Transesterifi cation of Fatty Acids in Freeze-Dried Forage Samples

Procedures for preparing fatty acid methyl esters from for-ages using single-step transesterifi cation have been well doc-umented. Outen et al. (1976) demonstrated a one-step extraction and esterifi cation method using benzene and 5 % methanolic HCl. Sukhija and Palmquist (1988) later recommended that benzene or toluene be used for extraction and formation of methyl esters. The use of benzene and toluene is now discouraged due to high toxicity and carcinogenic properties (USEPA, 2002). Recogniz-ing the hazards of benzene and toluene, Whitney et al. (1999) substituted these solvents with 7 % (1.03 M) BF3 in CH3OH for direct transesterifi cation of feedstuff s. Although BF3 has proven to be an eff ective catalyst for direct transesterifi cation of fatty acids in forages (Whitney et al., 1999) and animal tissues (Rule, 1997), it is highly volatile and can be toxic if inhaled (NIOSH, 2004). Boron-trifl uoride also must be sealed with N2 and stored in a cool, dark environment to main-tain reactivity. In contrast, methanolic HCl is less volatile than BF3, maintains shelf-life longevity without special preparation, and costs substantially less than BF3. Garcés and Mancha (1993) demonstrated that methanolic HCl is an effi cient catalyst for preparing fatty acid methyl esters, with solubility of transmethylated end products a factor limiting its utility. However, we have overcome potential problems wit