Progress 09/01/05 to 08/31/10
Outputs
OUTPUTS: Hydrogenation is used in the fats and oil industry to improve the consistency and oxidative stability of fats used in margarines, shortenings, spreads, and confectioneries to improve texture, and modify their melting and crystal behavior. Unfortunately, hydrogenation results in the production of trans fatty acids (FA). We proposed to synthesize structured lipids (SL) as a viable alternative to achieve the desired properties of fats without the trans FA. Our overall project goal was to use enzyme to produce trans-free SL for margarine application. Our overall aim was to use canola oil and stearic acid as substrates for the enzymatic production of trans-free SL as value-added products with potential for possible food and nutritional applications. The following objectives were explored: 1) synthesize canola oil-based SL using two lipases, an immobilized lipase, RM IM from Rhizomucor miehei and Candida rugosa lipase isoform (LIP1) as the biocatalysts with stearic acids incorporated into the canola oil 2) compare the physical properties of enzymatically produced SLs with chemically produced SL of canola oil and stearic acid, and 3) study food application of the trans-free SLs in model margarine (water-in-oil emulsion system) and compare the physical properties of the margarine made with enzymatically and chemically synthesized SLs. The most significant findings and dissemination activities from our research are: Response surface methodology was used to study the effects of Lipozyme Rm IM from Rhizomucor miehei and Candida rugosa lipase isoform 1 (LIP1) on the preparationof SLs. We found that ethyl stearate was a better acyl donor compared with stearic acid for Lipozyme RM IM lipase and that LIP1 was not as effective. The solid fat content (SFC) and crystal properties of trans-free SL synthesized from stearic acid and canola oil was optimized by blending with different amounts of palm mid-fraction (PMF). Both acidolysis and interesterification reactions were successfully employed to synthesize SLs for the formulation of trans-free margarines with desirable textural properties, SFC, and low atherogenicity. Various emulsifiers were used to improve the polymorphic behavior of the SLs. We also evaluated the physical and sensory attributes of an experimental trans-free margarine spread (MG-X) and two commercial margarine spreads (MG-A and MG-B). The trans-free margarine spread was formulated with a blend containing a SL synthesized by reacting canola oil with 40% stearic acid (w/w), PMF, and cottonseed oil (CTO). No trans fatty acids were detected in MG-X, whereas the trans fatty acid contents of MG-A and MG-B were 0.3 and 3.7% (w/w), respectively. MG-X was considerably firmer than MG-A and MG-B, less cohesive, and its adhesiveness was intermediate between those of MG-A and MG-B. MG-X's stability to syneresis was also intermediate between those of MG-A and MG-B. Sensory evaluation indicated that MG-X was comparable to MG-A in spreadability and texture only, but was significantly different from MG-B in all other attributes. PARTICIPANTS: Thomas A. Foglia Lead Scientist, Lipids Research ERRC/ARS/USDA 600 East Mermaid Lane Wyndmoor, PA 19038 USA Dr. Foglia's laboratory helped with the structural analysis of structured lipids synthesized in our laboratory using their GC-MS, HPLC-MS-APCI, NMR,etc and training the graduate student Stephen E. Lumor while at ERRC. William Kerr Department of Food Science and Technology University of Georgia Athens, GA 30602 Dr. Kerr was a co-PD for this project. He helped with the physical and rheological properties analyses. Training and Development: Graduate students and postdoc trained are: Stephen E. Lumor, PhD Byung Hee Kim, PhD Byung Hee Kim, Postdoctoral Research Associate TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Through this proposal, we were able to enzymatically synthesize trans-free SLs for margarine and spreads formulations. We believe these SLs can be used as trans-free alternatives to hydrogenated fats in baking shortenings, margarines, and spreads. Blending various fats and canola oil at different ratios helped us achieve desirable trans-free fats which were then used to produce trans-free margarines with low atherogenic, and desirable sensory and textural properties. The low atherogenic index of the SLs means they may help reduce LDL-cholesterol and lipoprotein levels in the blood thereby reducing the incidence of coronary heart disease among Americans. The resources provided by this grant allowed us to support and graduate 2 PhD students and train 1 postdoctoral student. Several invited and voluntered professional presentations were made in US and abroad to disemminate the results of our studies.
Publications
- Lumor, S.E., Pina-Rodriguez, A.M., Shewfelt, R.L. and Akoh, C.C. (2010). Physical and sensory attributes of a trans-free spread formulated with a blend containing a structured lipid, palm mid-fraction, and cottonseed oil. J. Am. Oil Chem. Soc. 87:69-74.
