Terbutaline sulphate (±)-α-[(tert- butylamino)methyl]-3,5-dihydroxybenzyl alcohol sulfate (I) (Scheme 1) is a beta-adrenergic radioactive substance that can be taken orally . It was used in bronchitis, breathing problems, sinusitis, obstructive airway diseases, uncomplicated preterm labor, and other conditions . In addition to what has been mentioned, it is a muscle relaxation that opens the air passage in the lungs to facilitate [3, 4].
To determine terbutaline sulfate, various analytical techniques have been reported such as HPLC [5-8], LC–MS, CE , CE–MS , chemiluminscense [11-13], and voltammetry [14, 15]. These techniques are cumbersome, time-consuming, and expensive. The spectrophotometry has been used for its low cost, simplicity, and versatility. Different spectrophotometric methods using various reagents are described for assay of Terbutaline sulfate, such as. 4-amino antipyrine  and iron  in the presence of potassium ferricyanide, eosin y , sodium borate, and treatment acetylacetone , p-aminophenol in basic medium of sodium hydroxide , 3-methyl-2- Benzothiozolone hydrochloride hydrochloride in the presence of ferric chloride , and p-chloranilic acid . This paper describes sensitive, fast, and simple methods to determine terbutaline sulfate based on the redox reactions.
Scheme 1: Terbutaline sulphate (±)-α-[(tert- butylamino)methyl]-3,5-dihydroxybenzyl alcohol sulfate (I)
Materials and Methods
Perkin-Elmer and Lambda 25 double-beam UV-visible and Genway 6300 were used as a single-beam UV-visible spectrometer, with identical 1 cm silica cells. The PGE453e type sensor scale with four digital weighing numbers was used. With a Cyber Scan 510 computer, pH measurements were performed using PH-meter with built-in glass electrode. The solutions are heated in a water bath 1003 Germany.
1,10-Phenanthroline solution (0.025 M) was prepared by dissolving of 0.450 g of 1,10- phenanthroline with distilled water and completing the volume to 100 mL in a calibrated flask. This solution was prepared daily and used immediately.
2,2'-bipyridyl solution (0.025 M) was prepared by dissolving 0.390 g of 2,2'-bipyridyl with distilled water and completing the volume to 100 mL in a calibrated flask. This solution was prepared daily and used immediately.
Standard solution of terbutaline sulfate (100 μg mL-1) was prepared by dissolving 0.01 g of pure terbutaline sulfate, which is supplied by Samara Pharmaceutical Industries (SDI), with distilled water and the volume was completed to 100 mL in a calibrated flask with distilled water and keeping in a refrigerator, and then, the solution was diluted as needed.
Fe(III) solution (0.025M) was prepared by dissolving 1.1945 g of ferric ammonium sulfate (Fe(SO4)2NH4) in a mixture of 5 mL of distilled water and 5 mL of 0.05 M HNO3, and then and diluted to 100 mL with distilled water in a calibrated flask.
Surfactant solutions, 0.1%: 0.1 g of various surfactants (neutral, positive, and negative) dissolved in 100 mL of ethanol.
Nitric acid solution (0.1 M) was prepared by diluting the concentrated solution with distilled water. The chemicals used are of the highest purity supplied by Lab Pak Chemicals (LTD), UNI-Chem, and 1,10-phenanthroline from Fluka Co.
In two series of 25 mL volumetric flasks, 0.1-2.0 and 0.1-1.2 μg mL-1 terbutaline sulfate were added separately followed by addition of 0.3 mL of Fe(SO4) 2NH4 and 2.4 mL of 1,10-Phenanthroline solutions in method A, 0.2 mL of Fe(SO4) 2NH4, 2.8 mL of 2,2'-bipyridyl solutions were added and followed by the addition of 0.4 mL of HNO3 in method B. The volumes were completed with distilled water to the mark, then mixed and placed in a water bath its temperature is set at 80 °C for 40 min in method A and at 70 °C for 80 min in method B, and then cooling and the absorbance of the complexes was measured at 510 nm and at 522 nm against corresponding reagent blank for the two methods, respectively.
Analysis of dosage forms
10 tablets were carefully powdered (each including 5 mg Terbutaline sulfate). The quantity of powder equivalent to one tablet was precisely transferred into a 50 mL beaker and dissolved in 20 mL pure water. After that, the solution was filtered through Whatman filter paper no.41. The filtrate was completed to 100 mL by pure water in a calibrated flask to obtain 50 μg mL-1 of Terbutaline sulfate in the final dilution. The analysis was performed due to the general procedure.
A bottle containing 20 mL of syrup (30 mg/100 mL terbutaline sulfate) was diluted to 100 mL with distilled water in a calibrated flask to get 60 µg mL-1 Terbutaline sulfate. Then, the stock solution was diluted and followed the general procedure.
