Environmental Friendly Method for Novel Estimation of Chlordiazepoxide in Pharmaceutical Preparations and Environmental Wastewater Samples: Application to Content Uniformity Testing

Ahmed NR and Al-Etewi MJE

Published on: 2023-10-18


A simple, accurate, precise, rapid, economical and sensitive ultra violet method has been developed for the estimation of chlordiazepoxide in pharmaceutical preparations and environmental wastewater samples, which shows maximum absorbance at 245 nm in 0.1M hydrochloric acid. Beer's law was obeyed in the range of 0.5-10mg/ ml, with molar absorptivity of 3.282X104 L.mol-1.cm-1, relative standard deviation of the method was less than 1. 5%, and accuracy (average recovery %) was 100 ± 1.0  .The method was successfully applied to the estimation of chlordiazepoxide in pharmaceutical formulations ,content uniformity testing and  industrial wastewater samples The proposed method was  validated by sensitivity and precision which proves suitability for the routine analysis of chlordiazepoxide  in true pharmaceutical formulations samples and environmental wastewater samples : Application to content uniformity testing.


Ultra violet; Chlordiazepoxide; Pharmaceutical formulations


Chlordiazepoxide (Figure 1) ia 7-Chloro-N-methyl-5-phenyl-3H-1,4-benzodiazepin-2-amine 4-oxide [1].

Figure-1: Chemical Structure of Chlordiazepoxide.

7-Chloro-2-methylamino-5-phenyl-3H-1,4-benzodiazepin- 4-oxide C16H14ClN3O   ?299.75

It used as an anxiolytic, sedative-hypnotic, tranquilizer, and anti-depressant. It shares the actions of other benzodiazepines and is used for the management of anxiety disorders or for short-term relief of symptoms of anxiety and for the management of agitation associated with acute alcohol withdrawal [2-3].Several methods have been reported for the quantitative determination of chlordiazepoxide including titrimetric [1]. HPLC, gas chromatography (GC), spectrophotometry, ion-selective electrodes and voltammetry [4-18]. This paper reports a simple, sensitive and accurate new ultra violet method for determination of chlordiazepoxide in pure form pharmaceutical preparations and environmental wastewater samples: Application to content uniformity testing.



Shimadzu UV- 1700 pharmaspec (double beam) spectrophotometer with 1.0 cm quartz cells was used for absorption measurement.


All chemical used were of analytical or pharmaceutical grade and chlordiazepoxide standard material was provided from AL-hokamaa company for pharmaceutical industries (HPI) Mosul-Iraq.

Chlordiazepoxide stock solution (1000 ppm) was prepared by dissolving 0.1g of chlordiazepoxide in 100 ml of (0.1M) hydrochloric acid in a volumetric flask.                                                                                                 

Chlordiazepoxide standard solution (100 ppm) was prepared by diluting 10 ml of stock solution to 100 ml by (0.1M) hydrochloric acid in a volumetric flask.      

Determination of Absorption Maxima

The standard solution of chlordiazepoxide (200µg/ml) was scanned in the range of 220-350 nm which shows maxima located at 245 nm Figure 2. Therefore, 245 nm wavelength was selected for the construction of calibration curve.

Figure 2: Absorption spectra of chlordiazepoxide 200 (µg/ml) against blank.

Recommended Procedure

From the absorption maxima, calibration curve was prepared in the concentration range of 0.5-10.0 mg/ml. The absorbance was measured at 245nm against (0.1M) hydrochloric acid as a blank. The concentration of the sample solution can be determined by using the calibration curve.

Procedure for Environmental Wastewater Samples

To demonstrate the practical applicability of the proposed method. Industrial waste water from AL- hokamaa company for pharmaceutical industries (HPI) Mosul-Iraq, were fortified with the concentrations in the range of 2,5,6 mg\ml of chlordiazepioxide. The fortified water samples were analyzed as described above for recommended procedure and the concentration was calculated by using the calibration curve of this method.

Procedure for Pharmaceutical Preparations (Tablets)

Weight and powder 10 tablets. Dissolve a quantity of the powdered tablets containing 0.01 gm. of chlordiazepoxide in about 100 ml of (0.1M) hydrochloric acid and mixed for 20 mint and then filtered. The filtrate was mad up to 100 ml with (0.1M) hydrochloric acid and aliquot of this solution was treated as described above for recommended procedure and the concentration was calculated by using the calibration curve of this method.

Result And Discussion

UV Visible spectrophotometry is still considered to be a convenient and low cost method for the determination of pharmaceuticals. The method used for the determination of chlordiazepoxide in pharmaceutical preparations and environmental wastewater samples was found to be sensitive, simple, accurate, and reproducible. Beer s law was obeyed in the concentration range of 0.5-10.0 mg/ml Figure 3 with correlation coefficient of 0.9999, intercept of - 0.0006 and slope of 0.1095 .The conditional molar absorptive was found to be 3.282X104  l/mol.cm.

Figure 3: Calibration curve for chlordiazepoxide.

The accuracy and precision of the method, a pure drug solution was analyzed at three different concentrations, each determination being repeated six times. The relative error (%) and relative standard deviation values are summarized in table 1.From Table 1 the values of standard deviation were satisfactory and the recovery studies were close to 100%. The RSD% value is less than 1.5 indicative of accuracy of the method.

Table 1: Accuracy and precision of the proposed method.

Chlordiazepoxide  taken( µg/ml)

Er (%)a











 *Mean of six determinations

The proposed method was satisfactorily applied to the determination of chlordiazepoxide in its pharmaceutical preparations tablets and wastewater samples, the results of the assay of the pharmaceutical preparations revels that there is close agreement between the results obtained by the proposed method and the label claim Table 2 and the results of water samples Table 3 show that recovery values obtained were closed to 100%.

Table 2: Assay of chlordiazepoxide pharmaceutical formulations.

Pharmaceutical formulation supplied by HPI

Amount of chlordiazepoxide *

Label claim


Tablet 1mg


1 mg


*Mean of ten determinations.

Table 3: Determination of chlordiazepoxide in spiked industrial wastewaster sample.

Water sample

Trifluogerazine (µg/ml) *

% Recovery

Industrial wastewater













Table 4: Content uniformity testing of chlodiazepoxide tablets using the propose method.


% of the label claim

Tablet NO. 1 Tablet NO. 2 Tablet NO. 3 Tablet NO. 4 Tablet NO. 5
Tablet NO. 6
Tablet NO. 7

Tablet NO. 8
Tablet NO. 9
Tablet NO. 10 Mean (x)
Max. allowed unit 1201

100. 2
100. 1

99. 65
100. 15
100. 16 99.76 99.9 0.518



In this work, a simple, rapid, precise and accurate UV-Spectrophotometric method was developed and validated for the determination of chlordiazepoxide in pharmaceutical preparations and industrial waste water samples. Themethod free from such experimental variables as heating or solvent extraction steps. The method rely on the use of simple and cheap chemicals and techniques and can be used for rapid routine determination and quality control of chlordiazepoxide in pure form, pharmaceutical preparations and real industrial waste water sample.                                             


The first author[Nief Rahman Ahmad] wishes to express gratitude to the state company of  drug industries and medical appliance (HPI)  Nineveh – Iraq  for providing gift samples of chlordiazepoxide standard  materials and pharmaceutical preparations (tablets) and for permission and facilities to carry out the research work .


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