Spectrophotometric determination of Chlorpromazine Hydrochloride by oxidative coupling reaction with P- Bromo aniline using Ammonium Ceric Sulphate Dihydrate

A new sensitive spectrophotometric method for the determination of chlorpromazine hydrochloride (CPZH) was developed. The method is based on an oxidative coupling reaction with p-bromoaniline using ammonium ceric sulphate dihydrate as oxidizing agent to produce a violet color, soluble in water, stable product and absorbs at 553 nm. Beer's law was in the linear range 2.5 30 μg/ml of CPZH, the molar absorptivity, Sandellʼs sensitivity index and detection limit were 6.5731 × 10 3 liter. mol -1 .cm -1 , 0.0541 μg.cm -2 and 0.1361 μg/ml respectively. The RSD value was 1.20 1.36 % depending on the concentration. This method was applied successfully to the determination of chlorpromazine hydrochloride in two pharmaceutical formulations (tablets and injection).

The chemical structure is:

M. wt = 355.3g/mol
Chlorpromazine hydrochloride (CPZH), white or almost white, crystalline powder, very soluble in water, freely soluble in ethanol. It decomposes on exposure to air and light [1].
Chlorpromazine is a psychiatric medication that belongs to the class of drugs called phenothiazine antipsychotics, it works by helping to restore the balance of certain natural substances in the brain [2]. It is used to treat various problems such as severe depression or behavioural disturbances and it is used to treat schizophrenia [3] and it also used to treat nausea, vomiting, severe pain, relieve prolonged hiccups, relieve restlessness/anxiety before surgery, and help treat tetanus [4]. side effects of chlorpromazine, HCl is low heat and drowsiness and nasal congestion and difficulty in urination and other side effects of low blood pressure and accelerated heart rate (in the case of intramuscular injection) [5].

Aim of research
In this research a rapid, sensitive and simple spectrophotometric method for determining of chlorpromazine hydrochloride (CPZH) in pure form as well as in pharmaceutical formulations based on the oxidative coupling using p-bromoaniline in presence of ammonium ceric sulphate dihydrate.

Experimental Apparatus
The apparatuses used shown in the Table (1).

Reagents and chemicals used
All chemicals and analytical reagents used in this research are high purity and shown in the

1-Standard CPZH solution,250 μg/ml
This solution is prepared by dissolving 0.025 g of CPZH in amount of distilled water and the volume is diluted to 100 ml with distilled water in a volumetric flask. This solution is kept in a brown bottle, where it is stable for one week, at least.

2-P-bromoaniline reagent solution (1 ×10 -2 M)
This solution is prepared by dissolving 0.1720m g of p-bromoaniline in 5 ml of ethanol and the volume is completed to 100 ml in a volumetric flask with distilled water. A 0.1000 g of each foreign compounds is dissolved in distilled water then the volume is completed to 100 ml in a volumetric flask with distilled water.

7-Solution of CPZH tablets formulation (250 µg/ml (
Largactil Chlorpromazine -HCl (100mg), Oubari Pharma -Aleppo-Syrie, every tablet contains 100 mg of CPZH and the solution is prepared as follows: Ten tablets are weighed (4.1649g) and powdered well, then a weight of 0.1041 g of this powder is dissolved in an amount of distilled water, and then the solution filtered by paper filtration (604, RUNFILTER, Q240 mm), the volume is completed with distilled water in a volumetric flask of 100 ml , which gives the concentration of 250 µg/ml.

8-Solution of CPZH injection formulation (250µg/ml)
Largactil Chlorpromazine -HCl (25mg/5ml), Oubari Pharma -Aleppo -Syrie, each injection contain of 25 mg of chlorpromazine and this solution is prepared by transferred of 50 ml of this formulation to volumetric flask and the volume was completed to 100 ml with distilled water to obtain a solution with a concentration of 250 µg/ml.

Optimization of the experimental conditions
To establish the optimum conditions, the effect of various variables on the intensity of the absorption was studied by adding ..0 ml of standard CPZH solution (250µg/ml),1.0 ml of pbromoaniline and 0.5ml of Ce +4 and measuring the absorption at 553 nm versus the blank reagent.