Results and Discussion
In general, in the spectroscopic determination of many drugs, Iron salts play an important role [22-24], as they act as an oxidizing agent, the ferric ion is reduced to an amount of ferrous ion corresponding to the drug concentration. The amount of resulting Fe(II) can be estimated by complexation with 1,10-Phenanthroline in method A and 2,2'-bipyridyl. in method B. However, methods A and B are depended on the oxidation of Terbutaline sulfate drug with Fe(III) in an acidic medium and Fe(II) librated. Fe(II) interacts with 1,10-phen. to produce a red colored compound of tris-1,10-phen-Fe(II) chelate (ferroin) with maximum absorbance at 510 nm in method A, and interacts with 2,2'-bipyridyl. to produce a red-colored compound of tris-2,2'-bipy-iron(II) chelate [Fe(bipy)3]+2, with maximum absorbance at 522 nm in method B (Figure 1).
Figure 1: Absorption spectra of a) Fe(III)-o-phen with terbutaline sulphate (2 μg. mL-1) against, b) reagent blank, c) Fe(III)- 2,2'-bipyridyl with terbutaline sulphate (2 μg. mL-1) against, d) reagent blank
Optimization of reaction conditions
Several variables that affect the absorption intensity of the complexes formed, in both methods, have been studied to reach the best conditions for the estimation of Terbutaline sulfate.
Effect of temperature and reaction time
The time required for the reaction was determined by following the formation of the complexes color at different temperatures for both methods A and B. The absorbance was measured at 5- and 10-min intervals against the similarly treated blank reagent. It was found that these colored complexes form slowly at laboratory temperature and demanded a longer time for completion. The reaction was accelerated by performing the reaction at greater temperature levels. It was observed that the maximum absorption was obtained when the reaction mixture heated at 80 °C for 40 min and remained constant for 20 minutes in method A and at 70 °C for 80 min and remained constant for 80 min in method B (Figure 2). The complexes were found steady after being cooled at room temperature for more than 24 hours.
Effect of acid, pH, and buffer solution
The effect of acid on the color intensity was studied by examining 0.1 M of different acids including HNO3, HCl, H2SO4, and CH3COOH. The results indicate that the complexes in both methods A and B are affected by the addition of nitric acid, and as for the uptake it is high, as shown in Figure 3.
Figure 2: Effect of the time and temperature on the absorbance of 1 μg. mL-1 terbutaline sulphate in methods A and B
Figure 3: Effect of acid on the absorbance of complexes in the presence of 1 μg. mL-1 terbutaline sulphate
Figure 4: Effect of nitric acid volume of 0.1 M concentration on the absorbance of complex in the presence of 1 μg. mL-1 terbutaline sulphate
Figure 5: Effect of pH on the absorbance of complexes in method A and B
The effect of pH in the presence of different concentrations of nitric acid on the absorption of the complexes has been studied. The results indicated that 0.6-2.0 mL and 0.4 mL at pH 3.43-3.14 and 3.51 gave it maximum absorption in method A and B. respectively (Figure 4 and 5). The effect of buffer solution is investigated via testing different types of the regular function of pH 3.4 and 3.5 for method A and B, respectively. However, the solution became turbid.
Effect of ferric ammonium sulphate
The effect of 1 mL of different concentrations of ferric ammonium sulfate solution, while keeping a fixed concentration of Terbutaline sulfate and 1,10-Phenanthroline or 2,2'-bipyridyl were investigated on the absorption of the compounds in both methods A and B. As depicted in Figure 6, it was found that 0.3 and 0.2 mL of 0.025 M concentration of ferric ammonium sulfate gave the maximum absorption as the solution became cloudy for both methods, respectively. Therefore, these quantities were approved as ideal.
Effect of 1,10-Phenanthroline and 2,2'-bipyridyl reagents concentration
The effect of 1,10-Phenanthroline and 2,2'-bipyridyl concentrations on the absorbance of the complexes in both methods A and B, respectively, were investigated. It was found that 2.4-3.0 mL and 2.6-3.0 mL of 0.025 M of 1,10-Phenanthroline and 2,2'-bipyridyl gives, respectively, the maximum absorption. Above the mentioned concentrations, there is a decrease in the absorbance, as demonstrated in Figure 7.
Figure 6: Effect of ferric ammonium sulphate on the absorbance of complexes in method A and B in the presence of 1 μg. mL-1 terbutaline sulphate
Figure 7: Effect of reagent concentration of a) 2,2'-bipyridyl and b) 1,10-phen in the presence of terbutaline sulphate (1 μg. mL-1)
Effect of solvent dilution
The dilution effect with various types of solvents (water, methanol, ethanol, and acetonitrile) are used for the final dilution to obtain the maximum sensitivity to the compound and it has been observed that using distilled water for dilution gives the maximum absorption (Figure 8).
