Spectrophotometric Determination of Carbofuran with Diazotized Benzidine in Environmental Water Samples

A simple, rapid, accurate and sensitive spectrophotometric method has been developed for the determing carbamate pesticides in both pure and water samples. The method is appropriate for the determination of carbofuran in the presence of other ingredients that are usually available in dosage forms. The effect of organic solvents on the spectrophotometric properties of the azo dye and the structure of the resulting product have also been worked out and it is found to be 1:1 benzidine :carbofuran. The method can be successfully applied to determination of carbofuran in water samples. The method is based on diazotization of Benzidine (4, 4 – diamino biphenyl) with sodium nitrite and hydrochloric acid followed by coupling with carbofuran in alkaline medium to form a yellow colored azo dye having the absorption maximum at 429nm against reagent blank solution. Beer’s law is obeyed in the concentration range of (0-14) μg of 10mL carbofuran. Molar absorptivity of 1×10 4 L.mol -1 .cm -1 which depend on the concentration level of carbofuran.


Introduction:
Carbofuran (2, 3-dihydro-2, 2-dimethyl-7-benzofuranol N-methy carbamate) is one of the carbamate pesticides, a large family of pesticides derived from carbamic acid [1] is shown in Figure (1) The toxicity of pesticides and their degradation products are making these chemical substances a potential hazard by contaminating our environment [2][3] Due to a highly toxicity of carbofuran , it is essential to develope accurate and reliable methods of monitoring their levels for safety purposes.Earlier techniques used for carbofuran assassment were including [4][5] Gas Chromatography(GC) [6][7] ,GLC [8][9] .High Performance Liquid Chromatography (HPLC) [10], and spectrophotometric techniques [11].Some of these reported methods need costly instruments, laborious procedure and less sensitive these facts have promoted the authors to develop sensitive and cost effective method.The proposed method is based on the coupling of hydrolyzed carbofuran with diazotized benzidine in alkaline medium which gives a yellow product having the absorption as medium at 429nm.

Open Access
Fig. (1) The Structure of Carbofuran [12] Materials and Methods:

Apparatus
UV-Visible recording spectrophotometer Shimadzu Model 160A (Japan) with a response time of 0.1s ,is used for spectrophotomatric determination A quartz cell of 5 mL internal volume and 1cm path length is used for absorbance measurements .
All other chemicals used in the study are of analytical reagent (AR) grade.

General
Procedure for Direct Determination of Carbofuran Using Diazotization Coupling Reaction 0.5mL from 100ppm of Benzidine transferred to 10 mL volumetric flask then 0.5mL of concentrated of 1% NaNO 2 is added Then, 0.5 mL of 1M of HCI is added.The mixture is shaken and cooled in ice bath for 5 min then added The following 0.5mL of sulphamic acid ,10ppm of carbofuran , 0.5 mL of 1M of Na 2 CO 3 are added and 0.5 mL of 0.1% of Tween 20 is added and the volume was made up to10mL with deionized water.Absorbance measurements are carried out at 429 nm for the yellow color by using a 1.0 cm quartz cell against reagent blank which is prepared in the same way but without carbofuran.

Results and Discussion:
Absorption Spectra A yellow -colored oxidizing coupling product with maximum absorption at 429 nm after studing of the Optimum Reaction Conditions is formed when carbofuran is allowed to react with benzidine in basic medium (sodium carbonate).Figure (2) shows the absorption spectra of yellow product formed so the absorption maximum at 429nm is used in all subsequent experiments.

Study of the Optimum Reaction Conditions
The effect of various parameters on the absorption intensity of the aso dye formed is studied and the reaction conditions are optimized.The factor affecting color development, reproducibility, sensitivity and conformity with Beers law is carbofuran with reagent benizdine.

