From time to time being one of the speedily developing worldwide issues is fungicide remains which are present in essential cereals items cause’s health issues on human beings¹. Estimation of fungicides to plants and animals hazard can be determined over limits present in the surroundings². It has been reported that causes of pathogenic moulds in radish, carrot, bottle gourd so that cultivator used fungicides practically in frequent for protecting of their crops³. It is appropriate to develop raise awareness to user for certainty that to known how much quantity of fungicides present in their edible product⁴. Dithiocarbamates having thiocarbonyl group such as Ziram which are frequently used fungicides against wide scope of domestic, vegetable⁵. It was serious that fungicides act as neurotoxin and attack to nervous system of several animals and human beings⁶. The varieties of procedure have been used for the persistence of fungicide⁷. These including HPLC-MS, TLC, GC-MS, AAS, NMR, FTIR, IR, Polarography, Spectrophotometers, and several methods have also been reported for the persistence of fungicide⁸. The aim of proposed work is to develop simple, rapid and precise methods for analysing for the persistence of globally used Ziram fungicide at µg levels⁹. The developed method is based on acidic hydrolysis of Ziram fungicide to carbon disulphide gas, further react with ethanol sodium hydroxide to form xanthate, xanthate further react with potassium iodated to librated free iodine and this librated iodine selective oxidize Rhodamine-B dye pink colour to rhodamine-B colourless¹⁰. The proposed method has been satisfactory applied to the persistence of Ziram fungicide in several agricultural sampling¹¹.

Experimental Section   

Apparatus:-A Systronics UV-Vis spectrophotometer model no-104 with 2cm cubate and _PH meter model no-335 were used for analytical parameters measurements.

Reagents:-reagents and distilled water was used throughout the experiment.

Solution of Ziram:-2µg Ziram was dissolved in 100ml of acetonitrile in volumetric flask.

Sodium hydroxide:-5g sodium hydroxide dissolved in 100ml ethanol.

Potassium iodated (Merck):-0.1M of 0.713g potassium iodate dissolved in 100 ml water.

N-chlorosuccinimide solution (Aldrich):-2.5g N-chlorosuccinimide dissolved in 100ml water in 250 ml volumetric flask volume was made up to the mark with water.

 Acetate buffer solution prepared:-13.6g (1M) sodium acetate trihydrate dissolved in 80ml of water, solution _PH was adjusted 3-6.8 with acetic acid.

Sodium salt of EDTA:-5g EDTA dissolved in100ml water, 3g Meta phosphoric acid dissolved in 100ml water. 

Procedure:-Stock solution of Ziram prepared in acetonitrile was introduced in the digestion flask¹². Then 5M H₂SO₄ was added drop wise to the above solution for hydrolysis during which CS₂ gas was librated, sodium hydroxide dissolved in 5ml of ethanol was taken as an absorbing  for carbon disulphide gas. Experiment was completed for 4 min, the yellow colour  appeared and diluted with water then added 2.5ml of Ziram and 2ml of acetate buffer solution then 2ml of potassium iodated, 1ml of N-chlorosuccinimide were added in above solution then shaken for few min thereafter 2ml of rhodamine-B dye was added  pink colour of rhodamine-B dye is disappeared. The reaction mixture was neutralized slightly alkaline with 3% of metaphosphoric acid with sodium salt of EDTA solution, the absorbance was measured at 545nm (Fig.-2). 

Result and discussion

Absorption Spectra of Ziram:-Ziram is decomposed by acidic hydrolysis to give carbon disulphide which trapped in methyl potassium hydroxide to give potassium methyl xanthate, which is then titrated iodometrically¹³. The librated iodine selectively reduced rhodamine-B dye pink colour complex to rhodamine-B dye colourless complex (Scheme-1); rhodamine-B showed maximum absorbance(λ_max) at 545nm, Value of maximum absorbance (λ_max) is decreases due to blue shift and  intensity of absorbance (E_max) is also decreases it is called hypo chromic shift because of colour of rhodamine-B dye is discharged shown in (Fig.-1).

Colour reaction:-The following steps as described in

 (1) Liberation of carbon disulphide from Ziram fungicide by acidic hydrolysis at 25-35 ºC temperature. (2) Formation of xanthate with the treatment of CS₂ with alcoholic sodium hydroxide. (3) Iodine is librated as a result of reaction between potassium iodated act as oxidizing agent and the xanthate in the presence of N-chlorosuccinimide¹⁴.  Potassium iodated has +5 oxidation states it changes into zero oxidation state in free iodine in step first. But in step second free iodine has zero oxidation state it changes into +1 oxidation state here iodine is selectively oxidized. (4) Formation of reduced rhodamine-B dye quinine structure through selective reduction reaction of rhodamine-B dye to formed benzenoid structure by librated iodine (Scheme-1).                                                      

Calibration curve of Ziram fungicide:-This graph is plotted between concentration of Ziram (µg) and absorbance. A straight line is obtained. It follows Beer-Lambert’s law at 10-2µg shown in (Fig.-2).

