Mw. Trucksess et al., HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC DETERMINATION OF 1,1'-ETHYLIDENEBIS(L-TRYPTOPHAN) IN L-TRYPTOPHAN PREPARATIONS, Journal of pharmaceutical sciences, 83(5), 1994, pp. 720-722
In studies to determine the cause or causes of the eosinophilia myalgi
c syndrome (EMS) and to monitor the purity of L-tryptophan preparation
s, an HPLC method has been developed for determining 1,1'-ethylidenebi
s(L-tryptophan) (EBT) in L-tryptophan (W) preparations. The W preparat
ions are extracted with 0.1% trifluoroacetic acid (TFA) and filtered,
and the EBT is purified by passage through a Sep-Pak C-18 cartridge. T
he cartridge is washed with water and 6% acetonitrile in water, and EB
T is eluted with methanol. The water-diluted eluate is then chromatogr
aphed on a silica-based, reversed-phase HPLC column with a gradient of
water and 80% acetonitrile, both solvents containing 0.1% TFA. EBT ab
sorbance is measured at 280 nm. The average recovery of EBT from L-try
ptophan powder, spiked over the range 1.2-4.8 mu g/g, was 91%. The lim
it of determination was similar to 0.6 mu g/g. Sixteen test samples of
W products manufactured by the company to which most of the cases of
EMS have been traced contained >70 mu g of EBT/g. Three nonpatient-rel
ated test samples either did not contain EBT or contained <2 mu g of E
BT/g.