Prepararion Of Iminodiacetic Acid By Immobilized Cells

Iminodiacetic acid(IDA)is an important intermediate for the production of glyphosate.And glyphosate is the most commonly used herbicides in the word,with 30%consumption of the global herbicides.Till we see its active demand with the rising of annual sales.Now,the mainly manufacturing process used for glyphosate utilise IDA as intermidates.Almost all the production of IDA,however,employs chemical methods,that bring out many byproducts and toxic,causing problems of cost and environment.While,enzymatic hydrolysis of nitrilase meets the "green chemistry" for the mild condition,low cost and ecofriendly nature.Lately,there have been some successful applications of nitrilase in hydrolyzing nitriles,but few researches were found in iminodiaectonitrile.The line of this study was the immobilization of the nitrilease-producing strain Alcaligenes faecalis ZJB09133,stored in our laboratory,and build of bioreactor.The main object was to research the biotransformation of iminodiacetonitrile to iminodiacetic acid by immobilized cells.The all research contents and results are as flows.The first,established the analytical method of iminodiacetonitrile and iminodiacetic acid.The optimal conditions was found:chromatographic colum,Hypersil SAX 5 ?m 4.6mm ×200 mm;mobile phase,20mM(NH4)2 HPO4,pH = 4;room temperature;detect wavelength,210 nm;flow rate,1.0 mL/min;sample size,20uL.Secondly,Screened out the optimal carrier,and optimized the condition of preparation of immobilized cells and its biocatalysis.Moreover,the reusability and stability of the immobilized cells were inspected.Polyvinyl alcohol-sodium alginate(PVA-SA)was chosen as the best carrier,and its optimal immobilization method was found:sodium alginate 2.0%;Polyvinyl alcohol 8.0%;embedding quantity 8.0(g/ml);4.0%calcium chloride with saturated boric acid solution(pH was adjusted to 7.0 by calcium hydroxide);sodifying 12h.The best catalyzing condition for immobilized cells obtained by the method above was as follows:iminodiacetonitrile 1.0%;immobilized cells(free cells embeded)0.2g;Tris-HCl buffer,pH 8.0;temperature 35 ?,150rpm.For 8 hour's conversion,15%yield was achieved.The immobilized cells also showed better reusability than free cells,as immobilized cells can be reused 10 and with 37%initial activity,while free cells can only be reused twice with 20%of its initial activity left.And both the free and immobilized cells had satisfied stability in a month.The last,built the bioreactor and optimized the operating process,for the feasibility research of producing IDA by immobilized cells in large scale.The experimental results showed that,the yield of IDA maintained around 20%in a week,with the catalization of immobilized cells,and PVA-SA as carrier,in packed-bed reactor,when 0.8%substrate was fed continuously.