Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate) as efficient Pepsin Inhibitor and a synthesis process thereof

Abstract

The present invention discloses a novel Pepsin inhibitor with excellent inhibition activity against the Pepsin enzyme that logically diminishes the activation of Pepsinogen into Pepsin. The present invention also discloses a process for preparing the novel Pepsin inhibitor, wherein process methodology used is highly economic as it uses no metal catalysts, acid or base and eco-friendly by using water as the green solvent. The process comprises preparing a carbohydrazide solution by mixing at least one hydrazide species in double distilled water. Then adding an aldehyde solution in the carbohydrazide solution at room temperature with continuous stirring to prepare the precipitates of the novel Pepsin inhibitor compound. Filtering and washing the precipitates with cold water at least one or two times to obtain the washed precipitates. Drying the washed precipitate under vacuum to obtain the novel Pepsin inhibitor compound.

Description

FIELD OF THE INVENTION:The present invention relates to the field of enzyme inhibitors. Specifically, the present invention relates to a novel class of compounds as Pepsin inhibitor with excellent inhibition activity in order to treat Pepsin-induced disorders. More specifically, the present invention discloses an ecofriendly and green process for synthesis of a novel compound as Pepsin inhibitor.BACKGROUND OF THE INVENTION:Pepsin, an aspartic protease, is one of the digestive enzymes found in stomach that breaks down proteins into smaller peptides and amino acids. Pepsin is synthesized by the chief cells of the stomach lining in an inactive form referred to as Pepsinogen. Upon encountering hydrochloric acid within the stomach, Pepsinogen is activated and transformed into active Pepsin. Pepsin is involved in the breakdown of proteins into peptides and becomes higher in activity at the acidity of gastric juice with pH 1.5-2.5. The excessive activity of Pepsin contributes to various complications such as damage of stomach lining, ulcers, gastritis, mucosal erosion and acid reflux. Pepsin, the most vital digestion catalyst, is associated with many diseases viz tumor progression, inflammation, hypothyroidism, gastroesophageal reflux and most evident for laryngopharyngeal reflux.There are several reported Pepsin inhibitors that serve as treatment therapies for the conditions worsened from chronic inflammation, recurrent respiratory papilloma, vocal fold polyp and chronic reflux. However, treatment therapies to prevent damage occurring due to overexpression of Pepsin activity have been developed. But there is always a surge for better inhibitors. In this context, there is a constant need to explore novel chemical entity with potential medicinal effects. Different compounds related to Pepsin inhibition are presented in table 1.Table 1However, the Pepsin inhibition efficiency of the known inhibitors is still limited. To meet the challenges posed by increased Pepsin activity, it is necessary to develop a novel class of compounds as Pepsin inhibitor, wherein the said novel class of compounds can pave the way for better health and vitality. Further, there is a need to develop advanced Pepsin inhibitors, to effectively address Pepsin-induced disorders. Accordingly, there is a need for novel class of compounds as Pepsin inhibitors having enhanced inhibition capabilities, and wherein such Pepsin inhibitors also significantly reduce side effects, leading to better patient outcomes.Further, there is a need for advanced Pepsin inhibitors which can be easily prepared with an ecofriendly and more economic synthetic approach.OBJECTIVES OF THE PRESENT INVENTION:The primary objective of the present invention is to provide a novel class of compounds as Pepsin inhibitor.Another objective of the present invention is to provide the Pepsin inhibitor with excellent inhibition activity in order to treat Pepsin-induced disorders.Another objective of the present invention is the use of invented molecule as Pepsin inhibitor, with small molecular weight and higher inhibition capacity than the standard.Another objective of the present invention is the utilization of ecofriendly and more economic synthetic approach for the preparation of the Pepsin inhibitor. SUMMARY OF THE INVENTIONThe present invention discloses a bis-hydrazide-hydrazone compound of formula 1 for pepsin inhibition,Formula 1wherein,R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.In an aspect, the present invention discloses a bis-hydrazide-hydrazone compound of formula 1 for pepsin inhibition, wherein R is selected from CH2-C6H5, and the compound of formula 1 is Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate).The present invention also discloses a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, Formula 1wherein R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof,wherein the process comprises:preparing a carbohydrazide solution by mixing at least one hydrazide species in double distilled water;adding an aldehyde solution in the carbohydrazide solution at room temperature with continuous stirring to prepare the precipitates of compound of formula 1; filtering and washing the precipitates with cold water for one or two times to obtain the washed precipitates; and drying the washed precipitate under vacuum to obtain