2. Fundamentals and State of Knowledge
2.5.2 MAGs and DAGs synthesis
Tranditionally MAGs and DAGs are synthesized via glycerolysis of triglycerides, employing alkaline catalysts under a nitrogen gas atmosphere. Although these processes are widely used, there still remain two disadvantages: low purity and high temperature demand.
These processes have a low yield and require a temperature up to 220 - 250°C. Therefore, the products have several drawbacks with dark color and burnt taste [105]. Because MAGs have better emulsifying properties than a mixture of different acylglycerols, a molecular distillation step is required to obtain higher pure MAGs in food industry.
b) Enzymatic synsthesis
When glycerides with a high purity are needed, the alternative route is the use of lipase as a catalyst. By this way, high specificity of the catalyst and mild conditions are obtained. In general, there are several approaches for the enzymatic synthesis of MAGs and DAGs, involving the esterification of free fatty acids with glycerol, transesterification of methyl or ethyl ester with
3 Source: http://www.lipidlibrary.co.uk
glycerol, glycerolysis of a fat and oil, or selective hydrolysis of fat or oil. To improve the solubility, glycerol can be preadsorbed on the solid support. Water as a byproduct of the synthesis can be removed by the adsorption on the support or molecular sieve and evaporation under vacuum.
In many studies, the synthesis of MAGs and DAGs has been reported so far. Glycerolysis of tuna oil enriched in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) with Pseudomonas fluorescence lipase was studied by Pawongrat [106]. The optimum conditions for MAG production were reported with 10% w/v of tuna oil in MTBE. When the glycerolysis was conducted with Novozym 435, the maximum yield of MAG was up to 90% [107].
A screening process of many commercially available lipases for diacylglycerol synthesis by enzymatic glycerolysis was conducted by Kristensen [108]. The study suggested that contact between the lipases and the hydrophobic oil phase was limited because glycerol forms a layer around the hydrophilic lipase particles.
In the other method, a derivative form of glycerol is used as the raw material for MAG and and DAG synthesis. Isopropylidene glycerol was estererified with free fatty acids in the absence of an organic solvent, or transesterified with methyl esters in hexane. Oleic and eicosapentaenoic acid were successfully incorporated into glycerides from 46.9 to 96.9% [109].
Because valorisation of glycerol is needed, enzymatic esterification of glycerol and fatty acids and theirs derivatives are drawing much attention. Kwon (1995) esterified palmitic acid with glycerol in n-hexane. To increase the equilibrium conversion and the interfacial area between glycerol and fatty acid, various inert solid supports were directly introduced into the reaction system [110]. Silica gel showed a good effectiveness with 60% palmitate conversion when Mucor miehei lipase was used as a catalyst. The results also showed that temperature and molar ratio of palmitate to glycerol greatly influenced selectivity and yield of mono- and dipalmitin.
Glycerol can be preadsorbed onto silicagel to improve the solubility of the substrate.
Bellot et al. used Rhyzomucor miehei lipase for direct esterification between silica gel-adsorbed glycerol and oleic acid [111]. In pure n-hexane, at thermodynamic equilibrium, the total products included 6 molar % MAG and 34% DAG. When an equivolume mixture of n-hexane and 2- methyl-2butanol was used, 94% MAG and 2.4% DAG could be obtained. The partition of water between solvent and immobilized catalyst, and the partition of glycerol between solvent and
silica gel have been proven to be important for the productivity and selectivity of the synthesis reaction.
A mixture of free fatty acids can also be used as an alkyl source for the reaction. Lo et al.
reported DAG synthesis by Rhizomucor miehei lipase-catylsed esterification of glycerol with fatty acids from the palm oil deodorizer distillate [112]. No solvent is required for the esterification. A maximum yield of 52% (w/w) DAG was observed after 6h reaction time. When a mixture of fatty acids from corn oil deodorizer distillate was used, with Rhizomucor miehei lipase, 70% (w/w) yield could be obtained after 5h [113].
Esterification between glycerol and fatty acids is summarized in Table 2.8. Sanjib Kumar Karmee has recently reported a good literature review about lipase catalyzed synthesis of MAGs from biomass derivatives [83].
Table 2.8: Emzymatic esterification of MAG and DAG – Literature review.
Year, Cite
BLipase Subtrates Solvent Water
removal 2008
[114]
Candida antarctica glycerol caprylic acid
Free solvent Microwave irradiation 2007
[115]
Candida antarctica Glycerol
Conjugated linoleic acid
Free solvent Vacuum- driven N2 2006
[105]
Staphylococcus simulans glycerol oleic acid
Free solvent CaCO3 Celite 545 2004
[112]
[113]
Rhizomucor miehei Glycerol
Fatty acids from palm oil and corn oil deodoriser distillate
Free solvent Molecular sieve
2003 [116]
Candida antarctica Rhizomucor miehei Rhizopus delemer Pseudomonas cepacia
Glycerol Fatty acids (C8-C22)
Acetone, hexane, isooctane,
acetonitrile, cloroform
Molecular sieve
2003 [117]
Rhizomucor miehei Glycerol oleic acid
Octane, hexane
Molecular sieve 2003
[118] [88]
Candida rugosa,
Penicillium camembertii
Glycerol
Conjugated linoleic acid
Free solvent Evaporation
2001 [111]
Rhyzomucor miehei silica gel-adsorbed glycerol
oleic acid
Hexane and 2-methyl-2- butanol
Silica gel adsorbtion 1998
[119]
Rhyzomucor miehei silica gel-adsorbed glycerol
oleic acid
2-methyl-2- butanol amended n-hexane
Molecular sieve 1995
[110]
Mucor miehei
Pseudomonas fluorescens Rhizopus delemar
glycerol and palmitic acid
Hexane Silica gel
Celite Active charcoal Florisil Al2O3
1994 [120]
Pseudomonas cepacia Glycerol lauric acid
bis-(2-ethylhexyl) sulfosuccinate
No 1992
[121]
Chromobacterium viscosum, Rhizopus delemar, Rhizomucor miehei
silica gel-adsorbed glycerol
various free fatty acids, fatty acid alkyl esters, vinyl esters
Tert-methyl-buthyl ester
Silica gel adsorbtion