Degradation study of Doxycycline in bulk and formulation by UV-Visible spectrophotometry

Introduction: Forced degradation is a process that involves degradation of drug products and drug substances at condition more severe than accelerated conditions and thus generates degradation products that can be studied to determine the stability of the molecule. Aim and objective: To investigate the forced degradation study for the determination of degradation of Doxycycline by UV-Visible spectrophotometric method. Methods: Doxycycline sample and standard were exposed to different stress conditions (hydrolytic and oxidative degradation). Both standard drug and marketed formulations were used for the degradation study. The amount of percentage degradation of each standard and samples were calculated by taking absorbances at their λ max with the help of UV-Visible spectrophotometer. Forced degradation of drug substance was done by exposing to acidic, basic and to medium of hydrogen peroxide. The degradation results of each condition were compared with that of standard. This method can be used successfully for studying the stress degradation factors. Because this method is less time consuming and simple and cost effective also. Results: Forced degradation of selected drugs performed using HCl, NaOH and H 2 O 2 . Degraded sample were quantified by UV visible spectroscopy. In all the methods used in degradation study, sample undergoes greater degradation compared with that of standard. Among the degradation conditions used in the study it is found that NaOH produce more degradation


Introduction
Analytical chemistry is the study of separation, quantification and identification of chemical components of natural and artificial materials constituted with one or more compounds or elements. Analytical chemistry is separated into two main categories; qualitative analysis that is to say the identification with regard to the chemical components exists in the sample, whereas quantitative analysis estimates the amount of elements or compounds in the substance i.e., sample.
UV-visible spectrophotometry is one of the most frequently employed techniques in pharmaceutical analysis. It involves measuring the amount of ultraviolet or visible radiation absorbed by a substance in the solution [1][2][3][4][5].
Antibiotic, chemical substance produced by a living organism, generally a microorganism, that is detrimental to other microorganisms. Antibiotics commonly are produced by soil microorganisms and probably represent a means by which organisms in a complex environment, such as soil, control the growth of competing microorganisms.
Doxycycline is a broad-spectrum antibiotic of the tetracycline class used in the treatment of infections caused by bacteria and certain parasites. It is used to treat bacterial pneumonia, acne, chlamydia infections, lyme disease, cholera, typhus, and syphilis. It is also used to prevent malaria in combination with quinine. Doxycycline may be taken by mouth or by injection into a vein.
Common side effects include diarrhoea, nausea, vomiting, abdominal pain, and an increased risk of sunburn. Use during pregnancy is not recommended. Like other agents of the tetracycline class, it either slows or kills bacteria by inhibiting protein production. It kills malaria by targeting a plastid organelle, the apicoplast [6][7][8].

Standard drugs
Doxycycline RS obtained from Cipla Ltd.

Test product
Doxycycline capsule (Doxicip 100 mg) was purchased from local medical shop.

Preparation of reagents
Preparation of 0.1M sodium hydroxide.
4mg of sodium hydroxide pellets were weighed and dissolved in small amount of distilled water then made up the volume to 1000 ml.
Preparation of 0.1M Hydrochloric acid 8.33ml of concentrated hydrochloric acid was measured and diluted with distilled water to 1000ml.

Degradation study of Doxycycline
Standard prepration 100 mg of Doxycycline was transferred to volumetric flask and dissolved in 100 ml of distilled water to achieve a concentration of 1 mg/ml. The solution was kept at room temperature. An aliquot solution was then diluted with distilled water to get final concentration of 20 µg/ml, 40 µg/ml, 60 µg/ml, 80 µg/ml, 100 µg/ml. The solution was scanned in the UV region and the maximum absorbance was recorded at 274 nm.

Sample preparation
Weighed and transferred 0.518 g of doxycycline capsule which contains 100 mg of Doxycycline to a 100 ml standard flask and made up the volume with distilled water. Solution was filtered and prepared five concentrations such as 20 µg/ml, 40 µg/ml, 60 µg/ml, 80 µg/ml & 100 µg/ml. Absorbance was measured at 274 nm.

Blank preparation
Blank solution was prepared by pipetting 6 ml NaOH to volumetric flask & made up to 100 ml with distilled water.

Intraday oxidative degradation study using 3% H2O2
Standard preparation (stress) 100 mg of Doxycycline standard was weighed and transferred to volumetric flask, containing 3% H2O2 & dissolved then make up the solution to 100 ml. 6 ml of the resulting solution was diluted to 100 ml to produce 60 µg/ml. The absorbance of the solution was taken at 274 nm in zero minutes. The same procedure was repeated for 30 min, 60 min and 90 min time interval Sample preparation (stress) 100 mg equivalent of Doxycycline capsule (0.518 g) were crushed weighed and transferred to volumetric flask, dissolved in 3% H2O2 & made up the solution to100 ml. 6 ml of the resulting solution was diluted to 100 ml to achieve a concentration of 60 µg/ml. The absorbance of the solution was taken at 274 nm in zero minutes. The same procedure was repeated for 30 min, 60 min and 90 min time interval

Conclusion
The present study involves the forced degradation studies such as acid and alkali hydrolytic degradation and oxidative degradation of Doxycycline bulk and formulation. Doxycycline is soluble in water, so λmax of drug was identified by measuring UV visible spectrum in the range of 800-200 nm in distilled water. Calibration curve of drug plotted in the obtained λmax (274 nm) and the calibration curve found to be linear in the selected concentration range (20-100 µg/mL). Forced degradation of drug was performed by using HCl (0.1 M), NaOH (0.1 M) and 3% H2O2. Degraded sample were quantified by UV visible spectroscopic method and the percentage degradation of each drug was re calculated. In all the methods used in degradation study, sample undergoes greater degradation when compared with that of standard. Among the methods selected for the study it was found that hydrolytic degradation using NaOH cause greater degradation.