Synthesis And Application Of Indoxyl Chromogenic Substrate

In the application of microbiological detection in food and clinic industries,the chromogenic medium can not only ensure the accuracy of the test results,but also reduce the detection time greatly.In this paper,2,4-dichlorobenzoic acid and methyl 2-aminobenzoate were used as the starting materials,and the target compounds–a series of 3-indole glucosides were synthesized and characterized.The synthetic conditions of the target compounds were further explored and optimized.The application as chromogenic agent in the microbiological detection was also investigated.1)Through the study on the synthetic conditions of the 3-indole esters including the reaction time,reaction temperature,the rate of material,and the amount of catalyst and the conclusion were as follows:（1）Using I₂ and 30%H₂O₂,the suitable reaction condition for 2-（（carboxymethyl）amino）-5-iodobenzoic acid is 1.2 times of the equivalent of 2-amino-5-iodobenzoic acid,the reaction temperature 75℃,the reaction time 12 h,and the pH 8.（2）The suitable conditions for the synthesis of 1-acetyl-6-chloro-1H-indol-3-yl acetate were as follows:1 grams of 2-（（carboxymethyl）amino）-4-chlorobenzoic acid and 16 milliliters of acetic anhydride（2-（（carboxymethyl）amino）-4-chlorobenzoic acid:Ac₂O=1:38.8）,sodium acetate is 4times the equivalent of 2-（（carboxymethyl）amino）-4-chlorobenzoic acid.The reaction temperature of the synthesis of 1-acetyl-6-chloro-1H-indol-3-yl acetate is 120℃and the reaction time 2 h.（3）The suitable conditions for the synthesis of 6-chloro-1H-indol-3-yl octanoate:sodium hydroxide is 12 times equivalent of 1-acetyl-6-chloro-1H-indol-3-yl acetate,the temperature of deacetylation 120℃,reaction time 2 h.Octanoyl chloride is 5 times equivalent of 6-chloro-1H-indol-3-yl octanoate,the temperature of reaction of esterification0℃,and reaction time 2 h.The target product was characterized by NMR.The synthesis of 3-indole-beta-D-glucoside was investigated.There are some conclusions:（1）The suitable conditions for the synthesis of（1-acetyl-6-chloroindole-3-group）-2,3,4,6-four acetyl-beta-D-glucoside are that 2,3,4,6-Tetra-O-acetyl-alpha-D-glucopyranosyl bromide is twice the equivalent of 1-acetyl-6-chloroindolin-3-one,and the potassium hydroxide is twice the equivalent of it.The reaction time 4 h,and the reaction temperature is room temperature,（2）The suitable conditions for the synthesis of 6-chloride-3-indole-beta-D-glucoside are suitable.Sodium methanol was selected to hydrolyze it,and the amount of sodium methoxide was 1times the equivalent of it,the reaction time 2 h,and the reaction temperature is room temperature.The target products are characterized by NMR.2)Using the above compounds to explore the application of salmonella detection,the following conclusions are drawn:The detection effect is good and the growth of salmonella is good too.The chromogenic medium solution consisted of salmonella chromogenic substrate0.5 g/L,X-Gal 0.6 g/L,they are dissolved by Dimethyl sulfoxide and N,N-Dimethylformamide,Sodium deoxycholate 1 g/L,cefsulodine sodium 0.004 g/L.Salmonella chromogenic medium can determine Salmonella according to the color of colony,and the color of colonies is smart and easy to distinguish.At the same time,it has good specificity,which can exclude Pseudomonas aeruginosa,which is false-positive,and has a good inhibitory effect on most gram positive bacteria.It also can distinguish Salmonella and Escherichia coli.The chromogenic effect of 6-chloro-3-indole-beta-d-glucoside was excellent.