Purification of PAGs
To verify the universality of the enzymatic
esterification method, the optimal
conditions of synthesis of CG obtained above were used to
synthesize FG and p -HCG. The
optimal conditions were as follows: 0.01 mol PAs (CA, FA, orp -HCA) was reacted with 1.5 mol of glycerol catalyzed by 25 % of
Lipozyme 435 (relative to the weight of total substrates) at 80oC under 86.7 kPa for 10 h. Then the enzyme was
filtered off. The filtrate was extracted with ethyl acetate (reaction
mixture/ethyl acetate, 1:1, V/V) for three times. The extracted organic
phase was combined and concentrated using a rotary evaporator at 50oC. When the liquid change to be turbid, the liquid
was crystallized at ambient temperature for 24 h. The solid was
carefully separated after centrifugation (3,000 rpm) for 10 min. Then
washed with distilled water and subsequently freezing dried. The liquid
was concentrated and separated by silica gel column chromatography using
chloroform/ toluene/ methanol/ acetate acid (CG, 32:20:10:1; FG andp -HCG, 32:20:1.5:1) as mobile phase. After vacuum evaporation,
the white solid was obtained. Combined with the freeze dried solid, 2.16
g of CG (85.0 % of isolated yield), 2.20 g of FG (82.1 % of isolated
yield) and 1.92 g of p -HCG (80.7% of isolated yield) were
obtained respectively. The purity of isolated PAGs were reconfirmed by
TLC and HPLC.