Effect of Type of Acid on Diazotization
In order to select the most suitable acid used in the diazotization of carbofuran, different types of 1M acids (HCl, H 2 SO 4 , HNO 3 , H 3 PO 4 , and CH 3 COOH) are investigated in the presence of 0.5mL of (100ppm) benzidine, 0.5mL of (1%) NaNO 2 , 0.5mL of (0.1%) sulphamic acid, 1mL of carbofuran and 0.5mL of (1M) Na 2 CO 3 The maximum sensitivity and stability are obtained, when the reaction is carried out in the presence of 1mL of 1M sodium carbonate solution as shown in (Table3).

Effect of Volume of Sodium Carbonate
The effect of different volumes of ( 1M) Na 2 CO 3 solution on the absorbance of the colored product is studied keeping other conditions constant.It was found that a maximum absorbance and stable color is established with 0.5 mL of sodium carbonate (Table 4).Thus 0.5 mL of 1M Na 2 CO 3 is chosen in all subsequent experiments as shown in Figure (4).

Effect of Nitrite Time and Amount on Absorbance of Azo Dye
The effect of different amounts 0.1-0.9mL of 1% sodium nitrite with time on the absorbance of the consequential azo dye has undergone a careful study.The results in (Table 5) show that 0.4 mL of 1% sodium nitrite solution at 5 min as a reaction time is optimal and it is recommended for the subsequent experiments shown in Figure (5).

Effect of Sulphamic Acid Amount and Time
The effect of different amounts 0.1-0.9mL of 0.1% sulphamic acid with time on the absorbance of the resulting azo dye is studied The excess of nitrite must be removed by adding sulphamic solution due to high blank values.

Effect of coupling reagent
The effect of varying concentration of coupling reagent is studied using the proposed procedure and adding 0.1-0.9mL of 100 ppm of benzidine to a series solution.

Effect of Surfactant Type
The experiments are conducted at optimum established conditions according to the general procedure by using various type of surfactants such as, Tween20, SDS.CTAP and Triton X-100 at optimum concentration of 0.1% for each species.In each case, the respective λmax for each surfactant used and absorbance are calculated to choose the best one.It is observed that Tween 20 which has maximum absorbance at 429 nm is the best one for further study as shown in (Table 8)

Fig. (8).The Effect of Tween 20 Volume on Absorbance of the Complex of Carbofuran Formed with Benzidine.
The data above of

Effect of Organic Solvents
Different organic solvents are examined to evaluate their effects on the intensity of the azo dye which form the data as shown in( Table 9).

Effect of Interference
The effect of some foreign organic compounds and Inorganic compounds, often found in environmental, are studied by adding equal amount of 1mL of (10ppm) of organic compounds, Inorganic compounds and phenols to 1mL of (10ppm) of carbofuran.The color is developed by following the recommended procedure described earlier.

Effect of Temperature
The effect of temperature on the color intensity of the product is studied (Table 12).Maximum absorbance is obtained when the color is developed at room temperature From the data in Table (12) it is recommended that the color reaction should be carried out at room temperature (25°C).Maximum absorbance is obtained when the color is developed at room temperature (25°C) more than when the color is developed in an ice-bath (0°C) or in a water-bath (40 °C).and in a water-bath (50 °C).In addition a loss in color intensity and stability is observed with increasing the temperature .

Selected Optimum Conditions
After studying the effect of different parameters that have impact on the absorbance intensity of the colored product, the optimum conditions for the proposed procedure are summarized in (Table 13) and are used in all subsequent experiments.

Structure of the Dye
The stoicheiometry of the complex formed between carbofuran and benzidine is studied ,it is observed that benzidine reacts with increasing amount of nitrite ion in the acidic medium to from Diazonium ion

N -
Adding sulphamic acid to prevent the side reactions with the dye formed which may be caused by the increase of sodium nitrite, which reacts with the increase of nitrite and produce free nitrogen gas Nitrose acid Sulphamic acid Then coupling Diazonium salt with carbofuran in the alkaline medium to get the yellow Azo Dye