Effect of surfactant
Surfactant effect different surfactants (positive, negative, and neutral) affect the color intensity. The results revealed a negative effect on the absorption of the complexes in both methods A and B.
By plotting the absorptions against the concentrations, the standard curves displayed in Figure 8 were obtained, which indicate that the method follows Beer's law within the ranges reported in Table 1 for Terbutaline, which shows the possibility of estimating infinitesimal quantities, and that there is a deviation negatively about Beer's law after discretionary upper limit. The molar absorptivity, detection limit, quantitative limit, recovery %, and relative standard deviation (RSD) values indicating good sensitivity, and reproducibility. The values of the square correlation coefficient statistically, which are greater than 0.99, listed in Table 1, indicate that the standard curves have the excellent linear characteristics.
Figure 8: Effect of solvent on the absorption of the complexes in the presence of 1 μg. mL-1 terbutaline sulphate
Figure 9: Calibration graphs for the determination of terbutaline sulphate
Validity of the method
The proposed methods were applied successfully for the determination of terbutaline sulfate in its dosage forms as tablet and syrup. As presented in Table 2, the results were in agreement with the certified values indicating good recovery. To prove the efficiency of the developed methods, the standard addition procedure had been applied to the pharmaceutical preparations for Terbutaline sulfate as tablet and syrup. The method was summarized by adding increasing amounts of the pure drug standard solution, in the range of Beer’s law, to the known amount of the pharmaceutical preparation. By following the general procedure, the absorption was measured at the wavelength of 510 nm and 522 nm for methods A and B, respectively, and the obtained results were included in Figure 9 and Table 2, which indicate that the method has a good selectivity.
To know the reliability of the developed methods, both methods were compared statistically by a Student's t-test for accuracy and a variance ratio F-test for precision with the official method  at the 95% confidence level with six degrees of freedom. It was found from the results presented in Table 2, that the experimental t and F values are less than the tabular values (t=2.45 and F=6.39) at the 95% confidence level and for six degrees of freedom, indicating there was not much difference between the proposed methods and the official method.
Stoichiometry and mechanism
One of the most commonly used methods were that of continuous variation presented by Job's and molar ratio methods . In the proposed methods, the stoichiometric ratio for the oxidation of Terbutaline sulfate by Fe(III) was investigated applying the above methods using identical molar concentrations (2×10-3 M) of the drug and Fe(III) in methods A and B. The results indicated that the product was formed in the ratio of 1:4 for Terbutaline sulfate: Fe(III) in both methods. This indicates that number of moles of ferric ion depend on the number of aromatic hydroxyl groups present in the drug structure. Accordingly, the proposed mechanisms can be explained in Scheme 2.
Figure 10: Standard addition plots of terbutaline sulphate as tablet a) and syrup b) in methods A and B
Comparison of the present methods with some literature spectrophotometric methods
Table 3 presents the comparison among a number of the analytical variables for the proposed methods with that of different literature spectrophotometric methods. As listed in Table 3, the sensitivity of the proposed methods is superior to those reagents in the other reported methods that were carried in an aqueous medium. Heating was used also to prove the sensitivity.
Scheme 2: Probable mechanism of the redox reactions of terbutaline sulphate with ferric ion and complexation with 1,10-Phenanthroline and 2,2'-bipyridyl
Figure 11: The stoichiometry reaction between terbutaline and Fe(III) by a) Job's and b) mole ratio methods
It can be concluded that the suggested methods are sensitive, accurate, and precise. Method A (used 1,10-Phenanthroline) was found to be more sensitive compared with Method B (used 2,2'-bipyridyl) for the Terbutaline sulfate assay. By analyzing the dosage form of Terbutaline sulfate (tablet and syrup); the correctness of the proposed methods had been well-demonstrated. Moreover, the methods are free from interference by the common additives and excipient.
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.
All authors contributed to data analysis, drafting, and revising of the paper and agreed to be responsible for all the aspects of this work.
Conflict of Interest
The author declared that they have no conflict of interest.
Intisar Adil Shihab Al-Hammoodi
Mohammed salim Al-Enizzi
Abdussamed M. A. Saeed
HOW TO CITE THIS ARTICLE
Intisar Adil Shihab Al-Hammoodi, Mohammed salim Al-Enizz, Abdussamed M. A. Saeed. Spectorphotometric Methods for Determination of Terbutaline Sulphate in Pure and Pharamaceutical Formulation. J. Med. Chem. Sci., 2023, 6(5) 1032-1043