Flow injection–chemiluminescence determination of sulfadiazine in compound naristillae

Talanta 72 (2007) 1036–1041

Flow injection–chemiluminescence determination of sulfadiazine in compound naristillae Haiyan Liu a , Juanjuan Ren b , Yuhong Hao b , Pingang He b,∗ , Yuzhi Fang b,∗ a

School of Chemistry and Molecular Engineering, East China University of Science and Technology, Shanghai 200237, China b Department of Chemistry, East China Normal University, Shanghai 200062, China Received 6 September 2006; received in revised form 18 December 2006; accepted 18 December 2006 Available online 12 January 2007

Abstract A simple, sensitive and selective flow injection-chemiluminescence method for the determination of sulfadiazine in compound naristillae has been investigated. It is based upon the chemilimunescence reaction of sulfadiazine, formaldehyde and potassium permanganate in polyphosphate acid medium. The optimum conditions for the chemiluminescence emission were investigated. Under the optimum conditions, the linear range for the determination of sulfadiazine was 8.0 × 10−7 to 2.0 × 10−4 mol/L with a detection limit of 2.0 × 10−7 mol/L calculated as proposed by IUPAC and a relative standard deviation of 2.53% for 11 solutions of 5.0 × 10−5 mol/L sulfadiazine on the same day. It was also found that the coexisting ephedrine hydrochloride did not interfere with this determination. This led to the successful application of the proposed method for the direct and selective determination of sulfadiazine in compound naristillae. © 2007 Published by Elsevier B.V. Keywords: Flow injection; Chemiluminescence; Permanganate; Compound pharmaceutical; Sulfadiazine

1. Introduction Chemiluminescence (CL) is known as high sensitivity, low detection limit and wide linear ranges with very simple instrument. Flow-injection analysis (FIA) coupled with CL (FIA–CL) has grown up rapidly in pharmaceutical analysis for rapid, automated and precise analysis at present. However, the only apparent limitation is the fact that FIA–CL manifolds do not allow for the analysis of multi-component samples thus limiting their practical utility due to its poor selectivity of the CL detection itself because many coexisting substances can react with CL reagents to give light emission which resulted in interference. In order to improve selectivity of CL, pre-treatment or separation procedures are usually required for the selective determination in FIA–CL. Many analysts made great efforts including coupling with high performance liquid chromatography [1,2] or capillary electrophoresis [3–6], using molecularly imprinted polymer [7], using partial least squares (PLS) cali-

∗

Corresponding authors. Tel.: +86 21 62233508/798; fax: +86 21 62233508. E-mail addresses: [email protected] (P. He), [email protected] (Y. Fang). 0039-9140/$ – see front matter © 2007 Published by Elsevier B.V. doi:10.1016/j.talanta.2006.12.048

bration [8] or high selective CL system [9]. Knecht et al. [10] presented a parallel affinity sensor array (PASA) for the rapid automated analysis of 10 antibiotics in milk, using multianalyte immunoassays with an indirect competitive ELISA format. Wang and co-workers [11] reported a novel configuration of a lab-on-valve (LOV) system, which was applied for CL detection tetracycline in milk by integrating a demountable Z-type flow cell onto the LOV unit. In addition, the selectivity for tetracycline was improved by employing solid-phase extraction sample pretreatment. Sulfadiazine (Fig. 1a) and ephedrine hydrochloride (Fig. 1b) compound naristillae is one kind of hospital preparation, which includes sulfadiazine and ephedrine hydrochloride as its effective components. Sulfadiazine is a kind of sulfonamide with medium effect and broad spectrum antibacterial effect. Ephedrine hydrochloride is a kind of imitating epinephrine drug. Sulfadiazine can be determined using acidic KMnO4 /H2 SO4 CL system after online photochemical reaction with a detection limit of 80 ␮g/L [12] and ephedrine hydrochloride can be determined using KMnO4 /H6 P4 O13 /␤-cyclodextrin after HPLC separation with a detection limit of 10 ␮g/mL [13]. Acidic potassium permanganate is one of the most important oxidants applied in CL reactions and a great number

H. Liu et al. / Talanta 72 (2007) 1036–1041

Fig. 1. The structures of sulfadiazine (a) and ephedrine hydrochloride (b).

