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CHEMICAL PROBLEMS 2025 no. 2 (23) ISSN 2221-8688
OXIDATIVE COUPLING REACTION FOR THE SPECTROPHOTOMETRY DETERMINATION OF FUROSEMIDE USING CHLORPROMAZINE HYDROCHLORIDE AS A REAGENT
Mohammed S. Al-Enizzia, Omar A. Sheej Ahmadb*, Mohamed Y. Dhamrab
aDepartment of Chemistry, College of Education for Girls, University of Mosul, Iraq bDepartment of Chemistry, College of Education for Pure Science, University of Mosul, Iraq
*e-mail: [email protected]
Received 23.06.2024 Accepted 09.08.2024
Abstract: A simple, economical, and rapid method has been developed for the determination of the pharmaceutical compound (furosemide) in its pure forms and several pharmaceutical formulations using oxidative coupling reactions. This method focuses on the reaction of the drug compounds with chlorpromazine hydrochloride as a reagent in an acidic medium in the presence of the appropriate oxidizing compound. The absorbance of the colored product was measured at 525 nm and molar absorptivity was 138891 L/mol.cm, compatible with Beer law in concentrations (2-32) ¡¡g/ml, the detection, and the quantification limits were (0.471, 1.571) fg/ml respectively, with a recovery rate of 101.15% and the relative standard deviation was less than 0.5%. The great correlation coefficient value (R2 = 0.9953) and insignificant values of intercept (0.067) validated the good linearity of the calibration curve and agreement with Beer's law. The created method was applied successfully to determine furosemide in pharmaceutical formulation, and the method was in agreement with the standard addition method. Keywords: Spectroscopic Determination, Furosemide, Chlorpromazine Hydrochloride. DOI: 10.32737/2221-8688-2025-2-256-265
1. Introduction
Furosemide (FA), scientifically named "5-(aminosulfonyl)-4-chloro-2-[(2 furanylmethyl) amino]benzoic acid". The FA has a chemical formula as C12H11QN2O5S, with a molecular weight of 330.7 g/mol. It is a white, slightly yellowish crystalline powder [1, 2]. It is considered a diuretic and used to treat liver,
high blood pressure, and kidney diseases and is taken orally, either alone or with other blood pressure medications [3-5]. Furosemide was estimated by several methods, including spectrophotometry [6-10], electrical [11, 12] and chromatographic [13, 14]. Figure 1 shows the chemical structure of furosemide.
H,N
Fig. 1. Furosemide Chemical Structure
In this research, furosemide was estimated spectrophotometrically by oxidative coupling interaction with chlorpromazine hydrochloride as a reagent and in the existence
of an appropriate oxidizing agent. The method was successfully applied to pharmaceutical preparations.
CHEMICAL PROBLEMS 2025 no. 2 (23)
www .chemprob.org
The current study aims to create a modest technique that can be conducted for the quality
and reproducible approach for the selective control process (CIP) of the drug or determine
spectrophotometry determination of FA in the pure FA in different drug formulations. tablet and injection using a simple UV-VIS
2. Experimental part
Instruments. The spectral measurements were carried out using Shimadzu UV-1800 double-beam spectrophotometry using glass cells with a width of 1 cm.
Chemicals and Reagents. Reagents solution: All chemicals used were of analytical purity grade. Detail information are given in Tables 1 and 2.
Table 1. Preparing the chemical compounds involved in the experiments
Chemicals Cons. Preparation Final dilution with water
Furosemide (100 ^g/ml) prepared freshly by dissolving 10 mg of the pure substance in 5 ml of Abs. ethanol 100 mL
Chlorpromazine hydrochloride (0.1%) prepared by dissolving 10 mg of the pure substance in pure water 100 mL
Potassium dichromate (5x 105 M) prepared by dissolving 1.4709 mg of the pure substance in pure water 100 mL
Copper sulfate pentahydrate (1x 102 M) prepared by dissolving 249.685 mg of the pure substance in pure water 100 mL
Hydrochloric acid (1x 101 M) prepared by diluting the concentrated acid with pure water 100 mL
Table 2. Preparation of drug solution (Furosemide) from the pharmaceutical preparation
Injection A 2ml vial containing 20mg of FA was added into a precise volume and diluted up to the mark (1 litter volumetric flask) with distilled water.
Tablets Ten tablets were weighed, and after grinding and mixing them well, the equivalent of the weight of one tablet was taken and dissolved well in an appropriate volume of ethanol, filtered then supplemented the volume to the mark (100 mL) with distilled water
3. Result and discussion
The absorption spectrum of the reagent absorbance of the colored product was 525 nm and furosemide complex. The maximum as shown in Fig. 2.
