Analysis of Copper (II) in Pharmaceutical Products using Micelle form with the Turbidity Method

 

Shaymaa Ibrahim Saeed¹, Ahmed Fadhil Khudhair²*, Sawsan Khudhair Abbas³ , Hasan F. Alesary⁴

Chemistry Department, College of Science, University of Kerbala, Karbala, Iraq

 

ABSTRACT:

A modern and easy procedure performed using the turbidity method for the pre-concentration and estimating of Cu (II) in pharmaceutical samples. The limited solubility complex formed by the reaction of copper ions with cysteine and SDS as the surfactant. The copper content measured using the turbidity method. The effect of chemical and physical parameters including grouping of the ligand, SDS, pH, equilibration temperature and time and effect of salting are studied. The analytical qualities of the technique have been determined, which includes linear range (the range was between 0.158 -47.625µgmL^‑1), the coefficient of turbidity (k=13.21), limit of detection (LOD=0.1767 µg ml^‑1nd LOQ=0.5355 µg mL^‑1).All values are calculated using statistical programs, but the detection limit for Cu(II) ions based on eight times the standard deviation of the blanks (N:8) characterized as 3Sb/m (where Sb is the standard deviation of the blank and m is the slope of the calibration curve) is 0.0096μg mL⁻¹ (3σ). The method employed successfully for the determination of Cu²⁺ in different local pharmaceutical formations.

 

 

 

INTRODUCTION:

Copper is an important element for plants, animals, and humans. The human body contains between 1.4 to 2.1 milligrams of copper per kilogram of body mass [1]. It undergoes a number of processes in the liver then distributed to other tissues in the body, including the ceruloplasmin protein, which transfers most of the copper in the blood [2]. High levels of copper can also lead to the development of Alzheimer's disease [3]. Also one of the elements that have the ability to interact with certain sulphur compounds and form different types of copper sulphate [4]. In addition to its behaviour as a Lewis acid in many complexes when interacting with appropriate reagents, amino acids considered to have the ability to form stable complexes with copper [5]. The composition of the complex is therefore a common way of estimating low concentrations of copper (II) using different techniques.

 

 

Some researchers have used the chromatic method to detect copper ions via cysteine oxidation [6]. In literature survey, the cloud point extraction (CEP) method used successfully to pre-concentrate the organic compounds and the inorganic compound in natural and the biological sample like measurement of strontium (II) ion as trace amount in water samples [7]. Determination trace iron (III) concentrations in urine samples [8]. Cloud point extraction incorporated the turbidity technique to estimate Bi³⁺ in medicinal samples [9]. While copper ion was determined in urine sample by FAAS method after pre-concentration by CPE method [10], solid phase extraction as a pre-concentration copper (II) in gasoline [11]. In addition, the copper determine by CPE-FAAS that depend on formation of slightly water-soluble complex [12]. Electrochemical determination in spirit drinks [13] and by flow injection analysis after extraction using solid phase in fresh water samples [14], where the copper ion determine in pharmaceutical product by deferent instrumental techniques for example extraction and spectrophotometric determination [15, 16], Liquid micro-extraction with FAAS [17,18] and determine by turbidity method in water sample [19]. Where using the micelle formation in the many adsorption methods [20,21], determine trace Cu (II) as micelle-mediated extraction [22], surfactant micelles mode coupling with spectrofluorimetric analysis [23] and coupling with turbidity analysis [24]. Also the micelle form used for brominating reactions [25] and as delivery system for some drugs [26-29].

 

Here, copper ion concentrations estimated in pharmaceutical preparations using the turbidity technique through a complex composition that is very sparingly soluble with cysteine and the presence of SDS that keeps the complex suspended in solution for a long time.

 

EXPERIMENTAL PART:

^{The chemicals used in this study are of high purity and distilled water used to make up all solutions. Stock copper (II) ions was formed by dissolving 0.19 g copper (II) sulphate (CuSO4.5H2O, Merck, 100 µgmL-1) in 500 mL of distilled water. An aqueous solution of cysteine (2-Amino-3-sulfhydrylpropionic acid), (HO₂CCHCH₂SH, was formed by dissolving 1g in 100mL of distilled water. Sodium dodecyl sulphate stock solution (SDS, 288.3 g mol-1, 4% w/v) was made up by dissolving 10g in 250 mL of distilled water, whilst sodium hydroxide solution (NaOH, Fluka, 1.0 mol L-1) was made up by dissolving 4 g in 100 mL (standardized with HCl solution).}

 

^{An FT-IR Shimadzu model 8400 device was used to record infrared spectrum utilizing a KBr disc (4000-400 cm-1), and a Lovibond turbidimeter (Germany) device was used to measure the turbidity of the solution while the acid function was measured by a pH meter (a Bench Top).}

 

Turbidity and Nephelometry Unites (NTU):

The Formazine polymer compound used as standard compound to calibration a turbiditmeter, so it is use as the same unit of attenuation of source light. In 1926, King Sbury and Clark discovered formazan, which formulated completely of traceable raw material, which drastically improved the consistency in standard formulation. Formazine is suitable for the suspension of turbidity stands when prepared by the reaction of hydrazine sulphate with hexamethylene tetraamine, as shown in Figure 1[30].

