Saccharomyces cerevisiae Catalyzed Cyclocondensation Reaction: Synthesis of Pyrazoline

A simple, efficient, and environment friendlymethod has been developed for the synthesis of pyrazoline by the cyclocondensation of chalcones and hydrazine hydrate using very cheaper catalyst, baker’s yeast (Saccharomyces cerevisiae).The 3,5-diphenyl pyrazolines are synthesized in moderate to good yield. The method also overcomes drawbacks of the previous pyrazoline synthetic methods and provides a new efficient route to 3,5-diphenyl pyrazoline derivatives.


Introduction
The pyrazolines are important nitrogen containing heterocyclic compounds possessing various pharmacological activities and therefore they are useful material in pharmaceutical industry for drug development [1][2][3].
Triaryl pyrazoline is showing antidepressant activity and also inhibits the flavivirus infection of cell culture and was identified through high-throughput screening of compound library using luciferase west Nile virus [6].
In view of the applications of 3,5-diphenyl pyrazolines, various methods are available for their synthesis using different catalysts like acetic anhydride [10], pyridine [11], and acetic acid [12].
The above-mentioned catalysts suffer from drawbacks like need of elevated temperature, longer time, and some catalysts which are hazardous; that is why green method for the synthesis of pyrazoline is required. Till date there is not any biocatalytical method reported; therefore we have developed baker's yeast catalyzed protocol for the cyclocondensation of chalcone and hydrazine hydrate to obtain pyrazolines.
Considering the above facts, we have developed an efficient methodology for synthesis of pyrazoline from chalcone and hydrazine hydrate in organic solvent by using inexpensive biocatalyst, baker's yeast.

Materials.
All chemicals were purchased from commercial suppliers and used as they are. 1 H NMR and 13 C NMR Spectra were recorded on Bucker Advance II 400 MHz spectrometer at ambient temperatures in CDCl 3 as solvent.
Thin layer chromatography was carried out using Merk's silica plates. The dry baker's yeast is purchased from local market.

Method: General Experimental Procedure for Synthesis of Pyrazolines (3a-m).
To the mixture of chalcone (5 mmol) and hydrazine hydrate (10 mmol) in methanol (15 mL), baker's yeast (2 g) was added. The resulting reaction mixture was continuously stirred at room temperature on magnetic stirrer. The progress of the reaction was monitored by thin layer chromatography using petroleum ether : ethyl acetate (3 : 1). After completion of the reaction (32 h), reaction mixture was filtered using silica bed to remove the catalyst and washed with methanol (50 mL). The reaction mixture was concentrated under vacuum and obtained crude products. The purification was done by recrystallization in ethanol.

Result and Discussion
Herein, we report an efficient and economic synthesis of pyrazoline from chalcone and hydrazine hydrate under mild conditions catalyzed by baker's yeast. Resulting products are obtained in high yields in methanol. The chalcones are prepared by the reaction of substituted acetophenones and benzaldehydes in the presence of sodium hydroxide in aqueous ethanol [23,24]. In order to get best experimental condition, we considered the reaction of chalcone and hydrazine hydrate using baker's yeast catalyst as model reaction (Scheme 1).
From Table 1 it is concluded that the protic solvents are best for this reaction and among those methanol was found to be best solvent for this reaction because reactions required less time and high yield of product is obtained (Table 1, entry 2) as compared to other solvents. Therefore, methanol was selected as a solvent for further study. Model reaction was carried out in the absence of catalyst in methanol but there no conversion takes place even after 32 hrs (Table 2, entry 1).
Then model reaction was forwarded using different amount of catalyst, starting with 0.5 g baker's yeast but there was no formation of product in 32 hours. When 1 g of baker's yeast was used, 35% yield in 32 hours is obtained. 2 g baker's yeast gives 89% yield, and 3 g and 4 g baker's yeast get 90% yield in 32 hrs. Though increased amount of catalysts yielded 90% of product but it required more solvent (20 mL) because 3 g and 4 g of catalyst were difficult to stir in 15 mL of methanol. We analyzed the yield after isolation of product (3a); we considered that model reactions with 2 g of baker's Journal of Applied Chemistry

1(a-m) 3(a-m)
Room temperature MeOH NH 2 NH 2 ·H 2 O Scheme 2: Synthesis of pyrazoline derivatives.  yeast in methanol are the best because of higher yield in less amount of solvent and catalyst ( Table 2, entries 2-5).
A variety of chalcones containing electron donating and electron withdrawing group attached to either acetophenone ring or benzaldehyde ring or both the rings were successfully employed to prepare corresponding products. Firstly simple acetophenone with benzaldehydes, 4-chloro, 4-nitro, 4-N,N dimethyl amine, and 4-methoxy substituted benzaldehydes containing chalcone to give desired products in good to better yields, ( Table 3, entries 1-5), and then 4-hydroxyacetophenone with benzaldehyde and 4-chloro and 4-methoxy substituted benzaldehyde ring containing chalcone to give good yield of the products, ( Table 3, entries 6-8) and then the chalcones of 2-hydroxyacetophenone and 4-chloro and 4-N,N-dimethyl substituted benzaldehyde successfully and yielded the pyrazolines, (Table 3, entries 9-11) and when the chalcone prepared from 4-methoxy acetophenone and 4-chlorobenzaldehyde is allowed to react with hydrazine hydrate in the presence of baker's yeast, it gives good yield of the corresponding pyrazoline (Table 3, entries 12-13 and Scheme 2).

Conclusion
In summary, here baker's yeast is successfully employed to synthesize value added heterocycle, pyrazoline from chalcone in organic solvent, with moderate to good yield under mild conditions. The highlight of this work is the cost effectiveness and eco-friendliness.