of investigations have been published [14–22]. Pasekov´a et al. [20] reported a sequential injection-CL method to individually determine a group of sulfonamides using potassium permanganate–glutaraldehyde–sulfuric acid system. A detailed review reported by Hindson and Barnett [21] showed that most of the compounds containing a phenolic and/or amine moiety elicited a CL response in acidic potassium permanganate system. It was also reported that formaldehyde could enhance the CL emission intensity of such system. Liao et al. [22] investigated the CL reaction mechanism, they supposed that HCHO did not change the emitter of the CL reaction, but increased the luminescence quantum number, the proper HCHO concentration increased the number of singlet oxygen of the CL reaction system, and excited molecular oxygen is a more likely emitter. To the best of our knowledge, there is not any report about selective determination of sulfadiazine when ephedrine hydrochloride was present by FIA–CL up to now. Then a simple, rapid FIA–CL method was developed, which was successfully applied to selectively determine the content of sulfadiazine in the compound naristillae. Under the optimum conditions, the relative CL intensity is linear with the concentration of sulfadiazine in the range of 8.0 × 10−7 to 2.0 × 10−4 mol/L with a detection limit of 2.0 × 10−7 mol/L (S/N = 3), giving a throughout of about 120/h. 2. Experimental 2.1. Apparatus and reagents CL intensity was recorded by IFFL-D flow-injection luminescent analyzer (Xi’an Ruike Electronic equipment Corporation, Xi’an, China). The schematic diagram is shown in Fig. 2. Two peristaltic pumps were used to deliver flow streams in this system. Polytetrafluoroethylene tube (0.8 mm i.d.) was used as

Fig. 2. Schematic diagram of the flow-injection CL system for the determination of sulfadiazine: (a) sample solution; (b) HCHO carrier stream solution; (c) polyphosphoric solution; (d) KMnO4 ; P1 and P2, peristaltic pump; V, eight-way injection valve; Y, Y-shaped mixing element, F, CL flow cell; PMT, photomultiplier tube; HV, negative high-voltage supply; PC, computer; W, waste solution.

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connection material in the flow system. A portion of sample or work solution of sulfadiazine was injected into the HCHO carrier stream using an eight-way injection valve equipped with a sample loop, merged with the mixture solution of KMnO4 solution and polyphosphoric solution just before a spiral flow cell, and then CL was emitted in the flow cell. The CL signal in the flow cell was detected with an R456 Photomultiplier tube, the negative high voltage of which was set at −800 V, and collected with the computer employing IFFL-D flow-injection CL analysis system software. All reagents were of analytical grade, purchased from Shanghai chemical reagent company and used without further purification. The water used throughout was double distilled. Sulfadiazine stock solutions (1.0 mg/mL), stored at 4 ◦ C, were prepared everyday by dissolving 0.0050 g standard sample (≥99.5%, friendly provided by Shanghai Sunve Pharmaceutical Co. Ltd.) in 2.0 mL of 1.0 mol/L HCl solution, sonicated, diluted to 5.0 mL in a brown measuring flask with water. Working solutions were prepared from the stock solution by appropriate dilution with water before use. A stock solution of KMnO4 (0.05 mol/L) was prepared by dissolving 3.951 g KMnO4 in 500.0 mL of distilled water, which was heated up to boiling and maintained slight boiling status for 1 h, then cooled, filtrated with fiberglass and stocked in a brown reagent bottle. A solution of HCHO (0.56 mol/L) was daily prepared. The stock solutions of HNO3 , H3 PO4 , H2 SO4 , HCl (1.0–5.0 mol/L) were prepared separately. The stock solution of H6 P4 O13 was daily prepared. Sulfadiazine and ephedrine compound naristillae, the prescription of which includes 50 g of sulfadiazine, 10 g of ephedrine hydrochloride, 6 g of sodium chloride, 5 g of sodium carboxymethyl cellulose, 3 g of sodium benzoic, was purchased from Eye Ear Nose and Throat Hospital of Fudan University. 2.2. Procedures 2.2.1. Procedures for FIA and static CL test In order to obtain good stability, the instruments were run for at least 10 min before the first measurement. Flow tubes were inserted into (a) water; (b) 0.56 mol/L HCHO carrier stream solution; (c) 0.7 mol/L polyphosphoric solution; (d) 0.5 mmol/L KMnO4 , respectively. The flow rate of P1 was set at 2.0 mL/min for line (a) the flow rate of P2 was set at 2.5 mL/min for lines (b–d). Pumps were started to wash the whole system until a stable blank signal was recorded. A 100 ␮L of sample solution was injected into a carrier stream. This stream was merged with KMnO4 /polyphosphoric solution, which was mixed on line, and then reached the flow cell. The concentration of sample was quantified by the relative CL intensity I (defined as the difference of CL intensity between in the presence and in the absence of sulfadiazine, respectively). The static system for the detecting kinetics characteristic of the CL reaction was composed of a quartz cell (2 mL) and the IFFL-D ultra-sensitive luminescence analyzer. The cell was located directly on the window of the R456 Photomultiplier tube in the luminescence analyzer. 100 ␮L of 0.5 mmol/L KMnO4 was injected into the cell, which contained 100 ␮L of 1.0 × 10−4 mol/L sulfadiazine or water, 100 ␮L of 0.7 mol/L