 

 

Figure 1 Synthesis of Formazin.

 

 

Figure 2 The proposed mechanism of rich phase of micelle formation

Table.1: Solubility information of Cu-Cys complex

Solvent	Water	ethanol	methanol	acetone	Ether	DMF	DMSO	CCL4	Benzene
Cu-Cys	IS.	IS.	IS.	IS.	IS.	IS.	IS.	IS.	IS.

IS: slightly soluble

 

Cu(II)-Cys Complex Formation:

For the complex emergence, 10, 000µLaquatic solution including Cu(II) ions(15µg mL^-1), 0.7 µL of 1% Cysteine solution, 200 µL of 4.0% (m/v) SDS solution, the volume of which is made up by deionised water to 1000 µL. The turbidity of the solution was determined by transferring of the solution to the cell of the turbidmeter at room temperature, but In the case of solubility tests and FT-IR, the complex was formed by mixing 3ml for a 5×10^-3M of Cu(II) ion with 2 mL of 1% Cysteine solution, then making the volume up to 10, 000 µL with deionized water. The complex was then isolated by a centrifuge, filtered and left to dry at room temperature. Figure 2 shows the suggested mechanism of the complex formation with the turbid solution after addition of the SDS surfactant.

 

Complex Solubility test:

The Cu(II)-Cys complexes, as reported in Table 1, were insoluble in water, ethanol, methanol, acetone ether, dimethyl formamide (DMF) dimethyl sulfoxide (DMSO), CCl₄ and benzene. All solubility results are identical to the results of previous research.

 

FT-IR Spectra:

The FT-IR spectra were taken for each the complex Cu (II)-Cys and L-cysteine reagent were taken from that can be observed that all the values found in the ligand were present within the complex and some of them had higher or lower frequency for the ligand spectrum. Some beams, such as thiol and some new bundles, through a sulphuratom, not oxidized by copper, and converted to cysteine. The IR spectra matched those obtained by other researchers[31]. The FT-IR assignments of the complex and ligand is shown in Figures 3 and 4.

 

Table 2: Infrared band assignments of L-Cysteine and Cu-Cy complex

L-Cysteine ν, cm-1	Cu-Cy complex ν, cm-1	Assignments
3039	--	-OH stretch
--	3433	-OH stretch
2708	--	-NH stretch
--	2924	-NH stretch
2554	--	-SH stretch
1340	1344	-CO stretch

 

The conditions of measurement:

In order to maximum extraction efficiency of complex Cu-Cy using the turbidity method. The most important effects were found to be order of addition, pH of the solution, ligand concentration, SDS concentration, effect of the salt, effects of temperature, and time.

 

Order Addition arrangement:

The order arrangement of the reactants was studied by varies arrangement of the addition to obtain the rich phase product at the experimental options where the outcomes are appeared in Table 3 from that can appear the arrangement of solution number 1 is the best chose.

 

 

Figure 3: Spectrum FT-IR of Ligand

 

Figure 4: Spectrum FT-IR of complex

 

Cysteine concentration:

The effect of the concentration of cysteine on attenuation of incident light was estimated by varying from 0.01% to 0.15 w/v%. Table 4 outlines the outcomes obtained and Figure 5(a) demonstrates that 0.07% of cysteine is the ideal focus.

 

Table.4 the effect Cysteine concentration on the attenuation of light [Conditions: 0.25 mmol L^-1=15.875 of Cu (II); 0.08% w/v SDS]

Copper (M), SDS(S), Cysteine (Cy)

 

 

Figure 5(a) Varying of Cysteine concentration on the measurement of turbidity

 

The SDS concentration:

The concentration of surfactant utilized as a part of the turbidity estimation is a vital variable of SDS surfactant. Additionally, it gives a richly complex phase in a suspended state. The effect of surfactant concentrate on turbidity estimation was analysed after surfactant added to increase the homogenously of the turbid solution. Table 5 and Figure 5(b) show signal of attenuation of incident light of the rich phase with expanding grouping of the SDS molecules. 0.08% is indicating the best concentration could be chose because the amount of turbid product is increase.

 

 

Figure 5(b) Turbidity measurement with concentration of surfactant

 

pH study:

The acidic or basic medium for metallic complex formation is imperative requiredin metal particles at the turbidity measurements. On the other handthe reaction should be having enough hydrophobicity to remove the little amount of the surfactant-. The pH effect measurement assumesthata metallic complex to form which is preferred the acidic medium and consequent extraction.Therefore this test- was done within pH range from 2.0 to 12.0 by using distinctive pH support arrangements 0.1N HCl and 0.1N NaOH.