3*0 DO «90 00 MO 00 "M 00 «00 00
Wavelenghth(nm)
Fig. 2. Absorbance spectrum of 4 |ig/ml of FA against blank (A); 4 |ig/ml of FA against water
(B); Blank against water (C)
Method Validation. Under the optimized correlation coefficient was 0.9953 the slope of
conditions, Fig. 3 shows a linear correlation the curve was 0.042. After that, a negative
between absorbance and furosemide deviation from Beer's law occurred. concentration in the range of (2-32) pg/ml. The
Fig. 3. Standard calibration curve for furosemide
Reagent Volume. Chlorpromazine 2.5 mL) was examined. Fig. 4 shows the reagent was prepared in a concentration of optimal amount of the reagent on the reaction 0.1%. The effect of the reagent volume (0.25- product which gives the highest color intensity.
S
1 (A
S
0.10
0.05
O.OO
Reagent Volume
Lrilll
0.25 0.5 1 1.5 2 2.S mL of Rcagont (O.I %)
Fig. 4. The effect of Chlorpromazine volume (0.1%)
Typa of oxidizing agont
Fig. 5. Volume of oxidizing agent (A); Type of oxidizing agent (B)
Study the type & volume of oxidizing agents on the adsorption of the resulting agent. The effect of different types of oxidizing complex was studied by using potassium iodate
(KIO3), aqueous copper sulfate (CuSO4.5H2O), potassium dichromate (K2&2O7), potassium periodate (KIO4) and Potassium permanganate (KMnO4). The study showed that the best oxidizing agent is potassium dichromate (1.0 ml), as shown in Fig. 5. Potassium dichromate was chosen to be the powerful oxidizing agent due to its ability to readily donate oxygen atoms.
Studying the type & volume of acid. To
select the best acid that can give the highest absorbance, different types of acids starting from HCl, H2SO4, and H3PO4 with a
concentration (0.1 M) were tried and added to the volumetric flask. It was indicated that the use of HCl (2.5 ml) gives higher absorption as shown in Fig. 6. The effect of the base NaOH was also studied at a concentration of (0.1 M) and a decrease was observed in the absorption value, so the addition of the base was excluded. The choice of acid can indeed influence the intensity of the color produced. So, hydrochloric acid, being a strong acid, can effectively protonate leading to vivid color changes or intense hues. It indeed offers advantages in color intensity.
Fig. 6. Type of acid (A); Volume of acid (B)
Addition sequence effect. The effect of separately studied with variable order additions. the addition sequence on the product intensity of The result in Fig. 7 confirmed the best and most absorption of the two colored solutions was reliable sequence in subsequent measurements.
'ilill
I
II
III
IV
Order No.
Fig. 7. Order of Addition
I Drug+ Reagent+ Oxidizing agent +Acid
II Drug + Oxidizing agent +Acid+ Reagent
III Drug + Acid+ Reagent+ Oxidizing agent
IV Reagent + Oxidizing agent + Acid + Drug
V Reagent + Acid + Oxidizing agent + Drug
Effect of temperature and product stability. The effect of different temperatures ranging between 40 and 60 °C on the stability and absorption intensity of the resulting complex was studied using the optimal conditions obtained from previous experiments.
Under optimum reaction conditions, the results shown in Fig. 8 indicated that the laboratory temperature (40°) was the best and it gives the highest absorption value. In addition, the complex remains stable for 90 minutes (Table 3).
Fig. 8. Effect temperature and time stability
Table 3. Optimal conditions and regression parameters of the created method for furosemide
determination
X max (nm) 525
Reagent 0.1 % (ml) 1.5
Oxidizing agent (K2&2O7) (5x10"3 M) 1.0 mL
HCl 0.1 M (ml) 2.5
Temperature (°C) 40
Development Time (min.) 5
Product Stability Time (min.) 90
Method Linearity range (^g /mL) 2 - 32
Detection Limit Value (LOD) (^g /mL) 0.471
Quantitation Limit Value (LOQ) (^g /mL) 1.571
Molar absorptivity (l.mol-1.cm-1) 13889
The accuracy and precision test for the suggested method. The accuracy and precision of the suggested approach were evaluated by calculating the recovery rate and relative
standard deviation (R%, RSD) using 5 replicates for 3 different concentrations of each of the medicinal compounds, as shown in Table 4.
Table 4. Method accuracy and precision
Added quantity Recovery* Average recovery RSD*
(^g.ml-1) (%) (%) (%)
8 96.29 101.15 0.20
16 104.00 0.10
24 103.17 0.45
*Average of Five Determination
Study the nature of the resulting between furosemide and chlorpromazine in the complex. The continuous changes method was presence of the oxidizing agent in the acidic applied to find out the stoichiometry ratio medium [15]. The results in Fig. 9a indicate that
the ratio was 1:1 for the compound with the react. To confirm the composition ratio of the product, the Job method [16] has been applied as well. The results were compatible with the previous result which was calculated by the
continuous changes method. Based on the results, a reaction mechanism was suggested and the formation of the color product probably occurs as follows [17].
