Analytical chemistry IV

ACHM412 DETERMINATION OF ASCORBIC ACID

Determination of ascorbic acid | Aadil khan 21416042

Title:

The determination of ascorbic acid (vitamin c) in fruit juices using cyclic voltammetry and
differential pulse polarography.

Aim:
To perform the analyses of fruit juices to determine ascorbic acid (vitamin c) in fruit juices
and exposure to technique of voltammetry.

Theory & Intro:

Ascorbic acid (AA) because of its antioxidant and therapeutic properties, is a valuable food
component. Unfortunately, it is also a very labile substance which easily undergoes
degradation caused by enzymes liberated from the raw material during technological
processes, too high temperature or access of air. The losses of vitamin C during
technological procedures forming part of the processing of juices can exceed even 50%
That is why manufacturers of food products bearing in mind the need to replenish vitamin C
losses caused by the technological treatment and also to enhance the nutritive value of the
product characterized by low content of this vitamin
or to extend the shelf-life of food products – frequently add vitamin C to them.
It is worth emphasizing that, in some cases, excessive quantities of AA may result
in the inhibition of natural processes occurring in food and, consequently, contribute to
the deterioration of its taste

Ascorbic acid is easily oxidized to de-hydro ascorbic acid, this makes electrochemical
methods a useful method in determining Vitamin C levels in foods. Electrochemical methods
have important applications in organic and inorganic synthesis and in sample analysis. This
method is fast, sensitive, selective and gives linear response at low concentration range.
The dropping mercury electrode (DME) surface used is a vital part of the
determination. In this investigation, we use voltammetry in the determination of
ascorbic acid levels in Liqui fruit and Ceres juice samples.
 OH

O
O
HO

HO OH

Structure of Ascorbic Acid

Reagents & Apparatus:

 Oxalic acid solution

 1.4000g oxalic acid di-hydrate is dissolved in distilled water (made up to 1L)

 Acetate buffer pH = 4.64

 8.2g of sodium acetate, dissolved in distilled water followed by addition of 6 mL glacial

acetic acid, made up to 100 ml with distilled water.

 500 mg/L Vitamin C standard, ρ Vitamin C

 50.0 mg Vitamin C is dissolved in oxalic acid solution and made up to 100 mL 1 mL

standard corresponds to 0.5 mg Vitamin C

 Grape juice sample

 Orange 100% fruit juice blend sample

Working Parameters:
Working electrode DME / HMDE

Stirrer speed 2000rpm

Mode DP

Purge time 300 s

Cleaning Potential -0.1V

Cleaning time 0

Deposition potential -1.15V

Deposition time 90 s

Equilibration time 10 s

Pulse amplitude 0.5 V

Start potential -1.2V

End potential -0.1 V

No. Of cycles 0

Sweep

Start Potential -0.1999V

End Potential 0.1999V

Pulse amplitude 0.05V

Pulse Time 0.04s

Voltage step 0.005951V

Voltage step time 0.5 s

Sweep rate (mV/s) 0.0119V/s

Peak potential Cd -5.8 V

Peak potential Cu -0.6 V

Table 1: Working parameters of the VA instrument

Schematic representation of the experimental setup

 Method:
 Electrolyte: Conc. (acetic acid)
 Working electrode (WE): MME (Multi Mode Electrode)
 Auxiliary electrode (AE): Pt
 Reference electrode (RE): Ag/AgCl/KCL (3mol/L)

Procedure:
Dissolved 5g of oxalic acid with deionised water up to 1 litre
Sodium acetate solution containing 10 % deionised water was prepared in the
following way:
40.9965 grams of sodium acetate with 30 ml glacial acid and made up to 500ml
deionised water.
Vitamin c standard was prepared (vitamin C) = 0.5g/L, 50 mg of vitamin C was
dissolved in oxalic acid solution and made up to 100ml. The solution had to be
prepared freshly. (0.5 mg of vitamin C equates to 1 ml standard)

I. Polarographic determination of ascorbic acid in grape and orange fruit juices

15 mL of sodium acetate buffer + 1 mL juice, measurement was done using

standard additions.
Voltammetry was carried out using 797 VA Computrace.

A calibration curve of vitamin C was prepared by diluting a 500 mg/L stock solution
of 300 µL-ascorbic acid in a15ml oxalic acid solution and 1ml pH 4.64 acetate
buffer. Calibration solutions were degassed by bubbling nitrogen for 300s prior to
each voltammetry experiment

C6H8O6 + H20 → C6H6O6 + 2H+ + 2e-

 Analyses of 4 sample were done separately using dropping mercury electrode on
797 VA Computrace and the i/E curves were obtained.

