Milk fat, true protein, and lactose contents were
determined by mid-infrared spectrophotometry using
a Milkoscan 4000 (Foss Electric, Hillerød, Denmark)
calibrated using historical samples of bovine and caprine
milk for which reference measurements had been
made (AOAC International, 1997). Total lipid in 100
mg of freeze-dried milk was converted to FAME by
incubation with 2 mL of 0.5 M sodium methoxide in
anhydrous methanol and 1 mL of hexane at 50°C for 15
min. After cooling, 1 mL of methanol and hydrochloric
acid (95:5 vol/vol) was added to the reaction mixture
and incubated at 50°C for 15 min. Methyl esters were
recovered in 1.5 mL of hexane, washed with 3 mL of
aqueous (6% wt/wt) potassium carbonate, and analyzed
using a GC (Agilent 7890A GC System, Santa
Clara, CA) equipped with a CP-Sil 88 capillary column.
The profile of FAME in a 2-μL sample at a split ratio of
1:50 was determined using a temperature gradient program
(Loor et al., 2005b), with hydrogen as the carrier
and fuel gas, operated at constant pressure (147 kPa).
Isomers of 18:1 were further resolved in an additional
analysis under the same conditions, with the exception
that a smaller sample volume (0.6-μL) was injected onto
the column. Peaks were routinely identified based on
retention time comparisons with commercial authentic