the compound of formula 1.In an aspect, the present invention also discloses a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the process has a reaction mechanism of,wherein, R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.BRIEF DESCRIPTION OF THE DRAWING:The detailed description below will be better understood when read in conjunction with the appended drawings. For the purpose of assisting in the explanation of the invention, there are shown in the drawings embodiments which are presently preferred and considered illustrative.It should be understood, however, that the invention is not limited to the precise arrangements and instrumentalities shown therein.The invention will now be described, by way of example, with reference to the accompanying drawings, in which:Figure 1: represents the 1H NMR spectrum of Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate) with structural formula F1.Figure 2: represents the IR spectrum of Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate) with structural formula F1.DESCRIPTION OF THE INVENTION:To meet the challenges posed by increased Pepsin activity, there is a need for innovative Pepsin inhibitors that can pave the way for better health and vitality. Therefore, the present disclosure provides a compound for effective Pepsin enzyme inhibition. It is identified that hydrazone-based inhibitors are useful for their significant efficiency in Pepsin inhibition. Hydrazides, standing at the forefront of pharmaceutical research, are recognized for their therapeutic potential in Pepsin inhibition. Accordingly, the present invention provides an excellent Pepsin inhibitor composed of hydrazide and hydrazone units. Further, water is used as a green solvent for preparing the Pepsin inhibitor as disclosed in the present invention, accordingly the process for preparing the disclosed Pepsin inhibitor is more eco-friendly and supports economic efficiency.The profile of the compound as disclosed in the present invention provides effective Pepsin inhibition and supports its potential utility in specific biochemical and biological function. Specifically, the presently disclosed Pepsin inhibitor provides significant potency in treatment and management of enzymatic dysregulation associated with inflammatory, digestive and metabolic disorders.Accordingly, the present invention is related to solving the problems resulting from the hyperactivity of Pepsin. The development of a new entity comprising bis-hydrazide-hydrazone exhibits remarkable inhibition efficiency against Pepsin enzyme. The novel compound was synthesized in purely aqueous medium in massive yield without the use of any metal or catalyst. Specifically, the present invention discloses a bis-hydrazide-hydrazone compound of formula 1 for pepsin inhibition,Formula 1wherein,R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.In an embodiment, the present invention provides a Pepsin inhibitor comprising of hydrazide and hydrazone units; wherein the compound is represented by the molecular formula C4H4N4O4R2 and with structural formula 1; wherein R is selected as -CH2-C6H5.Formula 1According to structural Formula 1, the Pepsin inhibitor is Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate). In an aspect, the disclosed Pepsin inhibitor according to structural Formula 1 has a low molecular weight of 354.13.The Pepsin inhibitor shows percent inhibition of 45.78 at 10-8 M concentration. The inhibition activity against the Pepsin enzyme was recorded in reference to Curcumin, as represented below with structural Formula 2.Formula 2In the present invention, Curcumin is chosen as standard reference to record comparative inhibition efficiency, represented by the molecular formula C21H20O6. According to structural Formula 2, standard is named as (1E,6E)-1,7-bis(4-hydroxy-3-methoxyphenyl)hepta-1,6-diene-3,5-dione. In an embodiment, the present invention provides the importance of small molecular weight compound as an effective Pepsin inhibitor. The compound penetrates easily into the active site of Pepsin, reacts quickly with catalytic residues and exhibits high specificity. In an embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, Formula 1wherein R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof,wherein, the process comprises:preparing a carbohydrazide solution by mixing at least one hydrazide species in double distilled water; andadding an aldehyde solution in the carbohydrazide solution at room temperature with continuous stirring to prepare the compound of formula 1.In another preferred embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, Formula 1wherein R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof,wherein the process comprises:preparing a carbohydrazide solution by mixing at least one hydrazide species in double distilled water;adding an aldehyde solution in the carbohydrazide solution at room temperature with continuous stirring to prepare the precipitates of compound of formula 1; filtering and washing the precipitates with cold water for one or two times to obtain the washed precipitates; and drying the washed precipitate under vacuum to obtain the compound of formula 1.In an embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the carbohydrazide solution is prepared by mixing 25 mmol of carbohydrazide in 10 ml of double distilled water at room temperature. In a preferred embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the carbohydrazide is selected from Carboaryloxyhydrazides, carboalkyloxyhydrazides and combination thereof.In an embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the aldehyde solution is prepared by adding 10 mmol of an aldehyde compound in 20% of water. In a preferred embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the aldehyde compound is a dialdehyde compound selected from Glyoxal.In an embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein the compound of formula 1 is a white powdered solid with 85% yield and the compound is stable at room temperature.In another preferred embodiment, the present invention provides a process for preparation of a bis-hydrazide-hydrazone compound of formula 1, wherein, the process has a reaction mechanism of,wherein, R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.The scope of the present invention is not limited to providing a better Pepsin inhibitor but also a compatible method of preparation. EXPERIMENTS:Method of Preparation: The active compound of structural formula 1 was prepared by a condensation method. The present invention involves the process for preparation of the Pepsin inhibitor as defined below, wherein the process comprising: adding 25 mmol of carbohydrazide in 10 ml of double distilled water to obtain a uniform carbohydrazide solution at room temp. Wherein; the aldehyde solution comprising 10 mmol of an appropriate aldehyde in 20% of water is added; stirring the mixture to obtain precipitates. The reaction progress monitored by TLC confirms the completion of reaction followed by the step of filtration; washing the precipitates with cold water for one or two times, to obtain the washed precipitates; and drying the washed precipitate under vacuum to obtain the Pepsin inhibitor. Characterization of Pepsin Inhibitor: The Pepsin inhibitor of Formula 1 has been characterized using 1H NMR and IR spectroscopy. The respective spectra are given in the accompanying figures i.e. Figure 1 and Figure 2.Compound of Formula 1:Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate); White powdered solid; 85% Yield; Stable at room temperature.1H NMR (500 MHz, DMSO-d6) δ = 11.44 (s, 2H), δ = 7.68 (s, 2H), δ = 7.40-7.33 (m, 10H), δ = 5.16 (s, 4H).ENZYME INHIBITORY ACTION (IN VITRO):As per the present invention, the Pepsin inhibition efficacy can be measured by any method known in the art. As an exemplary embodiment, the Pepsin inhibition efficacy of the presently provided Pepsin Inhibitor of Formula 1 was measured and compared with that of the standard Pepsin Inhibitor of Formula 2. The present invention evaluates the Pepsin inhibition activity using following methodology; 0.1 M phosphate buffer (Sodium dihydrogen orthophosphate dihydrate and di-Sodium hydrogen phosphate anhydrous) of pH 2.0 was used in this study. 1.920 ml of the buffer was utilized to incubate the 50 μl of Pepsin enzyme at 37°C for about 10 min. The addition of Bovine Serum Albumin (5mg / mL in water) prepared in DMSO, was added to the former reaction mixture and again incubated at 37°C for half an hour. After one hour, 3 mL of trichloroacetic acid was added. Curcumin was taken as control. Measurement of absorbance using a UV spectrophotometer was managed at 280 nm. The percentage of Pepsin inhibition was determined using the equation outlined below:% Inhibition of Pepsin = (Abc - Abs) / Abc x 100Where,Abc = Absorbance of controlAbs = Absorbance of sampleSubsequently, the results are compared with respect to the reference under similar conditions.The percent inhibition serves as a critical parameter to indicate the potency of a compound. The present disclosed compound serves a better Pepsin inhibitor with higher inhibition as that of reference (Table 2). The present invention suggests that very less amount of compound is needed to achieve the desired response of Pepsin inhibition.Table 2: Pepsin inhibition efficiencyIn vitro Pepsin inhibitory assay confirms the superiority of the inhibition efficiency of compound with structural formula 1, namely Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate); followed by the inhibition capacity of standard with formula 2, namely (1E,6E)-1,7-bis(4-hydroxy-3-methoxyphenyl)hepta-1,6-diene-3,5-dione.The in vitro Pepsin inhibitory assay experiments have also demonstrated that the compound with structural formula 1, possessing low molecular weight achieves more inhibition of Pepsin enzyme at 10-8 M concentration (Table 2) than compound with structural formula 2. The compound with structural formula 1 displayed percent inhibition of 45.78%, which is higher than the reference compound with formula 2 having 36.20% inhibition. The present disclosure serves an excellent Pepsin inhibitor with more pepsin inhibition as that of reference (Table 2). The compound of formula 1 is slightly better than the compound with formula 2.Accordingly, the present invention supports smaller molecules with higher medicinal efficacy by imparting excellent Pepsin inhibition capacity and diminishing Pepsin induced disorders. The advantage of the present invention is the utilization of ecofriendly and more economic synthetic approach for the preparation of Pepsin inhibitor. Accordingly, the present invention provides a green method of preparation of the said Pepsin inhibitor which not only uses water as green solvent but also results into higher yield.