Effect of acid
The effect of acid was studied by adding 0.5-3.5 ml of strong acids and weak acids (0.1 M). the results showed the H 2 SO 4 solution gives a higher absorption for colored product at a wavelength of 553 nm compared with other acids used , so this acid was chosen in subsequent experiments. the results are shown in Table (3).   The results shown in the Table (4) indicate that the volume of 1.0 ml of (Ce +4 ) (1×10 -2 M) is the optimum amount because of highest absorbance, so it was used in subsequent experiments.

Effect of the amount of coupling reagent
The effect of the amount of coupling reagent was studied by adding different volumes  Table (5).   Table (6) shows that 15 min is sufficient for the oxidation to be completed , so it is adopted in the subsequent experiments.

Effect of temperature
The effect of temperature(5-60°C) on the absorption of the formed colored product were studied by using 2.0 ml of CPZH solution (250µg/ml) and 1.0 ml of (Ce +4 ) ( 1 × 10 -2 M), then

Effect of the solvents
The effect of the solvents on the formed colored product was studied, the dilution was carried out by different organic solvents instead of water. The results shown in Table (9) indicate that the water is a good medium for reaction and gives good absorption value at the wavelength of 553nm and due to its availability, it was used as the best solvent in the subsequent experiments.

Final absorption spectrum
The spectrum of the formed colored product by coupling of CPZH with p-bromoaniline (1×10 -2 M) in the presence of Ce +4 (1×10 -2 M) in acidic medium (pH 1.8) and temperature 25°C against its corresponding reagent blank show a maximum absorption at 553 nm in contrast to the blank reagent of zero absorbance at λ max . The spectra are shown on Fig. (1).

Accuracy and precision
Accuracy and precision were studied by measuring absorption (n=5) at 553 nm for two different concentrations of the drug within the limits of Beer's law, the average recovery (99.75 %) and the relative standard deviation ( 1.39%) indicate that the method is of high accuracy and precision . The results are shown in Table (10).  1, March 2016 , p.p(1-19) ISSN 1992 -0849 Web Site: www.kujss.com Email: kirkukjoursci@yahoo.com, kirkukjoursci@gmail.com

Detection limit
Detection limit was calculated by measuring the absorption for the lower concentration 2.5 µg/ml at optimal conditions (ten times) at 553 nm. The results are shown in Table (11).   The volumes were completed to the mark with distilled water and the absorbance was measured at 553 nm against the blank reagent. Molar ratio was found to be 1:1. The results are shown in Fig.(4) which is in agreement with the Job's method results.

Effect of interferences
In order to test the efficiency and selectivity of the proposed method, the effect of some foreign substances (lactose, fructose , maltose, starch and glucose) that usually present in dosage forms are studied by taking volumetric flasks ( 25 ml) containing 2.0 ml of CPZH (250 µg /ml), then different volumes ( 2.5, 5.0 ,7.5 ml) of foreign substances (1000 µg/ml) have been added resulting in a final concentration of (100, 200 ,300 µg/ml ). The optimum conditions have been applied and the volumes have been completed to the mark with distilled water.The absorbance was measured at 553 nm versus blank reagent and recovery is calculated. The results showed that there is no interferences

Applications
This method was applied for the determination of CPZH in its pharmaceutical formulations (tablets and injection).

Direct method
In this method, different volumes (0.25, 0.5ml) of a pharmaceutical formulation solutions (250 µg/ml) were transferred to 25 ml volumetric flasks and the resulting concentrations (2.5, 5 µg/ml) and were treated as in construction of calibration curve. The absorbance was measured at 553 nm for five times. Recovery and RSD were calculated and the results are shown in Table (13).

standard additions method
To prove that the developed method is free from interferences , method of standard additions is applied for determining of CPZH in its pharmaceuticals. Different volumes  The results shown in table 12 indicate that method of standard additions is in agreement with the direct method within the acceptable range of error, indicating that the method is satisfactory and free from interferences.

4.Conclusions
The results obtained confirm that the proposed method is simple, rapid and of good sensitivity for the determination of chlorpromazine hydrochloride CPZH. The method is based on oxidative coupling between CPZH and p-bromoaniline reagent in presence of ammonium ceric sulphate dehydrate (Ce +4 ) in acidic medium to form a violet colored dye which is water soluble, stable and shows a maximum absorption at 553 nm. This method does not require temperature control, nor use of organic solvents, or solvent extraction and it can be applied successfully for determination of CPZH in pharmaceuticals formulation.