 

 

 

Table 5 The measured turbidity with the concentration of surfactant [Conditions: 0.25 mmol L^-1 of Cu (II); 0.07% aqueous solution of Cysteine]

 

 

Figure 6. Effect of varying pH on the formation of Cysteine-Cu²⁺ complex.[Conditions: Cu²⁺ = 0.25 mmol L^-1; 0.07% aqueous solution of Cysteine; (w/v) 0.08% w/v SDS]

 

Figure 6shows the values of turbidity increase with the pH of the solution increase to pH 4.42 then the turbidity of the product decrease with rising the pH that conform the complex decomposition, So that as evidenced by the incomplete extraction of the complex in the neutral and alkali mediums from the decrease in turbidity values. Subsequently, at pH 4.42 was picked as the ideal working pH to producethe final Cu(II)-Cysteine complex and consequently a reasonable yield.

 

Temperature and Time:

Temperature is one of the factors affecting extraction efficiency, so the effect of a range of temperatures was studied from 10°C to 80°C with most efficient extraction of the copper ion found at 35°C, which is satisfactory complete the extraction and formation of suspended complex. The effect of incubating the solution was also tested in the same optimum of temperature for interval sranging from 5min to 45 min, where it was ultimately found that 15 min is the best time to rich phase form. Figures 7 and 8 show the effect of temperature with incubation time on the extraction and development of the complex using the turbidity technique while Figure 9 shows that the complex is stable for a long period of time as a suspension in solution.

 

 

Figure 7 Change in turbidity with temperature [Conditions: Cu (II) = 0.25 mmol L^-1; 0.07% aqueous solution of Cysteine; (w/v) 0.08% w/v SDSat pH 4]

 

 

Figure 8 Change in turbidity with incubation time. Conditions: Cu (II) = 0.25 mmol L^-1; 0.07% aqueous solution of Cysteine; (w/v) 0.08% w/v SDS at pH 4 and 35 ͦC].

 

 

Figure 9: Effect of time of leaving after heating on the measurement of attenuation of incident light

The Salt Addition Effect:

The salt contamination effect is another key parameter in estimations of turbidity. The addition of salt can effect on the extraction where the fluid phase for most non-ionic surfactants and surprisingly encourage phase lack of involvement. Furthermore, non-polar analytes may turn out to be less solvent in the arrangement at higher salt fixations and in this way add to higher turbidity. The results show that the salt added creates an increment in the extraction. The efficiency of the suggested method in the extraction and determination of copper ion levels using the turbidity method in the presence of different concentrations of salts was examined using a solution containing mg.L^-1of copper with the addition of different concentrations of salts. It shown that KCl had no effect on the turbidity of the complex, but MgCl₂and NaCl increased turbidity, as shown in Figure10.

 

 

Figure 10 Effect of salt contamination on the turbidity of the complex (Cu (II) mg.L⁻¹ at 178 NTUand 35 ͦ C) using the turbidity method

 

Repeatability:

Table 6 shows the average values and RSD% of 15.89mgL^-1 Cu (II) and at the optimum conditions The RSD% values were less than 1% indicating a reliable measurement can be achieved for Cu (II).

 

 

 

 

Table.6Repeatability of Cu²⁺ at best conditions.

[Cu] mg L-1	Average	RSD%	ȳi ±t0.05/2, n-1 σn-1/at 95%
15.89	176.38	0.294	176.38±0.433

 

The Method Characteristics:

Under the ideal conditions determined above, the calibration curve of attenuation of incident light method was assessed. An 0.158 - 47.625µgml^‑1 Cu(II) solution was prepared. Table7 reports the results of the linear regression analysis

 

 

Figure 11: Calibration curve for the variation of Cu(II) ion concentration on NTU using the turbidity instrument

 

Application:

Two methods are used for the determination of the copper ion concentration in pharmaceutical preparations (copper, 0.002 g /0.6 g). The first was the turbidity method while the second method was performed via UV-Vis spectrophotometry. Table 9 shows the summary of results of the samples.

 

 

 

 

 

 

Table 7: Calibration Curve parameters

 

Table 8 Copper (II) ion determination in local pharmaceutical samplesusing the proposed turbidity method and standard UV-Vis method

CONCLUSION:

This paper uses the micelle form via a turbidity technique to allow for the determination of low concentrations of copper ions in complexes that were only slightly soluble in most of the available solvents. The new method is used to determine Cu (II) in pharmaceutical preparations after optimization of the formation of a suspension of complex in each solvent. This work represent proposed method have some the advantages like; simplicity, availability material, low consume of materials and the waste is friendly to– environment as a green chemistry, in addition to the possibility - to estimate copper ions in - slightly soluble complexes.

 

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Received on 12.01.2019          Modified on 21.02.2019

Accepted on 29.03.2019        © RJPT All right reserved