Stability Profiling of Amodiaquine under stress Degradation Conditions

 

Neethu Mathew*, Jane T Jacob

Department of Pharmaceutical Chemistry, NGSM IPS, Derlakatte, Mangalore.

 

ABSTRACT:

Degradation Product is an impurity which results from a chemical change in the drug substance during storage and/or manufacture of the new drug product due to some environmental changes or by reaction with an excipient and/or the immediate container closure system. Impurity development may be either during formulation or upon aging of both API’S and formulated API’S in medicines. The objective of the current investigation was to study the stress degradation behaviour of Amodiaquine under different ICH recommended stress conditions by HPLC method. The anti-malarial drug Amodiaquine, when subjected to stress degradation conditions of hydrolysis, oxidation, photolysis and thermal decomposition prescribed by ICH guideline Q1A (R2), it is found that, the drug Amodiaquine (API) was stable. The method was established using column C18 Hypersyl BDS (250* 4.6) mm, 5µ, mobile phase as monobasic potassium phosphate _PH 2.5 with orthophosphoric acid and methanol. Method was isocratic and composition 22: 78 (A: B), at a flow rate of 1.2ml/ min. To facilitate the development of analytical methodology, in order to obtain a better understanding of active pharmaceutical ingredient (API) and drug product (DP) stability, and also to provide information about degradation pathways and degradation products, this process is very useful. These types of studies will provide information about the degradation products that could form during storage and transportation.

 

 

 

1. INTRODUCTION:

Amodiaquine is an antimalarial medication of dipeptidylpeptidase-4 (DPP-4) inhibitor clas. It is chemically described as 4-[(7-chloroquinolin-4-yl) amino]-2[(diethylamino) methyl] phenol^1,2. It is recommended to be given with artesunate to reduce the risk of resistance. Amodiaquine is an orally active 4-aminoquinoline derivative with antimalarial and anti-inflammatory properties. Similar in structure and activity to chloroquine, amodiaquine is effective against some chloroquine resistant strains particularly; it is active against the asexual erythrocytic forms of, Plasmodium falciparum, the most deadly malaria parasite. The use of drug is recommended only as treatment and not for prophylaxis against malaria; the reason is that, the drug has been linked to severe cases of acute hepatitis which can be fatal.

 

plasmodicidal mechanism of action of amodiaquine is not completely certain. It inhibits heme polymerase activity, similar to other quinolone derivatives; this causes accumulation of free heme, which is toxic to the anti-malarial parasite. Amodiaquine binds to the free heme, preventing the parasite from converting it to, a form which is less toxic. This drug-heme complex is toxic to the parasite and disrupts membrane function and preventing DNA and RNA production and subsequent protein synthesis. The drug is official in the Indian Pharmacopoeias and USP-NF^3,4. The literature survey did not report any stability profiling of Amodiaquine hydrochloride, though methods are available for the combined form of drug^5,7. Studies involving techniques in combination like HPLC^8,6,9. RP-HPLC, UPLC methods have been reported for the determination of Amodiaquine hydrochloride in plasma and urine of humans, rats and dogs^10,11. Thus, an attempt was made stability profiling of Amodiaquine.

 

IUPAC Name: 4-[(7-chloroquinolin-4-yl) amino]-2-[(diethyl amino) methyl] phenol.

Fig .1. Amodiaquine chemical structure

 

2. MATERIALS AND METHOD:

2.1. Reagents:

 Analytical grade chemicals and reagents were used for the current work. Sodium hydroxide, hydrochloric acid procured from Rankem chemicals. HPLC grade water procured from Milli Q water system. HPLC grade Acetonitrile, potassium Dihydrogen phosphate and ortho phosphoric were procured from Merck chemicals.

 

2.2. Apparatus and equipment:

A dry-bath (Thermolyne, IA, USA) was used for solid state thermal stress studies. Precision water bath equipped with MV controller (Julabo,Seelbach, Germany) was used for solution degradation studies. For checking and adjusting the pH of buffer solutions a pH/Ion analyzer Metrohm, M780 was used. Other equipments like, sonicator Elma Elmasonic p300, and precision analytical balance Mettler Toledo, XP205 also used.

 

HPLC:

The separation of degradation products was achieved using a Prominence series liquid chromatograph (LC) equipped with photodiode array detector (ShimadzuKyoto, Japan) and Pursuit column C18 Hypersyl BDS (250×4.6) mm,5μ, Varian, North America). Aligent 1260 model HPLC system and software used was open labs.

 

2.3 Preparation of reagents:

Preparation of buffer:

Monobasic potassium phosphate buffer was prepared by taking 6.8gms in 200ml water and pH was adjusted to 2.5 with ortho phosphoric acid.

 

Preparation of 1 N HCl:

In a 100ml volumetric flask, taken 8.2 ml of Conc.HCl and volume was made up with distilled water.

 

Prepartion of 1 N NaOH:

1 N NaOH was prepared by taking 4gms of NaOH in a 100ml volumetric flask and volume was made up to 100ml with distilled water.

 

 

2.4 Preliminary analytical chromatographic method:

Method was obtained from USP monograph for related compound analysis for the initial identification of degradant impurities which are formed due to forced degradation. Many trials using various mobile phase compositions were tried and best composition for identification of degradants is mentioned below.