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Progress 09/01/08 to 08/31/09
Outputs
OUTPUTS: We optimized the solid fat content (SFC) and crystal properties of trans-free structured lipids (SL) synthesized by incorporating stearic acid into canola oil. The SLs were blended with varying amounts of palm mid-fraction (PMF). SFC and crystal polymorphism were improved. Addition of sucrose stearate (S-170), sorbitan tristearate (STS), and distilled monoglycerides (DMG) to one of the blends, SL40:PMF (70:30, w/w), did not improve crystal polymorphism, but had significant effects on crystal morphology. The emulsifiers significantly delayed crystal growth, resulting in smaller crystal sizes compared to the control. They were unable to inhibit the formation of granular crystals (30 to 140 μm), which are undesirable in margarine, after 4 weeks of storage at 0˚C. Blends treated with S-170 and STS showed many small evenly distributed crystals interspersed with large crystal aggregates (after 4 weeks of storage), whereas the blend treated with DMG and the control showed irregularly shaped globular crystals, also interspersed with large crystal aggregates. The esterification and hydrolytic activities of free and immobilized Candida rugosa lipase isoform 1 (LIP1) were also investigated. Results indicated that LIP1 had little esterification activity, which was not significantly improved upon immobilization. LIP1 showed hydrolytic activity comparable to Lipozyme RM IM. The hydrolytic activity improved significantly upon immobilization. Immobilization on Celite 545 produced the highest increase in hydrolytic activity. PARTICIPANTS: PhD student, Stephen E.Lumor has graduated from this project. Post doctoral training was given to Dr. Byung Hee Kim TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
We believe that reducing the amount of trans fatty acids in foods will help in improving the health of consumers especially by reducing the incidence of coronary heart disease. It will mean more productive life and more time spent in the work place and less trips to the doctor.
Publications
- Lumor, S.E., Kim, B.H., and Akoh, C.C. 2008. Optimization of solid fat content and crystal properties of a trans-free structured lipid by blending with palm midfraction. J. Agric. Food Chem. 56:9294-9298.
- Lumor, S.E. and Akoh, C.C. 2008. Esterification and hydrolytic activities of Candida rugosa lipase isoform1 (LIP 1) immobilized on Celite 545, Duolite A7, and Sephadex G-25. J. Agric. Food Chem. 56:10396-10398.
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Progress 09/01/07 to 08/31/08
Outputs
OUTPUTS: Two types of structured lipids (SLs) for formulating trans-free margarines were synthesized by (1) lipase-catalyzed acidolysis of canola oil (CO) with stearic acid (SA) and (2) lipase-catalyzed interesterification of CO with palm stearin (PS) and palm kernel oil (PKO), respectively. CO/SA-based SLs were further blended with palm mid-fraction (PMF) and were treated with emulsifiers, such as sucrose stearate (S-170), distilled monoglycerides (DMG), and sorbitan tristearate (STS). We determined the atherogenicity and physical properties of SLs and the textural properties of margarines made with the SLs. Among a series of blends of CO/SA-based SLs and PMF, the SLs prepared from the 40% blend (SA/CO, w/w%) and PMF blend in the weight ratio of 70:30 (SLs:PMF, w/w) had predominantly β′ polymorphs and had the most desirable solid fat content for margarine formulation. However, the addition of S-170, STS, and DMG to the blend did not further improve its polymorphic properties. Among a series of CO/PS/PKO-based SLs, the SLs prepared from 50:30:20 and 60:25:15 blends (CO:PS:PKO, w/w/w) had lower and similar atherogenic index than the commercial trans (CTMF) and trans-free margarine fats (CTFMF), respectively and had more β′ than β polymorphs. The margarines made with them had similar textural properties to margarines made with CTMF and CTFMF, respectively. Therefore, CO/SA-based SLs:PMF blends and CO/PS/PKO-based SLs were suitable for formulating trans-free margarines with desirable textural properties or low atherogenicity. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Blending various fats and canola oil at different ratios helped us achieve desirable trans-free fats which were then used to produce trans-free margarines with low atherogenic and desirable textural properties. Obviously, eliminating and/or reducing the amount of trans fats in foods will lead to healthier Americans.
Publications
- Lumor, S.E., Jones, K.C., Ashby, R., Strahan, G., Kim, B.H., Lee, G.C., Shaw, J.F., Kays, S.E., Chang, S.W., Foglia, T.A. and Akoh, C.C. 2007. Synthesis and characterization of canola oil-stearic acid-based trans-free structured lipids for possible margarine application. J. Agric. Food Chem. 55:10692-10702.
- Kim, B.H., Lumor, S.E., and Akoh, C.C. 2008. Trans-free margarines prepared with canola oil/palm stearin/palm kernel oil-based structured lipids. J. Agric. Food Chem. 56:8195-8205.
- Lumor, S.E., Kim, B.H., and Akoh, C.C. 2008. Optimization of solid fat content and crystal properties of a trans-free structured lipid by blending with palm mid-fraction. J. Agric. Food Chem. (in press).
