TITLE

Quantification of Sodium Benzoate in Food

OBJECTIVE

To measure the quantity of sodium benzoate in food.

INTRODUCTION

Food and beverage spoilage has been a problem throughout history. Most

food spoilage is due to enzyme action upon food. Enzymes are complex organic

compounds that may act as catalysts and cause a chemical change to occur. Most

enzymes that cause food to spoil are produced by living organisms, e.g., bacteria,

molds and yeast. Fresh foodstuffs may have some of these microorganisms present

and others may be encountered by exposure to the air or in processing. The

enzymes that cause spoilage may be generally described as two types;

“endoenzymes” which exist within the microorganism and “exoenzymes” which are

released by the microorganism. Consequently, the quality of microorganism

enzymes of both types which cause food to spoil is directly related to the

concentrations of the microorganism present, its species, and its general activity.

The addition of chemical preservatives to food is not new and been practiced

for centuries. Some of the most familiar preservation methods, those of brining,

pickling with vinegar, smoking, and preserving with sugar solutions, depend upon

chemical preservatives. These methods inhibit microorganism activity and retard

microorganism growth and multiplication. These methods act in one of two

generalized way is by physically increasing the density of the microorganisms

environment or chemically, by a direct inhibiting action on the microorganism

themselves. Consequently, chemical preservatives which perform by a direct

inhibiting action on the microorganisms themselves are not new.

Sodium benzoate is a chemical preservative which in very low concentration

inhibits the activity of the microorganisms themselves. It

is bacteriostatic and fungistatic under acidic conditions. It is most widely used in

acidic foods such as salad dressings (vinegar), carbonated drinks (carbonic acid),

jams and fruit juices (citric acid), pickles (vinegar), and condiments. It is also used

as a preservative in medicines and cosmetics. 

Sodium benzoate is produced by the neutralization of benzoic acid with

sodium hydroxide. Benzoic acid is detectable at low levels in

cranberries, prunes, greengage plums, cinnamon, ripe cloves, and apples. Though

benzoic acid is a more effective preservative, sodium benzoate is more commonly

used as a food additive because benzoic acid does not dissolve well in water.

MATERIALS

Deionized water, Stomacher bag, Whatman No.1 filter paper, micropipette, Nylon

syringe filter (0.45μm), C18 column, sodium acetate, acetic acid glacial, HPLC grade

methanol, micropipette tips, HPLC grade Sodium benzoate.

METHODS

Sample Preparation

1) 5ml of each samples is added to approximately 20 mL of water and then

homogenized on Stomacher for 10 minutes.

2) 35ml of methanol is mixed thoroughly with the mixture.

3) 100mL of water is used to brought up the volume of mixture and filtered through

Whatman paper No.1.

4) The 0.45μm syringe filter is used to filter the samples prior to injection.

HPLC Condition

1) The chromatographic separation was achieved with a C18 column

2) The injection volume was 20μL, mobile phase was 65:35, acetate buffer (pH 4.74)

and methanol with flow rate of 0.8mL/min for 20 minutes at wavelength of 235nm.

Preparation of Standard Curve

1) The external standard plot method was used.

2) Duplicate injections of 20μL sodium benzoate standard solutions were used to

construct linear regression lines (peek area versus concentration).

3) The peaks were identified based on the retention time.

4) The standard curves were obtained with five points (5,10,20,40 and 80mg/L) for

sodium benzoate.

RESULT

A = kc

k = A

c

k = 7.7cm

7.66337 mg/L

k = 1.0478

So, the concentration of sodium benzoate, c = A

k

c = 7.3487mg/L

0.05 mL

c = 146.97mg/L

c =0.14697 mg/ mL

DISCUSSION

In this experiment, the quantity of sodium benzoate in food (soft drink Sprite)

was measured. According the result that has been obtained the concentration of

Sodium Benzoate is 0.14697 mg/ mL. Theoretically the concentration of the sodium

benzoate should be 0.1 mg/ mL. Thus, the result is accepted. The k value is 1.0478.

The value of k is calculated from the formula A = kc. The formula is derived from

the Beer’s Law that is A=ebc, Since eb is a constant, it can be represented as k, and

the equation can be simplified to A = kc. According to the Beer’s Law A is the

absorbance of the sodium benzoate (proportional to the peak height of the sodium

benzoate on the chromatogram.), e is the constant particular to sodium benzoate, b

is the path length of the cell in the instrument (which remains unchanged), c is the

concentration of sodium benzoate. Beer's law  states that the absorbance is directly

proportional to the concentration of a solution. If you plot absorbance versus

concentration, the resulting graph yields a straight line.

HPLC is a chromatographic technique that can separate a mixture of

compounds and is used in biochemistry and analytical chemistry to identify,

quantify and purify the individual components of the mixture. HPLC typically utilizes

different types of stationary phases, a pump that moves the mobile phase(s) and

analyte through the column, and a detector to provide a characteristic retention

time for the analyte. The detector may also provide additional information related to

the analyte. Analyte retention time varies depending on the strength of its

interactions with the stationary phase, the ratio/composition of solvent(s) used, and

the flow rate of the mobile phase. It is a form of liquid chromatography that utilizes

smaller column size, smaller media inside the column, and higher mobile phase

pressures.

A buffer is an aqueous solution that has a highly stable pH. If you

add acid or base to a buffered solution, its pH will not change significantly. Similarly,

adding water to a buffer or allowing water to evaporate will not change the pH of a

buffer. A buffer is made by mixing a large volume of a weak acid or weak

base together with its conjugate. A weak acid and its conjugate base can remain in

solution without neutralizing each other. The same is true for a weak base and its

conjugate acid.

The buffer used in this experiment is acetate buffer. Acetate is a salt or ester

of acetic acid. Salts of acetic acid contain a metal attached to the acetic acid radical

CH3COO. In this experiment, the pH of the acetate buffer used is 4.74. This buffer

can be made using an equal volume of 0.10 M acetic acid, HC2H3O2, and 0.10 M

sodium acetate, NaC2H3O2. Typically the pH can be set to one pH unit above or one

pH unit below the pKa for the acid. The pH used is 4.74 because the pKa of acetic

acid is 4.7. Despite, a buffer solution resists change of pH, water does not. That is

the reason why acetate buffer solution is used instead of using water.

CONCLUSION

At the end of the experiments, the quantity of sodium benzoate in food (Sprite soft drink) was

able to be detected. The quantity of sodium benzoate calculated from the formula is 0.14697

mg/mL.

REFERENCE

Saad, B., 2004. Simultaneous determination of preservatives (benzoic acid, sorbic

acid,methylparaben and propylparaben) in foodstuffs usinghigh-performance

liquid

chromatography. Journal of Chromatography A, 393–397.

Sivasankar, B. (2008). Food Processing and Preservation ,5th ed., Prentice Hall of India

Private Limited, New Delhi.

UNIVERSITI MALAYSIA SABAH

SCHOOL OF FOOD SCIENCE AND NUTRITION

SEMESTER 1, SESION 2011/2012

NT 30703

FOOD SAFETY AND QUALITY CONTROL

PRACTICAL 1

QUANTIFICATION OF SODIUM BENZOATE IN FOOD

LECTURER

DR. CHYE FOOK YEE

DEMONSTRATOR

MR TIN

NAME: SHATESH KUMAR A/L CHANDRAHASAN

MATRIX NOMBOR: BN09110073