4. Spectrometric determination of phosphates
in apple juice

EXERCISE

Determine if the phosphate concentration in apple juice is between the permitted RSK values (100 – 350 mg PO₄^3-/L).

PRINCIPLE OF THE METHOD

Phosphates react with molybden ions in acid medium to phosphomolybdate, which is reduced with ascorbic acid to molybdic-blue. The final solution is photometrically measured at 720 nm.

MATERIALS AND REAGENTS

Spectrophotometer Spektra^TM

Measuring flasks: 50 in 100 mL

Pipettes: 2 mL, 4 mL, 20 mL, 25 mL

Pipetor 200 μL

Measuring cylinder

Ammonium heptamolybdate
(NH₄)₆Mo₇O₂₄·4 H₂O

Disodium-hydrogenphosphate
(Na₂HPO₄·12 H₂O)

L-ascorbic acid

Hydrochloric acid (2 mol/L)

Sulphuric acid (1 mol/L)

Phosphate stock solution P
(g = 1000 mg/L)

SAMPLE PREPARATION

Transfer 25 mL of juice into an incinerating dish, evaporate on the water bath and ignite the residual at 550°C, until white ash appears. Add 2-3 mL of hydrochloric acid and transfer to the 50 mL measuring flask. Quantitative transfer is achieved by rinsing with distilled water. Fill the measuring flask with distilled water to the marker line and mix well.

SAMPLE PREPARATION FOR PHOTOMETRIC DETERMINATION

Reagents (Pipeted in 100 mL measuring flask)	Sample	Standard low– Standard 1	Standard high– standard 2
Ash solution	5 mL	-	-
Phosphate stock solution	-	100 µL	300 µL
Distilled water (cca.)	50 mL	50 mL	50 mL
Sulphuric acid	20 mL	20 mL	20 mL
Ammonium heptamolybdate	4 mL	4 mL	4 mL
Ascorbic acid	2 mL	2 mL	2 mL

Heat opened measuring flask on water bath for 15 minutes, cool it down and fill it with distilled water to the marker line. Pipet 350 µL of well mixed solution into the blister and measure transmittance photometrically using the red LED.

EVALUATION

Measurements (transmittance readings) are signed in the table and absorbances are calculated.

	T (%)	A	gPO₄^3- (mg/L)
Standard 1			1.00
Standard 2			3.00
Sample

g_sample = A_sample . g_standard . 122.6 / A_standard

Result and final conclusion:........................................................................

SOLUTION PREPARATION

Ammonium heptamolybdate (NH₄)₆Mo₇O₂₄·4 H₂O (g =2 g/100 mL)

Dissolve 2.0 g of ammonium heptamolybdate in 60 mL of hot (60°C) destilled water. Fill in the 100 mL measuring flask with destilled water to the marker line.

Disodium-hydrogenphosphate (Na₂HPO₄·12 H₂O) (g =11.5000 g/L)

Dissolve 11.5000 g of disodium-hydrogenphosphate in destilled water and fill the 1000 mL measuring flask to the marker line.

L-ascorbic acid (g =0.353 g/100 mL)

Dissolve 0.353 g L-ascorbic acid in destilled water and fill the 100 mL measuring flask to the marker line.

Hydrochloric acid (2 mol/L)

Sulphuric acid (1 mol/L)

HAZARDS

	Ammonium heptamolybdate (NH₄)₆Mo₇O₂₄·4 H₂O R22, R36-38, R40
	Disodium-hydrogenphosphate Na₂HPO₄·12 H₂O R 36/37/38
	Hydrochloric acid R34, R37
	Sulphuric acid R35

Developed and prepared by:: Alenka HMELAK GORENJAK, Živilska šola Maribor

4. Spectrometric determination of phosphates in apple juice