Methods for purification of sophorolipids
The subject invention provides materials and methods for producing and purifying sophorolipids (SLP). More specifically, the subject invention provides materials and methods for the purification of both hydrophobic and hydrophilic SLP molecules to a purity of, for example, at least 80% by weight, preferably at least 95% by weight, without using solvents or centrifugation. Advantageously, the subject invention is suitable for industrial scale production of purified SLP for use in, for example, cleaning products and detergents, and uses safe and environmentally-friendly materials and processes.
1. A method for producing a purified sophorolipid (SLP) composition, the method comprising:
cultivating a sophorolipid-producing yeast in a submerged fermentation reactor to produce a yeast culture, said yeast culture comprising liquid fermentation medium, yeast cells and a mixture of hydrophobic SLP and hydrophilic SLP;
separating the hydrophobic and hydrophilic SLP by allowing the yeast culture to sit undisturbed at the end of the cultivation cycle in the fermentation reactor or in a first collection container for 12 to 50 hours, wherein a layer comprising hydrophobic SLP settles to the bottom of the yeast culture, and collecting the settled hydrophobic SLP layer into a second collection container, leaving behind a supernatant comprising dissolved hydrophilic SLP; and
purifying the hydrophobic SLP and/or the hydrophilic SLP;
wherein the hydrophobic SLP layer further comprises impurities, selected from yeast cells, glucose, fatty acids and other residual materials from fermentation, wherein purifying the hydrophobic SLP comprises water-washing the hydrophobic SLP layer according to the following steps:
a) mixing the hydrophobic SLP layer with deionized water continuously for about 30 to 300 minutes at a temperature of about 50 to 80° C.;
b) allowing the hydrophobic-water mixture to rest for about 8 to 24 hours, wherein the mixture stratifies into a bottom layer comprising hydrophobic SLP and water, a middle layer comprising water and impurities, and a top foamy layer comprising additional impurities; and
c) collecting the bottom layer into a third collection container to produce a water-washed SLP composition; and
wherein the purified hydrophobic SLP composition comprises lactonic SLP, di-acetylated linear SLP and/or mono-acetylated linear SLP.
2. The method of claim 1 , wherein the yeast is Starmerella bombicola.
3. The method of claim 1 , wherein the middle layer of the stratified mixture of the hydrophobic SLP-containing layer and water comprises water and impurities, said impurities comprising residual yeast cells, glucose and/or fatty acids.
4. The method of claim 1 , wherein the top layer of the stratified mixture of the hydrophobic SLP-containing layer and water comprises foam and impurities, said impurities comprising residual yeast cells, glucose and/or triglycerides from fermentation.
5. The method of claim 1 , wherein the percentage of water in the collected bottom layer is adjusted so that it is less than 50%.
6. The method of claim 1 , wherein the water-washed SLP composition comprises one or more fatty acid and/or oil impurities, and wherein the method further comprises removing the fatty acid and/or oil impurities.
7. The method of claim 6 , wherein the fatty acid and/or oil impurity is oleic acid.
8. The method of claim 1 , wherein the purified SLP composition has a hydrophile-lipophile balance (HLB) value of about 1 to 8.
9. The method of claim 1 , wherein the supernatant comprises hydrophilic SLP and glucose impurities dissolved therein, and wherein the method further comprises centrifuging the supernatant to remove cellular material and removing glucose impurities in the supernatant using a yeast digestion method and/or an enzymatic digestion method.
10. A method for producing a purified sophorolipid (SLP) composition, the method comprising:
cultivating a sophorolipid-producing yeast in a submerged fermentation reactor to produce a yeast culture, said yeast culture comprising liquid fermentation medium, yeast cells and a mixture of hydrophobic SLP and hydrophilic SLP;
separating the hydrophobic and hydrophilic SLP by allowing the yeast culture to sit undisturbed at the end of the cultivation cycle in the fermentation reactor or in a first collection container for 12 to 50 hours, wherein a layer comprising hydrophobic SLP settles to the bottom of the yeast culture, and collecting the settled hydrophobic SLP layer into a second collection container, leaving behind a supernatant comprising dissolved hydrophilic SLP;
purifying the hydrophobic SLP and/or the hydrophilic SLP;
wherein the supernatant comprises hydrophilic SLP and glucose impurities dissolved therein, and wherein the method further comprises centrifuging the supernatant to remove cellular material and removing glucose impurities in the supernatant using a yeast digestion method and/or an enzymatic digestion method.
11. The method of claim 10 , wherein the enzymatic digestion method comprises mixing glucose oxidase into the centrifuged supernatant to catalyze the conversion of glucose into gluconic acid and hydrogen peroxide; and, optionally, evaporating the hydrogen peroxide.
12. The method of claim 10 , wherein the purified hydrophilic SLP has an HLB of 10 or greater.