New Preservative Glucose Methyl Ester Of Fumaric Acid Synthesis And Antibacterial Properties

In the production of food, it is crucial that proper measures are taken to ensure the safety and stability of the product during its whole shelf life. Preservatives are still one kind of important food additives. Sorbate, benzoate and esters of p-hydroxybenzoic acid are the principally employed preservatives in food industry, but the antimicrobial activity of them are limited by solubility, the species of microbe, food components, pH and so on. The preservatives used at present still can not meet the need of developing food industry. So it is a hot issue in food science to develop high-performance, broad-spectrum and low toxicity preservatives. Having studied on the relationship between structure and antimicrobial activity of preservatives, a structure-function relationship theory was developed which regardsα,β-unsaturated carbonyl as an effective group of antimicrobial activity. Di-methyl fumarate is an effective preservative containing such group, but it is banned to used in food because of its skin allergy. In order to decrease its allergy, a new preservative was designed and synthesized and its antimicrbial activity were studied. Maleic anhydride, glucose and methol was choosed as reactant. According to structure chemistry calculate, 2-and 6-OH are more easy to react than others. The following synthesis route was adopted: maleic anhydride react with methol, then isomerized with fumaric acid monomethyl monochloride(FMMC) and monomethyl fumarate(MMF) was produced. FMMC chlorided with thioyl chloride produced MMF which react with glucose and the proposed production glucosyl methyl fumarate(GMF) was synthesized. We studied the optimum technology condition as following: 1. Synthesis of MMF: maleic anhydride and methol (molar ratio 1:1) reacted under 60℃for 1 hour before 6% FMMC was added. Then the mixture maintained at 70℃for 2 hours, and the yield of MMF reached 86%. 2. Synthesis of FMMC: MMF and thioyl chloride (molar ratio 1:6) reacted at 100℃for 40 minutes, and the yield of FMMC is above 90%. 3. Synthesis of GMF: the relationship of antimicrobial activity(Y₂) and three variable, ratio of reactant (X₁), temperature (X₂) and time (X₃), was studied by RSA ( Response Surface Analysis) and the following equation was developed: Y₂=29.7+6.38X₁+2.5X₂+2.13X₃-5.71X₁²-4.46X₂²-2.21X₃²-1.5X_1X2-0.25X₁X₃ -5.5X₂X₃. From RSA contour, the optimized technology condition is: molar ratio of FMMC to glucose 3:1, temperature 83⁹1℃　and reaction duration 98¹08min. The production extracted by ether can delay the growth of mixed microbe to 32 hours. The production can be deduced as prospected ester from its IR spectrum: it has typical absorption of ester carbonyl and the ester carbonyl conjugate with contiguous olefinic bond; there are both absorption of methyl and glucosyl. HPLC analysis showed there were three main elute suspected as GMF with different esterification degree as one, two and three. The degree of esterification(DE) of GMF is directly relevant with antimicrobial activity. GMF molecule with low DE has more free –OH and is easy to be dissolved in water but not easy to enter cell membrane to react with sub-structure in cell, which decrease its antimicrobial ability. However, too high DE is lipophilic and decreases solubility of GMF, which make GMF difficult to contact with microbe in media and low its antimicrobial activity. The results showed that GMF with degree of esterification has the highest antimicrobial ability than others. Antimicrobial activity of GMF was systematically studied and the results showed that GMF can delay the growth of bacteria, yeast and mold to more long duration than sorbate. Skin tests showed that GMF donot stimulate skin as dimethyl fumarate. So it may be used in food as a kind of effective, broad-spectrum and low toxicity food preservative. QSAR (Quantitative Structure-Activity Relationship) was introduced to research 17 preservatives which contain α,β-unsaturated carbonyl group. QSAR was analyzed with multiple stepwise linear regression. Molecular connectivity of preservativeswere adopted to describe molecular structure and equation of MIC and t0.5 were calculated as following respectively: lg1/MIC=2.693+0.284⁴Хν+0.523⁴Ð¥pcν; lgt_0.5=0.668+1.076³Ð¥cν+0.489⁴Ð¥pcν. The results showed that the principal structure parameters affecting MIC and t0.5 are 4^Хν,4^Ð¥_pc^ν,3^Ð¥_c^νand 4^Ð¥_pc^νrespectively. Both equations obtained can be used to predict the antimicrobial activity.