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H. Liu et al. / Talanta 72 (2007) 1036–1041

polyphosphoric acid and 100 ␮L of 0.56 mol/L HCHO mixture solutions for signal/blank respectively, and at the same time, the CL signal was recorded. Data acquisition and treatment were still performed with IFFL-D software. 2.2.2. Preparation of sample solution Sulfadiazine and ephedrine compound naristillae is a kind of suspension because sulfadiazine is undissolvable in water. A whole bottle of naristillae (10 mL, contained 0.5 g sulfadiazine) was quantitatively transferred into a 100 mL of brown measuring flask, then added 40.0 mL of 5.0 mol/L HCl solution, sonicated, diluted with water to scale, stored in refrigerator and avoid light. An appropriate volume of above sample solution was diluted further with water so that the concentration of sulfadiazine in the final solution was within the working range, and then analyzed according to the procedure described in Section 2.2.1. 3. Results and discussion 3.1. Characteristics of CL reaction The CL profiles using static method for the mixtures of sulfadiazine, HCHO, KMnO4 and polyphosphate acid solution are shown in Fig. 3. It is shown that the mixing of KMnO4 , H6 P4 O13 with sulfadiazine could give very weak CL emission (line a), while the mixing of KMnO4 , H6 P4 O13 with HCHO could give weak CL emission (line b), and when sulfadiazine was mixing with KMnO4 , HCHO and H6 P4 O13 , a strong CL emission was recorded (line c). Previous investigations showed that formaldehyde could significantly enhance the acidic KMnO4 CL reactions [19,22]. In our study, it was also found that HCHO can greatly enhance the CL emission of

KMnO4 –H6 P4 O13 –sulfadiazine (Fig. 3, line a and c). The possible mechanism may be that much more 1 O2 was generated when HCHO was present, and hence increased luminescence quantum number. Fig. 3 also indicated that the maximum emission intensity was attained within 1 s and then attenuated rapidly. 3.2. Optimization of the experimental conditions To select the reaction parameters that give the higher sensitivity and optimum signal/blank ratio which can further be used in the determination of sulfadiazine in real sample, a series of univariate searches were performed on FIA–CL system, including reagent concentration, reaction medium, reagent flow rate and injection sample volume. 3.2.1. The optimum of FIA–CL system In order to avoid the erosion to valve from acid in carrier, we devised three systems to test. Table 1 is the effect of different FIA–CL systems on the relative CL intensity and signal/blank ratio. It is shown that the system 1 can give the highest sensitivity and the best signal/blank ratio in the three kinds of FIA–CL system. Although the blank signal of system 3 was the smallest, its relative CL intensity and signal/noise was also the smallest owing to the dilution action of water carrier. So the system 1 was adopted in the following experiment. 3.2.2. Selection of acid medium and the optimum of their concentrations The kinds and concentration of acid in the reaction system influence the CL emission intensity. Therefore, five different acids including HCl, HNO3 , H2 SO4 , H3 PO4 and H6 P4 O13 of different concentration were tested respectively. It was shown in Table 2, the highest relative CL intensity and the best signal/blank ratio was obtained in 0.7 mol/L of H6 P4 O13 . Although it was reported that H6 P4 O13 was apt to hydrolysis [21,22], considering H6 P4 O13 was prepared just before use, its hydrolysis can be ignored. So 0.7 mol/L of H6 P4 O13 was selected as acid medium in the experiment. 3.2.3. The optimum of HCHO concentration From Fig. 4 line a, it can be concluded that when the concentration of HCHO is between 0.56 and 1.22 mol/L, the system Table 1 The effect of system on the CL signal