[Furosemidef]/lFurosemi<te}*(Chloropfomazirte HCl
Fig. 9a. Mole and Job Methods
o °
H-CI
Furosemide
l^J
CI
3-(2-chloro-1 OH-phenothiazin-10-yl)-N,N-dimethylpropan-1 -amine hydrochloride
H
H2N ^
° CI
Fig. 9b. Suggested reaction
Complex stability constant. Based on the was calculated at a ratio of 1:1 by applying the Job and continuous changes methods, the following equation: stability constant (Kst) of the formed product
Kst =
1 — a
a2C
Am — As
a =
Am
Where Kst is the stability constant, L/mol, a= the dissociation degree;
C is the concentration of the formed product which is the same concentration of Furosemide; Am is the absorbance of the complexes at the optimum amount of reagent (chlorpromazinej; As is the absorbance of the complexes at
stoichiometric amounts of chlorpromazine reagent according to the 1:1 ratio under the optimum conditions reaction.
The results illustrated in Table 5 indicate that the complex (Drug:Reagent) has good stability of the colored product.
Table 5. The stability constant of the resulting complex
Conc.(moM-1) Absorbance a Average Kst (l. mol-1)
As Am
2x10-6 0.017 0.025 0.320 1.525 x 106
4X10-6 0.019 0.034 0.441
6x10-6 0.030 0.052 0.423
Applications of Method. The proposed approach was conducted to estimate the FA in commercial dosage forms. The results in Table 6 show that the developed method has a very
good accuracy value and is in good agreement with the original content in pharmaceutical dosage forms.
Table 6. Determination of the drug compound in pharmaceutical preparations by the proposed
method
Pharmaceutical dosage Certified weight Amount present (^g. ml-1) Drug content found*(mg) Recovery* (%) Average recovery (%)
Furoject 20 mg 2 20.85 104.25 106.56
Injection Turkey 4 20.60 103.00
8 22.49 112.45
LASIMEX 40 mg 2 41.93 104.82 105.28
Tablets S.D.I. - 4 42.46 106.15
IRAQ 8 41.95 104.87
Evaluate the results with the standard addition method (SAM). To prove the efficiency of the recommended method and its success in estimation and that it is free from the effect of the additive compound, the standard addition method was applied to estimate the drug compound (furosemide) and due to the
lack of requirements for the standard method approved in the British Pharmacopoeia, it can be inferred from the results shown in Fig. 10 and Table 7 that the obtained method is compatible with the suggested method, which indicates that the method has good selectivity.
Table 7. Comparison of the accuracy of the suggested method for the micro-determination of furosemide in pharmaceutical dosage with the standard addition method
Pharmaceutic al preparation Certified value (mg) Amount present (^g. ml-1) Drug content found (mg) Recovery (%) of standard addition procedure
Present method* Standard addition procedure
Furoject Injection Turkey 20 2 20.85 20.93 104.65
4 20.60 20.10 100.50
iigmL1
Fig. 10. Standard addition curve for the determination of furosemide in a pharmaceutical
preparation
A comparison of the method. The
developed technique was compared with other published UV-Vis spectrophotometric methods based on the same reaction (oxidative coupling reaction). The results confirmed that the developed method has a greater sensitivity compared with published papers. Furthermore,
this method is straightforward and does not involve any extraction steps. In addition, the applications of the method are broader than those mentioned in the literature. It includes the determination of the drug compound in tablets and injections Table 8.
Table 8. Comparison of the suggested UV-Vis spectrophotometry methods with the published
methods for Furosemide determination.
Parameter Current method Literature method [18] Literature method [19]
Type of reaction oxidative coupling reaction oxidative coupling reaction oxidative coupling reaction
Reagent Chlorpromazine 1 -Naphthylamine-4-sulfonic acid 2,4-di-nitrophenyl hydrazine
Maximum wavelength (nm) 525 465 467
Medium of reaction Acidic Alkaline Basic
Beer's law fag /ml) 2-32 3-23 0.4-14
Molar absorptivity (l/mol.cm) 138891 10650 11890
Application Injection and Tablet Tablet Tablet
Conclusion
A rapid and simple spectrophotometric method was developed for the determination of furosemide based on the oxidative coupling reaction and measuring the product formed at
525 nm. The drug was estimated in the range of 2-36 |ig/ml of furosemide with good accuracy, as the recovery rate was 101.15% and precision was less than 0.5%. The method was applied
successfully in the estimation of pharmaceutical preparations, as it was found from the study of the nature of the complex that the complex is formed in a ratio of 1:1 (drug:reagent). The
developed method was characterized by ease, sensitivity, and the absence of the need for prior extraction
A cknowledgement
All authors would like to extend their thanks to the College of Education for Girls and the College of Education for Pure Science/ Mosul University for the purchase of chemicals and for providing the necessary laboratory facilities.
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