1. Proposal for Soybean Association to Reduce the Sterol Glucoside Content in Soy-based Biodiesel By Haiying Tang

2. Structures Plant Sterols exist as free sterols, steryl esters, steryl glycosides, and acylated steryl glycosides.

3. Toivo et al (Determination of sterols in foods: Recovery of free, esterified, and glycosidic sterols. Journal of Food Composition and Analysis 2001;14(6):631- 643) – Corn meal and onion contain an appreciable amount of steryl glycosides – Rapeseed oil did not contain steryl glycosides. Verleyen et al ( Influence of the vegetable oil refining process on free and esterified sterols. JAOCA, Vol.79, no.10, 2002) –Water degumming did not affect the sterol content or its composition. –A significant reduction in the free sterols during neutralization; a gradual reduction in the total sterol content as a function of temperature (220-260 0 C) during deodorization –A considerable increase in sterol ester fraction during physical refining; a slight increasing in the free sterols during acid degumming and bleaching Results:

4. Continued Kochhar ( Influence of Processing on Sterols of Edible Vegetable-Oils. Progress in Lipid Research 1983;22(3):161-188.) –Sterol occur in free and esterified form in refined vegetable oil process. –Conventional refining has minor effects on the sterols; however, high temperature physical (steam) refining of oil does affect the sterol composition. –Reduction in the levels of sterols during degumming, neutralization, bleaching, hydrogenation, deodorization,and steam refining. –The rate of reduction was depended on The amount of sodium hydroxide, Type of bleaching earth, Conditions of hydrogenation Time and temperature of steam deodorization Physical deacidification –Loss of sterols of between 10-70%

5. Continued: Sterol Glucosides –Sterol glycosides are removed during the oil refining process. –Degumming vegetable oils effectively removes sterol glucosides. –Fully refined oils are invariably free from sterol glucosides; –Alkali refines removes a portion of the sterol; –Both steam deodorization and physical refining further reduce the sterol glucosides.

6. Wedel ( Quantitative studies of the pHLip Test Kit for B100, AOCS presentation, May 2007)

7. Critical Issue No ASTM standard on the analytical methods to determine sterol glucosides in biodiesel; The effect of refining and processing strategy on sterol glucoside content in biodiesel has not been fully elucidated; Few studies on reducing the sterol glucoside content in biodiesel blends.

8. Objectives Evaluate different refining and processing strategy to reduce the sterol glucoside content in soy-based biodiesel –The effect of vacuum distillation on the sterol glucoside content in biodiesel; –The effect of cold filtering on the sterol glucoside content in biodiesel and blends Using filter membrane Using gravity to settle out, and separate

9. Chemical refining –Water degumming –Chemical neutralization –Bleaching –Deodorization Physical refining –Acid degumming –Bleaching –Deordorization

10. Process Water degumming: oil 70 o C at 175 rpm+water(3%) ----at 250 rpm for 30 min----Centrifuging-----supernatant Chemical neutralization: water-degummed oil at 55 o C + NaOH (10%) at 175 for 40 min----heated to 75 o C---- centrifuge----oil was collected and washed with water. Acid degumming:oil 60 o C+0.15% citric acid at 10000rpm for 1 min__ 60min at 40 o C--- cnetrifuging Bleachinng: The degummed oil at 95 o C under reduce pressure (50 mbar)+ bleaching earth --- mixed at 85 rpm for 30 mil--- filter Depdorization: at 240 o C (2 mbar) with injection of 1.5 and 1% steam, respectively.

11. Sterol Glycosides Crude soy oil: 2300ppm SG 19300 ppm SG in gum Hydrolysis may be carried out by the application of steam and alkali or acid: biodiesel made from acid oil can have high levels of SG

13. Analysis method Total sterols were analyzed by acid and alkaline hydrolysis, hydrolysis, derivatisation to trimethylsilyl(TMS)-esters and anlysis using GC-FID. The steryl glucosides were isolated by solid phase extraction (SPE), and then was hydrolysed with acid to release the free sterols

14. Extraction and Quantitation of Sterols

15. Analytical Steps