass titer of each solution is calculated using the relation :

t = avec t0 = 0,02 g.L-1


WINE ROOM

Ferric casse

t0 x V0

(V0 +V1)

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4- In number 6 test tube, introduce 10mL of discolored wine, 5- In each tubes , add 1,5mL of sulfuric acid and 1mL of potasium thiocyanate 6- Pour 10 drops of perhydrol and homogenize .

The scale of tints is ready. � DETERMINATION OF THE CONCENTATION IN IRON(III) ION IN WINE

Determine by comparison the mass concentration in iron (III) of the discolored wine in the test tube n°6 � SPECTROPHOTOMETRIC TITRATING

The spectrophotometric titrating allows to get the exact value of the concentration The more colored a solution is, the more light it absorbs.

The absorbance of a solution is proportional to its concentration.

Beer-Lambert law A=εεεε(λ).e.C

εεεε(λ) is the coefficient of molar extinction e is the thickness of the solution through which the light passes C is the concentration of the studied solution We measure the absorbance of all prepared solution. We select the 480nm wavelength, which corresponds to the maximum absorption of the studied color

• On the desktop log on the « spectro CCD » software • Adjust the wavelength • Setting zero: in the spectrophotometer add an absorption cell filled with distilled water ; the absorbance is

equal to zero. • Then measure the absorbance of all prepared solution

Test tube 1 2 3 4 5

T(mg.L ⁻¹)

A

1- Draw the absorbance in relation with the solution titer. 2- Measure the absorbance of the prepared solution from wine. 3- Using the pointer, position and determine the content in iron (III) ions 4- Check then print 5- Is wine safe ?

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The red cabbage owes its color to the presence of a natural pigment which belongs to the anthocyanin family. The

anthocyanins change color depending on the environment's acidity , this pigment can be extracted by a decoction. It will be used as a pH-meter.

Equipment - 1 burette 25 ml - 1 stir plates - white vinegar - 5beakers 50 ml - 1 wooden clamp - 0,1 mol.L sodium hydroxide - 1 beaker 400 ml - 1 heat resistent glove - Britton Robinson solution

- 1 beaker 250 ml - filter paper - pH-meter and pH7 and 4 buffer

solutions - 1 erlenmeyer flask - funnel cup - 1 dropping pipette - 1 calibrated pipette 20 ml - glass rod

- 1 propipette - felt tips pens - 10 test tubes - 1 red cabbage - 5 plugs - discoloured wine - 1 goblet - discoloured grape juice

I. Preparation of the decoction

1- In a 400mL beaker, introduce a cut of red cabbage leaf with 100mL of distilled water 2- Let it boil on a stir plate for 5min, while stirring with a glass rod 3- During the heating prepare the filtration material 4- After the boiling, percolate the beaker's content 5- We get a purple filtrate. Later on we'll call this solution Sc

II. Preparation of the pH scale

1- Fill the graduated burette with 0,1 mol.L sodium hydroxide 2- Pour in the seven 20ml test tubes the Britton Robinsin solution, with a propipette

The Britton Robinson solution is an acid mixture whose pH varies by the addition of a V volume of 0,1mol.L sodium hydroxide solution, according to the relation:

pH= 1,2+Vb with Vb in mL

3- Fill in the third line of the chart

4- Pour in each test tubes the Vb volume indicated in the chart 5- Add in each test tube 10 drops of Sc solution

6- Fill in the fourth line indicating the solution colour for each test tube.


WINE ROOM

Wine pH assessment and pH measure

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Tube 1 2 3 4 5 6 7

pH 1,2 2 3 4 5 6 7

Vb (ml)

Colour

III. pH assessment

1- In 3 test tubes , pour 20ml of grape juice , wine and vinegar measured with a graduated cyclinder. 2- Add 10 drops of Sc solution 3- Fill in the following chart

Grape juice Wine Vinegar

Colour

pH

IV. pH measure

1- Measure the 3 product pH with a calibrated pH-meter 2- Complete the following chart

Grape juice Wine Vinegar

pH

� Compare the results (assessment – measure) and conc lude

EXPERIMENTAL SESSION ORGANISATION · 2- Repeat with adding a drop of Sudan red III and compare both...

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