Fig. 3. The characteristics of CL reaction (a) CL intensity in the absence of HCHO; (b) CL intensity in the absence of sulfadiazine; (c) CL intensity in the presence of sulfadiazine and HCHO. Procedures for static CL test: 100 ␮L 0.5 mmol/L of KMnO4 was injected into the cell, which contained 100 ␮L 1.0 × 10−4 mol/L of sulfadiazine or water, 100 ␮L 0.7 mol/L polyphosphoric acid and 100 ␮L 0.56 mol/L HCHO or water mixture solution, the negative high voltage of PMT, −800 V.

System

I0

I

I

I/I0

1a 2b 3c

16 17 10

122 120 45

106 103 35

7.6 7.1 4.5

Conditions: KMnO4 , 0.5 mmol/L; H6 P4 O13 , 0.7 mol/L; sulfadiazine, 5.0 × 10−5 mol/L; HCHO, 0.56 mol/L; the flow rate of line b–d: 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V. a The schematic diagram is shown in Fig. 2. b In system 2, the sample is as carrier, and HCHO is injected into sample carrier. c In system 3, sample and HCHO are online mixed and injected into water carrier; I0 , the blank of CL intensity; I, the signal of CL intensity.

H. Liu et al. / Talanta 72 (2007) 1036–1041

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Table 2 The effect of acid medium and concentration on CL Acid kinds

Concentration range tested (mol/L)

Optimum concentration (mol/L)

I0

I

I

I/I0

HCl HNO3 H2 SO4 H3 PO4 H6 P4 O13

0.2–1.0 0.2–1.5 0.2–1.0 0.5–5.0 0.2–1.0

0.5 1.2 0.5 2.0 0.7

6 5 5 3 4

29 25 16 37 75

23 20 11 34 71

4.8 5 3.2 12.3 18.8

Other conditions: KMnO4 , 0.5 mmol/L; HCHO, 0.56 mol/L; sulfadiazine, 5.0 × 10−5 mol/L; the flow rate of line b–d, 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V.

3.3. Analytical characteristics Under the optimum conditions, the average value of relative CL intensity was linear with the concentration of sulfadiazine from 8.0 × 10−7 to 2.0 × 10−4 mol/L and three replicative injections were performed for each standard solution. The regression equation was I = 29.13 + 0.074 × 107 C (I is the relative CL intensity and C is the concentration of sulfadiazine, mol/L) with a correlation coefficient of 0.9986. The detection limit was 2.0 × 10−7 mol/L which was calculated according to IUPAC definition that is three times of standard deviation of blank value [23,24]. The relative standard deviation for 11 solutions of 5.0 × 10−5 mol/L sulfadiazine under the optimum conditions on the same day was 2.53%. 3.4. Interferences experiments Fig. 4. The effect of HCHO concentration on CL. Conditions: KMnO4 , 0.5 mmol/L; H6 P4 O13 , 0.7 mol/L; sulfadiazine, 5.0 × 10−5 mol/L the flow rate of line b–d: 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V.

can give higher relative CL intensity. However, the signal/blank ratio is decreased while the concentration of HCHO is increased (Fig. 4 line b). As a result, 0.56 mol/L of HCHO was selected at last as a compromise between sensitivity and signal/blank ratio.

Considering that the developed method would be applied to determine sulfadiazine in compound naristillae, the interference effect of common ions and all coexisting components in compound naristillae were assessed. Samples containing sulfadiazine at a fixed concentration of 5.0 × 10−5 mol/L and

3.2.4. The effect of KMnO4 concentration The effect of KMnO4 concentration on CL intensity is shown in Fig. 5. The relative CL intensity reaches a relative stable value when its concentration is between 0.5 and 0.9 mmol/L. So 0.5 mmol/L was accepted as an optimum concentration in this study. 3.2.5. The effect of physical parameter of FIA–CL The flow rate and sample volume can affect the CL intensity, precision, and the flow rate can also influence the rate of analysis. Since the CL reaction is rapid, the shortest distance between the valve and flow cell (6 cm) was firstly applied in this study. Then the flow rate of pump P2 from 1.5 to 3.5 mL/min and sample volume from 55 to 150 ␮L were separately investigated in order to collect the maximum CL signal. When the flow rate was 2.5 mL/min and sample volume was 100 ␮L, the relative CL intensity, signal/blank ratio, and reproducibility of signal was the best. So, the flow rate of 2.5 mL/min and sample volume of 100 ␮L was chosen as the optimum in the experiment.