Claims

1. A bis-hydrazide-hydrazone compound of formula 1 for pepsin inhibition, Formula 1 wherein, R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.

2. The compound as claimed in claim 1, wherein R is selected from CH2-C6H5, and the compound is Dibenzyl 2,2'-(ethane-1,2-diylidene)(2E,2'E)-bis(hydrazine-1-carboxylate).

3. The compound as claimed in claims 1 to 2, wherein the compound has a low molecular weight 354.

13.

4. The compound as claimed in claims 1 to 3, wherein the compound has percent inhibition of 45.78 at 10-8 M concentration.

5. A process for preparation of a bis-hydrazide-hydrazone compound of formula 1, Formula 1 wherein R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof, wherein the process comprises: preparing a carbohydrazide solution by mixing at least one hydrazide species in double distilled water; adding an aldehyde solution in the carbohydrazide solution at room temperature with continuous stirring to prepare the precipitates of compound of formula 1; filtering and washing the precipitates with cold water for one or two times to obtain the washed precipitates; and drying the washed precipitate under vacuum to obtain the compound of formula 1.

6. The process as claimed in claim 5, wherein the carbohydrazide solution is prepared by mixing 25 mmol of carbohydrazide in 10 ml of double distilled water at room temperature, wherein the carbohydrazide is selected from carboaryloxyhydrazides, carboalkyloxyhydrazides and combination thereof.

7. The process as claimed in claim 5, wherein the aldehyde solution is prepared by adding 10 mmol of an aldehyde compound in 20% of water.

8. The process as claimed in claims 5 to 7, wherein the aldehyde compound is a dialdehyde compound selected from Glyoxal.

9. The process as claimed in claim 5, wherein the compound of formula 1 is a white powdered solid with 85% yield and the compound is stable at room temperature.

10. The process as claimed in claims 5 to 9, wherein the process has a reaction mechanism of, wherein, R is selected from C1 to C2 alkyl group, an aryl group, and combination thereof.