Fig.(9) Carbofuran Coupling with of Reagent
The reaction shows an absorption maxima at 429nm named the molar absorpitivity of 10 ppm is (1×10 4 Lmol - 1 cm -1 )which means that the method can be used for direct determination of carbofuran depending on the absorption measurements of thecoupling product of carbofuran,

Calibration Curve
A series of solution is prepared with deionized water at a known concentration of carbofuran (0, 2,4,6,8,10,12,14 ppm) in to 10mL volumetric flasks and added 0.3mL of benzidine , 0.4 mL of 1% of NaNO 2 , 0.4 mL of 1M of HCl, 0.3 mL of 0.1% sulphamic acid and then added 0.3mL of 0.1% Tween and then added 0.5 mL of 1M Na 2 CO 3 and filled to mark with distilled water to form a spiked samples.The absorbance measurements are carried out at a wavelength at 429 nm.The concentrations are obtained from the calibration curve for the spiked solutions are shown in the (Table 14) The calibration curve is drawn by using the mean absorbance as a function of concentration (ppm) as shown in Figure (10).

Stability Constant of Reaction Product [13-14]
The conditional or apparent stability constant of the 1:1 (Reagent and carbofuran) product is evaluated as described Complete founding the stability constant [K] of colored product Formed imputation of ( Regent :carbofuran) as follows : A series of solution is prepared containing three different concentrations of reagent and carbofuran the concentration (5× 10 -4 ) mol L -1 for Each reagent and carbofuran when Forming imputation under this condition easily to Hydrolysis and the Intensity absorption are very low.Another series of solution is prepared containing three different concentration of reagent and carbofuran but with abundance of the reagent (the best concentration) The complex is prepared of the intensity absorption A m and application the relationship we can calculate the value degree of decomposition follows (α): Stability constant [K] as follows ;

Conclusion:
The proposed method is simple, sensitive and free from drastic experimental conditions such as heating.It is also accurate and precise enough to be successfully adopted as an alternative

Fig. ( 2
Fig. (2): A. The Absorption Spectrum of the Carbofuran Versus Distilled Water B. The Absorption Spectrum of the Benzidine Versus Distilled Water C. The Absorption Spectrum of Carbofuran With reagent Benzidine Versus Blank

Fig. ( 6 )
Fig. (6):The Effect of Salphamic Acid Volume on Absorbance of Complex of Carbofuran with Benzidine

Fig.( 12 )
Fig.(12): Mole Ratio Plot of Carbofuran and Benzidine Complex.CL: The concentration of the reagent(benzidine) CM: The concentration of the compound (carbofuran) ; stability constant C; the concentration of the product complex and it equivalence the concentration of carbofuran.areshown in the Table19 spectrophotometric method and evaluation of carbofuran in an electrophilic compound using Diazotization and in Environmental samples.

Preparation of Standard Solutions: All
glassware used are supplied and cleaned with distilled water and dried at50˚C for 30min prior to use.Batch experiments are carried out to ensure the reproducibility of results and the average value.All reagents used are of the highest purity and most solutions are prepared in ultra pure water and Dieonized water.A standard stock solution of 250 ppm of carbofuran is prepared by dissolving 0.25g of the solid carbofuran in 1000ml of deionized water.A working standard solution of 100 ppm of carbofuran is

Table (2): Effect of Volume of Hydrochloric Acid and Time in Absorbances intensity of aAzo Dye
It is found that 0.4mL of 1M HCl acid is adequate for completing diazotization, and it give a maximum absorbance at 5min.Increasing in the volume of HCl solution causes a decrease in the absorbance of the reaction product (

(3): The Effect of Hydrochloric Acid Volume The Absorbance of Carbofuran Which Demonstrates That The Best Volume of HCl is 0.4 mL. That Dependent for Further Optimization Effect of the Type of the Alkaline
Table(

Table ( 18) The Mole Ratio Method of The Carbofuran and Benzidine Complex.
( 12 )ng the value of absorbance versus the CL /CM is shown in Figure( 12 )