 

Table 1: Chromatographic parameters

Mobile phase A	Monobasic potassium phosphate pH 2.5 with orthophosphoric acid
Mobile phase B	Methanol
Method composition	Isocratic (22:78)
Column	C18 Hypersyl BDS( 250× 4.6 mm x 10cm) ,5µ
Column temperature	30oC
Flow	1.2 ml/ min
ƛ max	224 nm
Sample preparation	0.15 mg/ml
Injection volume	10µl

 

2.5 Stress degradation:

To identify the unknown degradants present in Amodiaquine drug was subjected to different stress degradations such as acid , alkali, Peroxide , and thermal conditions as per ICH Q1A (R2) recommendations. The guideline as per ICH entitled stability testing of new drug substances and products requires that stress testing be carried out to elucidate the inherent stability of the active substance¹¹. The aim of this work was to perform the stress degradation studies on amodiaquine in different molarityof HCl and NaOH solutions, for acid and alkali degradation . Peroxide degradation was carried out by 30% H₂ O₂ . Thermal stress was carried out by keeping about 200 mg API in the oven for about 5-6 hours at 105 ˚C.

 

1. Acid degradation

2. Alkali degradation

3. Peroxide degradation

4. Thermal degradation

 

Procedure for Acid degradation:

15 mg amodiaquine was taken in 100ml volumetric flask to which added about 10ml of 1N HCl and kept it on a water bath at about 80^oC for about 1hr, diluted to volume with water and the solution was thoroughly mixed.

 

Procedure for Alkali degradation:

15 mg amodiaquine was taken in a 100ml volumetric flask to this added about 10ml of 1N NaoH, kept it on a water bath at about 80^oC for about 1hr, and diluted to volume with water and mixed well.

 

Procedure for peroxide Degradation:

15 mg of Amodiaquine was taken in a 100ml of volumetric flask to which about 5ml of 30% H₂ O₂ was added and kept it on a bench for 1 hr , and then diluted to volume with water and mixed well.

 

Procedure for Thermal Degradation:

Kept about 200mg of amodiaquine in an oven 105 ^oC for about 5-6 hrs. From this 15mg of amodiaquine dissolved in water and diluted to volume with water and mixed well.

 

Fig.2. Amodiaquine blank chromatogram

 

Fig. 3. Amodiaquine API Chromatogram

 

Fig.4. Amodiaquine acid degradation Chromatogram

 

Fig.5. Amodiaquine alkali degradation Chromatogram

 

Fig.6.Amodiaquine peroxide degradation Chromatogram

 

Fig.7. Amodiaquine thermal degradation Chromatogram

 

3. RESULTS AND DISCUSSION:

Stress degradation studies were carried out under the conditions of acid , alkali, Peroxide , and thermal as per, ICH Q1A (R2) recommendations . The ICH guideline entitled by stability testing of new drug substances and products requires, that stress testing be carried out to elucidate the inherent stability characteristics of the active substance.Under all the forced stress degradation conditions , the anti- malarial drug Amodiaquine was found to be stable.

 

Table 2: Amodiaquine (API ) Retention Time

Drug	Retention Time
Amodiaquine (API )	6.25 mins

 

Table 3: Results & Discussion

Degradation condition	Retention time
Acid degradation	6.207 mins
Alkali degradation	6.32 mins
Peroxide degradation	6.200 mins
Thermal degradation	6.213 mins

 

4. CONCLUSION:

In this study, Anti-malarial drug Amodiaquine was subjected to stress studies under various ICH recommended conditions. Amodiaquine remained unaffected under different stress degradation conditions such as Acidic, basic, peroxide, and thermal. This method proved the inherent stability of the API Amodiaquine under different stress condition to be good.

 

5. ACKNOWLEDGEMENT:

The authors acknowledge the support and necessary facilities provided by NGSM IPS, Mangalore, Nitte deemed to be university, Mangalore, Vidgas Labs, Hydrabad to carry out this work.

 

6. REFERENCES:

1.        Martindale, The Extra Pharmacopoeia, The complete Drug Reference, 35th edn, Pharmaceutical Press: London; 2007. -33.

2.        The Merck Index, 13th ed. Whitehouse Station NJ: Merck & Co. Inc., 2003.

3.        Indian Pharmacopoeia (2014). 7th ed.vol. 2, Ghaziabad: Indian Pharmacopoeia Commission.

4.        USP (32) NF (27). Vol.2, Asian edition 2008: 1406-1407.

5.        P. R. Rao et al. Analytical method development and validation of Artesunate and Amodiaquine hydrochloride in tablet dosage form by RP-HPLC. J. Res. Pharm. Biotechnol.2013, 1: 822-827.

6.        A. Lawal et al. Development and validation of UV Spectrophotometric and HPLC Methods for Quantitative Determination of Chloroquine and Amodiaquine in Pharmaceutical Formulations. I. J. ChemTech Res2012, 4: 669-676

7.        M. T. Ansari et al. Spectrophotometric Determination of Amodiaquine and Sulfadoxine in Pharmaceutical Preparations. Chem. Anal 2008, 53: 305-313.

8.        N. G. Rathod et al.Identification of impurities related to Amodiaquine Hydrochloride by using some analytical techniques. WJPR, 2014,3:1739-1758.

9.        M. U. Phadke et al. Development and Validation of Stability indicating UPLC Method for Determination of Related Impurities in Artesunate and Amodiaquine Fixed Dose Tablets. Der Pharmacia Lettrre, 2010, 2: 333-341.

10.      S. Gandhi et al .A simple and sensitive RP-HPLC method for simultaneous estimation of Artesunate and Amodiaquine in combined tablet dosage form. J. Chem. Pharm. Res. 2010, 2: 429-434.

11.      ICH guidelines, Q1A (R2): Stability Testing of New Drug Substances and Products (revision2), International Conference on Harmonization.

 

 

 

 

 

Accepted on 17.05.2019           © RJPT All right reserved

Research J. Pharm. and Tech 2019; 12(8):3807-3810.