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Progress 09/01/06 to 08/31/07
Outputs
Lipases and a chemical catalyst were used to incorporate stearic acid into canola oil to produce trans-free structured lipid (SL) as a healthy alternative to partially hydrogenated fats for margarine formulation. Response surface methodology was used to study the effects of Lipozyme RM IM from Rhizomucor miehei and Candida rugosa lipase isoform 1 (LIP1) and two acyl donors, stearic acid and ethyl stearate, on the incorporation. Lipozyme RM IM and ethyl stearate gave the best result. More SLs were synthesized using Lipozyme RM IM and the products compared to SL made by chemical catalysis, and fat from commercial margarines. After short-path distillation, the products were characterized by GC and RPHPLC-MS to obtain fatty acid and triacylglycerol profiles, 13C NMR spectrometry for regiospecific analysis, X-ray diffraction for crystal forms, and DSC for melting profile. Stearic acid was incorporated into canola oil, mainly at the sn-1, 3 positions for the lipase reaction
and no new trans fatty acids were formed. Stearic acid icorporation increased with increase in substrate ratio for the lipase-catalyzed reaction but were less for the chemically synthesized SL. Most SL products did not have adequate solid fat content or β' crystal forms for tub margarine, although these may be suitable for light margarine formulation. Ongoing studies include increasing solid fat content and regulating the crystal forming habit of the SLs by using emulsifiers and/or by blending the SLs with palm oil fractions.
Impacts
Reducing the amount of trans fats or artery clogging fats in the diet will go a long way in decreasing the incidence of coronary heart disease in American. It will mean less trips to the hospital and a healthy workforce to power the economy and improvement in the quality of life for most Americans.
Publications
- Lumor, S.E., Jones, K.C., Ashby, R., Strahan, G., Kim, B.H., Lee, G.C., Kays, S.E., Chang, S.W., Foglia, T.A. and Akoh, C.C. 2007. Synthesis and characterization of canola oil-stearic acid-based structured lipids for possible margarine application. J. Agric. Food Chem. (in review).
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Progress 09/01/05 to 09/01/06
Outputs
Trans-free structured lipids (SLs) with potential for use in food applications such as margarine were synthesized. Stearic acid and ethyl stearate were the acyl donors and Lipozyme RMIM and LIP1 lipase isoform from Candida rugosa were the biocatalysts. Response surface methodology was used to model the response (stearic acid incorporation). The amount of each enzyme was 10% of reactant weight. The data obtained was fitted to a quadratic model. The reproducibility and validity of the model were 0.99 and 0.74, respectively. The normal probability and residuals plots showed no violations of model assumptions of normality and constant error variance, respectively. The results showed that ethyl stearate, as acyl donor led to more incorporation compared with stearic acid. Also, Lipozyme RM IM was found to strongly favor the process more than LIP1. Incorporation was found to increase linearly with substrate ratio. Increasing temperature had positive influence on the process
until about 50 degrees C with stearic acid as acyl donor. The effect of time on the process was not so obvious. Based on the above findings, 55-70g of SLs were synthesized with Lipozyme RMIM as biocatalyst, while stearic acid was the acyl donor for economical reasons. The amount of stearic acid used varied from 10-40% by weight of substrates. The reaction time and temperature were set at 12h and 50 degrees C, respectively. Free fatty acids were removed via short-path distillation from the products, and their melting profiles determined by differential scanning calorimetry. The products were analyzed for the presence of trans-fatty acids using GC. The thermograms showed the presence of three new peaks in the SL products compared with that of canola oil. These new peaks were due to the formation of higher melting triacylglycerol (TAG)species as a result of stearic acid incorporation into canola oil. These peaks were within the melting range of four commercial margarines. Our study
showed that SLs made with 10, 20, and 30% stearic acid would be most suitable for margarine. The total trans fat content of the SL samples and canola oil were between 0.07 and 0.08% by total weight of fatty acids. The presence of these trans fatty acids (18:1n-t and 18:2n-t) in the samples were most probably not due to the interesterification process, but rather, from canola oil since there were no significant differences between the trans fat contents of the canola oil and the SL samples. Commercial margarine samples were found to contain 0.23-3.3% trans fatty acids.
Impacts
Trans-free structured lipids were successfully synthesized. Trans faty acid contents of foods such as baking shortenings, margarines, spreads, and used frying oils are estimated between 0-35% of total fatty acids. Trans fatty acids are known to elevate LDL-cholesterol and lipoprotein levels in the blood. The use of enzymatically synthesized structured lipids to replace trans fatty acids in spreads, margarines, and other food products will lead to healthier diets for Americans. It is hoped that this goal is achievable, in part, through the completion of the current project.
Publications
- Lumor, S.E., Lee, G.C., Shaw, J.F. and Akoh, C.C. 2006. Enzymatic incorporation of stearic acid into canola oil: effects of lipase and acyl donor types. Presented at the American Chemical Society National Meeting, Atlanta, GA, March 26-30.
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