Fig. 5. The effect of KMnO4 concentration on CL intensity. Conditions: HCHO, 0.56 mol/L; H6 P4 O13 , 0.7 mol/L; sulfadiazine, 5.0 × 10−5 mol/L the flow rate of line b–d: 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V.

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H. Liu et al. / Talanta 72 (2007) 1036–1041

Table 3 Tolerance limit of some foreign substances on the determination of sulfadiazine Substances

Concentration ratio to sulfadiazine K+ ,

Na+ ,

Zn2+ ,

+,

Ni2+ ,

Cr3+ ,

Ca2+ ,

Al3+ ,

Cl− ,

Lactose, glucose, starch, citrate, PVP, NH4 C2 O4 Maltose, galactose, cyclodextrin, carboxymethyl cellulose, ephedrine hydrochloridea , Cu2+ Benzoic sodiuma Sucrose, riboflavin

≥200 50 10 5

2−

Conditions: KMnO4 , 0.5 mmol/L; HCHO, 0.56 mol/L; H6 P4 O13 , 0.7 mol/L; sulfadiazine, 5.0 × 10−5 mol/L; the flow rate of line b–d: 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V. a The maximum ratio tested. Table 4 Detrmination of sulfadiazine in compound naristillae Sample

Proposed methoda Sulfadiazine supplement (10−5 mol/L)

1 2 3

Pharmacopoeia methoda Found (10−5 mol/L)

Recovery (%)

R.S.D. (%)

0 4.00

4.06 8.09

100.8

2.3

0 6.00

6.61 12.5

98.2

2.2

0 8.00

8.04 16.25

102.6

2.0

Content labeled (g/bottle)

Content (g/bottle)

Content (g/bottle)

R.S.D. (%)

0.508

0.5026

0.25

0.50

0.491

0.4985

0.30

0.50

0.503

0.5010

0.40

0.50

Conditions: KMnO4 , 0.5 mmol/L; H6 P4 O13 , 0.7 mol/L; sulfadiazine, 5.0 × 10−5 mol/L; HCHO, 0.56 mol/L; the flow rate of line b–d: 2.5 mL/min; sample volume, 100 ␮L; the negative high voltage of PMT, −800 V. a Average of five measurements.

increasing concentration of the interferences were analyzed by the method under the optimum conditions. The tolerable limit of a foreign species was taken if it caused a relative error of less than 5%. The obtained results in the Table 3 showed that under the optimum conditions, the common ions, ephedrine hydrochloride and the other coexisting components at concentrations in the naristillae did not interfere with the determination of sulfadiazine. Although it was reported that ephedrine hydrochloride can be determined using KMnO4 /H6 P4 O13 /␤-cyclodextrin after HPLC separation with a detection limit of 10 ␮g/mL [12], in our proposed system, it did not interfere with the determination of sulfadiazine when its concentration was more than 50 times of sulfadiazine, which was only 1/5 quantity of sulfadiazine in the prescription. This may be due to the different sensitizer and other reagent conditions in the two systems. Owing to some sulfonamides could generate CL emission in KMnO4 /H2 SO4 /glutaraldehyde system [20], and in their structure, they have a same aniline group which was easily to be oxidized by acidic KMnO4 solution, so some of them, such as sulfanilamide, sulfacetamide, sulfadimidine and sulfamethoxazole may interfere with the determination of sulfadiazine in the proposed method. However, considering other sulfonamides were absent in the compound naristillae except for sulfadiazine, so the content of sulfadiazine in the compound naristillae could be determined after dilution without any other pretreatment. 3.5. Determination of sulfadiazine in compound naristillae Following the procedure described above, the proposed method was applied to the determination of sulfadiazine in com-

pound naristillae. The results (Table 4) agreed well with the content labeled and compared favorably with those obtained by pharmacopoeia [25]. The recovery test was also satisfactory. 4. Conclusions Based on the chemilimunescence reaction of sulfadiazine, formaldehyde and potassium permanganate in polyphosphate acid medium, a simple, rapid FIA–CL method has been developed for the selective determination of sulfadiazine. It was successfully applied to the determination of sulfadiazine in compound naristillae. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16]

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