Fifth stage second lab Pharmaceutical analysis

SPECTROPHOTOMETRIC ANALYSIS OF TETRACYCLINE

Introduction:
Tetracycline is a broad-spectrum polyketide antibiotic produced by the
Streptomycesgenus of Actinobacteria, indicated for use against many bacterial infections. It is
a protein synthesis inhibitor. It is commonly used to treat acne today, and, more recently,
rosacea, and is historically important in reducing the number of deaths from cholera.
Tetracycline is marketed under the brand names Sumycin, Tetracyn, and Panmycin, among
others. Actisite is a thread-like fiber formulation used in dental applications. It is also used to
produce several semisynthetic derivatives, which together are known as the tetracycline
antibiotics. The term "tetracycline" is also used to denote the four-ring system of this
compound; "tetracyclines" are related substances that contain the same four-ring system.

Systematic (IUPAC) name:- (4S,6S,12aS)-4-(dimethylamino)-

1,4,4a,5,5a,6,11,12a-octahydro-3,6,10,12,12a-pentahydroxy-6-methyl-1,11-
dioxonaphthacene-2-carboxamide.

Molecular formula : - C22H24N2O8

The tetracycline content of sample can be determined by one of three colorimetric

methods:

1-Ferric chloride method.

2-Acid method .

3-Base method.
 Fifth stage second lab Pharmaceutical analysis

Ferric chloride method

It may be used to determine any of these three antibiotics chlortetracycline,

oxytetracycline and tetracycline. all contain the same phenolic hydroxyl groups and they all
read with ferric chloride in acidic solution to give an orange – brown color.

A colored complex is formed between tetracycline and the iron (III) ion. The
intensity of the color is directly related to the concentration of tetracycline present; therefore,
spectrophotometric analysis can be used.

A series of solutions with different tetracycline concentrations will be prepared and

complexed. The absorbance of each solution will be measured and a calibration curve will be
constructed. Using the standard curve, the amount of tetracycline in a commercial
tetracycline product can be determined.

The general method applicable to all three tetracyclines. This method may be used to
determine the tetracyclines in such products as:

1-capsules.

2-ointments.

3-tablets.

And troches after suitable filtration or extraction to eliminate any insoluble or immiscible
material ( complex )
 Fifth stage second lab Pharmaceutical analysis

Purpose

The purpose of this lab is to determine the amount of tetracycline in a commercial tetracycline product.

Procedure:

Part I : Reagents:

Dilute 1ml of 10% Ferric chloride solution (Fecl3) to 200 ml with 0.01 N HCl.

Part II: Making Standards.

1 .Prepare 250 ppm by dissolving 250 mg of Tetracycline in a small amount of distilled
water.

2. Quantitatively transfer the solution to a 1000 mL volumetric flask, and complete the
volume with distilled water to the mark.

3. Transfer (2,4, and 6) ml of 250 ppm of tetracycline and add ( 8,6,4 ) ml of 0.01 N(HCl )
to a 25mL volumetric flask , respectively to give each total volume of ( 10 ml).

4. To each of these flasks add (10 ml) of FeCl3 (0.05 %). then, allow for each sample to stand
at room temperature for ( 10 ) min then complete to 25 ml by DW.;.

Part III Making Blank :

Transfer ( 10 ml ) of the ( 0.01 N HCl ) to flask and add ( 10 ml ) of ( FeCl 3 ) (0.05% ) in

volumetric ( 25 ml ).

Part IV: Making an unknown from a capsule:

1. Give amount of oxytetracycline to prepare solution of unknown concentration.

2. Pipet a 1mL sample of tetracycline solution to a 25 mL volumetric flask. add ( 9 ml ) (

0.01 N HCl ),then put 10 ml of ( FeCl3 ) (0.05% ) in volumetric ( 25 ml ).

3. Label this solution "unknown," and stand it at room temperature for 10 min.

Part VI: Testing the Solutions.

1. Turn on the spectrophotometer.
2. Adjust the wavelength to max= 490 nm and record the absorbance ( A ).
 Fifth stage second lab Pharmaceutical analysis

Calculation :

Calculate the tetracycline content of the sample under test from the standard curve .

C ( PPm )

Questions

Can be used basic medium in determination of tetracycline and why?

What is the linearity of the method?

Is ferric chloride oxidizing agent or reducing agent?

What is the effect of chelation of ions with tetracyclines?