Effect of concentration of rhodamine-B dye:-This graph is plotted between volume of rhodamine-B dye and absorbance.1.5 ml (0.1g/1000ml) of rhodamine-B dye is required for decolourizing itself (Fig.-3).

Effect of sodium hydroxide:-This graph is plotted between volume of sodium hydroxide and absorbance. Volume of reagent concentration is required for decolourising rhodamine-B dye is 1ml (5g/100ml).  It is responsible for maximum hydrolysis shown in (Fig.-4).

Effect of concentration of potassium Iodated:-This graph is plotted between volume of potassium iodated on x-axis and absorbance on y-axis.1.5 ml (0.713g/100ml) of potassium iodated is sufficient for decolourising rhodamine-B shown in (Fig.-5).

Effect of _PH:-This graph is plotted between pH on x-axis and absorbance on y-axis.PH 3-6.8 is sufficient for decolourising rhodamine-B dye and acetate buffer solution maintaining the above pH of the system shown in (Fig.-6).

Effect of time and temperature:-This graph is plotted between temperature and absorbance.It has observed that the hydrolysis of Ziram fungicide was studied at slightly acidic condition. The maximum hydrolysis of the decoloured reaction was observed.  The 25-35ºC temperature is sufficient for decolourising rhodamine-B dye. It was also found that a required time period is 5min for the complete decolouration of solution after dilution to 10ml water and formed product was stable for several hours shown in(Fig.-7).

Effect of Ziram Concentration:-This graph is plotted between volume of Ziram and absorbance, the 2.4ml volume of Ziram is enough for decolourising rhodamine-B dye shown in (Fig.-8).

Differentiation with reported methods:-The differentiation between reported method and proposed method, reported method are less sensitive, highly costly, it has extra steps hydrogen sulphide generation but proposed method is highly sensitive, low cost and can be applied to real sampling¹⁵(Table-1).

Effect of foreign species:-The fungicide were evaluated from several agricultural sampling and presence other  fungicides, pesticides and commons ions does not interfere in the procedure but hydrogen sulphide and some ions like  Cu²⁺, Hg²⁺, Fe³⁺ interfere in the persistence of Ziram, interference of other ions is inhibited by absorption in lead acetate¹⁶ (Table -2).

Analytical parameters characteristics of proposed method (Table-3)

Application of Ziram fungicide in various samples:-To check the credibility of proposed method and persistence of Ziram in10ml agricultural running water were taken from the region where Ziram has been sprayed such as Pihrid, Kalami¹⁷.  These agricultural sampling were found to be freed Ziram, known amount of ziram is added in agricultural running water sample taken in 50ml beaker 1.5ml of concentrated sulphuric acid mixed and _PH maintained at 3-6.8; these sampling were analyzed through proposed method¹⁸.Several vegetables, potato, lettuce, green leafy vegetables, wheat were collected from agricultural region where Ziram has been sprayed. 300g of tomato, potato, cabbage, taken and blended in a mixture;  known amount of ziram is added and kept some days;  the sample was digested with 100ml of sulphuric acid. The mixture was filtered and analysed by proposed method¹⁹ (Table-4).

Conclusion:-The proposed method is on the basis of use of reagent rhodamine-B dye for the persistence of ziram. Proposed method offers simple sensitive, selective and affordable. This developed method is much better than other sophisticated equip mental methods. Impact on community to spread awareness, amongst community, that have this fungicide of harmful to human health and environment. To give suggestion to the local farmers for tempted used for this fungicide. In present scenario development of method for analysis of any harmful chemical is important technique, through many sophisticated instrumental methods are available but this classical method have their own importance forever. Significance of method was many analytical parameters have been studied, results as approved by statics analysis accuracy and precision are determined. Results of application reflect the reliability to the method. In the development method no any harmful, chemicals were used; the proposed method is reliable, precise, affordable and easily available in small laboratory then other sophisticated instrumental reported method. 

Acknowledgement

       I express my gratitude to Principal Dr. B. D. Jangade Govt.Vedram College Malkharoda, District-Sakti (C.G.), and India. I am also thankful to University Grant Commission, New Delhi for providing laboratory facilities and financial assistance to carry out this experiment.

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