ABSTRACT: Steryl ester content of refined and interesterified corn, soybean, and rapeseed oils has been measured via clean- up on a short silica gel column, followed by high performance liquid chromatography with evaporative light-scattering mass detector. Chemical interesterification, catalyzed by sodium methoxide, led to random positional distribution of fatty acids in triacylglycerols and some increase in the steryl ester content of all three oils. Enzymatic interesterification, catalyzed by the immobilized lipase from Rhizomucor miehei (Lipozyme), re- sulted in a distinct reduction in steryl ester content, but essen- tially no alteration in positional distribution of fatty acids in tri- acylglycerols occurred. Formation of steryl esters during chemi- cal and enzymatic interesterification was also examined by radioactive tracer technique with [4– 14 C]β-sitosterol added as marker to refined rapeseed oil and measurement of the radioac- tive steryl esters formed. Chemical interesterification of rape- seed oil resulted in moderate formation (10% of total radioac- tivity) of radioactive β-sitosteryl esters. Enzymatic interesterifi- cation of the oil, catalyzed by Lipozyme, led to little formation of radioactive β-sitosteryl esters, whereas with the lipase from Candida cylindracea high proportions (>90% of total radioac- tivity) of 14 C-labeled β-sitosteryl esters were formed. JAOCS 74, 93–96 (1997). KEY WORDS: Candida cylindracea lipase, chemical inter- esterification, enzymatic interesterification, positional distribu- tion of fatty acids, radioactive tracer technique, Rhizomucor miehei lipase, β-sitosterol, steryl esters. Sterols occur in plant oils partly as unesterified sterols and partly as steryl esters of fatty acids, in addition to steryl gly- cosides and esterified steryl glycosides (1,2). We have re- cently developed a method for quantitative determination of steryl esters in oils by clean-up on a short silica gel column, followed by high-performance liquid chromatography (HPLC) with detection by means of an evaporative light-scat- tering mass detector (Ferrari, R., E. Schulte, and K.D. Mukherjee, manuscript in preparation). The above method was used to monitor changes in steryl esters in plant oils at different stages of industrial refining (Ferrari, R., E. Schulte, and K.D. Mukherjee, manuscript in preparation). Here we re- port the alterations in steryl ester content of plant oils by in- teresterification, e.g., chemical interesterification (random- ization) catalyzed by sodium methoxide, and enzymatic in- teresterification catalyzed by lipase from Rhizomucor miehei (Lipozyme). Moreover, we report the formation of steryl es- ters during chemical interesterification and enzymatic inter- esterification of rapeseed oil, catalyzed by Lipozyme and the lipase from Candida cylindracea, as monitored by a radioac- tive tracer technique. [4– 14 C]β-Sitosterol was added as marker to the oil, and the formation of radioactive steryl es- ters during interesterification was followed. EXPERIMENTAL PROCEDURES Materials. Refined corn oil was a product of Rafinacoes de milho Brasil Ltda. (São Paulo, Brazil), and soybean and rape- seed oils were provided by Cocamar Ltda. (Maringá, Brazil). Immobilized lipase from R. miehei (Lipozyme IM20, 25 batch interesterification units/g) was a generous gift of Novo Nordisk (Bagsvaerd, Denmark). Powdered lipase preparation from C. cylindracea (syn. C. rugosa) with an activity of 850 U/mg was purchased from Sigma (Deisenhofen, Germany). β-Sitosterol and [4– 14 C]β-sitosterol (56 mCi/mmol) were products of Sigma and Amersham (Braunschweig, Germany), respectively. β-Sitosteryl oleate was prepared according to Gupta et al. (3). All reagents and adsorbents were of analytical grade and pur- chased from E. Merck (Darmstadt, Germany). Chemical interesterification. Refined oil (10 g) and 15 mg sodium methoxide were placed in a round-bottom flask, and the mixture was stirred magnetically at 110˚C for 30 min under vacuum generated by a water jet aspirator. The reaction products were cooled to room temperature, dissolved in hexane, and washed repeatedly with an aqueous 1% (wt/vol) sodium chloride solution to remove the soaps completely. The hexane solution of the interesterified product was dried over sodium sulfate and stored refrigerated under nitrogen. Copyright © 1997 by AOCS Press 93 JAOCS, Vol. 74, no. 2 (1997) 1 Part of doctoral thesis of Roseli Ap. Ferrari to be submitted to Faculdade de Engenharia de Alimentos, Universidade de Campinas, Campinas, Brazil. *To whom correspondence should be addressed at Institute for Biochemistry and Technology of Lipids, H.P. Kaufmann-Institute, BAGKF, Piusallee 68, D-48147 Münster, Germany. Alteration of Steryl Ester Content and Positional Distribution of Fatty Acids in Triacylglycerols by Chemical and Enzymatic Interesterification of Plant Oils 1 R.Ap. Ferrari a , W. Esteves b , and K.D. Mukherjee a, * a Institute for Biochemistry and Technology of Lipids, H. P. Kaufmann-Institute, Münster, Germany, and b Laboratory of Oils and Fats, University of Campinas, Campinas, Brazil