Requirements:

Actual masses used

Oxalic: 0.7g
Sodium acetate: 41g
Acetic Acid: 30ml

Oxalic acid standard preparation:

0.72 𝑔
𝑚𝑜𝑙𝑒𝑠𝑜𝑥𝑎𝑙𝑖𝑐 𝑎𝑐𝑖𝑑 =
126.07 𝑔. 𝑚𝑜𝑙 −1

= 5.711 × 10−3 mol

5.711 × 10−3
𝑐𝑜𝑛𝑐𝑜𝑥𝑎𝑙𝑖𝑐 𝑎𝑐𝑖𝑑 = × 126.07 𝑔. 𝑚𝑜𝑙 −1
500 × 10−3 𝐿
= 1.44 𝑚𝑔. 𝐿−1

0.72 g oxalic salt was weigh to a 500-mL volumetric flask and made up to the mark
with deionized water

Ascorbic acid standard preparation:

0.05 𝑔
𝑚𝑜𝑙𝑒𝑠𝑎𝑠𝑐𝑜𝑟𝑏𝑖𝑐 𝑎𝑐𝑖𝑑 =
176.13 𝑔. 𝑚𝑜𝑙 −1

= 2.8388 × 10−4 mol

2.8388 × 10−4 𝑚𝑜𝑙

𝑐𝑜𝑛𝑐𝑎𝑠𝑐.𝑎𝑐𝑖𝑑 = × 176.13 𝑔. 𝑚𝑜𝑙 −1
500 × 10−3 𝐿
= 0.1 𝑚𝑔. 𝐿−1
50 mg ascorbic salt was weighed to a 100-mL volumetric flask and made up to the
mark with the oxalic acid standard prepared above Ascorbic acid standard was
analyzed using cyclic voltammetry and differential pulse polarography.

Discussion

It was noted that ascorbic acid is affected by a change in pH levels. Reduction is

favoured with a more basic solution.

This was observed (polarograms attached in appendix) when ascorbic acid with a pH
of 8.6 and only two peaks (not well resolved), thus we rejected the pH 8.6.

At a pH of 2.6 it was observed that the three peak overlaps and the voltage showed a
more positive value indicating a drop in the readings, which tells us that there is
oxidation taking place.

The optimum pH of 4.6 was selected for ascorbic acid for the quantification of vitamin
C in fruit juices since there was a steady increase in current after each injection of the
ascorbic acid std.

The percentage recovery from the orange juice and grape juice was 88.04% and
91.8% respectively.

Possible reasons for differences and inaccuracies include experimental errors, cross
contamination and technique errors with the injection or other errors made by the
analyst that injected the sample.

A change of analyst means that the specific style of the process of injecting the sample
would be changed, introducing a new variable to the experiment that could affect the
results.

The actual amount of vitamin C in apple, orange and grape juice was 41. mg and
22.6mg in the experiment we found 36.91mg and 20.72 mg respectively. If a closer
look at the polarogram of orange juice you will notice an uneven curve which caused
a lower value in the percentage of vitamin C and due to experiment errors

Conclusion

The developed voltammetric methods (differential pulse voltammetry and cyclic

voltammetry) for ascorbic acid determination are characterized by sensitivity, rapidity
and reproducibility. The degree of accuracy of the investigated voltammetric methods
is confirmed by the values
The linear character of the plot of peak currents (Ip– Itp) versus the ascorbic acid
concentration obtained for the ascorbic acid systems in the solution of the
supporting electrolyte supplemented with the citric acid indicates the possibility of
application of the cyclic voltammetry to assess vitamin C concentration.
.
The degree of accuracy of the investigated voltammetric methods is confirmed by
the values obtained for the degree of recovery, when compared to the accurate
values labelled on the fruit juices.

These experimental results suggest that because of the ease of use, accuracy,
minimal sample preparation, and rapid detection, voltammetry is a superior method
for vitamin C detection.

REFERENCES

1. Erdurak-Kiliç, C.S.; Uslu, B.; Dogan, B.; Ozgen, U.; Ozkan, S.A.; Coskun, M. Anodic
voltammetric behavior of ascorbic acid and its selective determination in pharmaceutical
dosage forms and some Rosa species of Turkey. J. Anal. Chem. 2006, 61, 1113-1120.
2. Popa, C.V.; Danet, A.F.; Jipa, S.; Zaharescu, T. Determination of total antioxidant activity
of wines using a flow injection method with chemiluminescence detection. Rev. Chim.
(Bucharest) 2010, 61, 11-16.
3. Pisoschi, A.M.; Cheregi, M.C.; Danet, A.F., Total antioxidant capacity of some
commercial fruit juices: electrochemical and spectrophotometrical approaches,
Molecules 2009, 14, 480-493.
4. Papuc, C.; Pop, A.; Serban, M. Metode Analitice in Biochimia Veterinara; Editura
Printech, Bucharest, Romania, 2001; pp. 167-169.