Iodine → L-Glutamic Acid

A stirred mixture of pGlu--His--Trp--Ser--Tyr--Gly--OH (XXXVIII) (16.6 gm) and 1-hydroxybenzotriazole (3 gm) in dry DMF (250 ml) was treated with DCI (4.5 gm) at 0°C and after 3 hours, the dihydrofluoride salt of Leu--Arg--Pro--Gly--NH2 (LXVI) (10.5 gm) and Et3N (3.1 ml) in dry DMF were added and stirring was continued at room temperature for a further 36 hous. The reaction mixture was evaporated in vacuo and the residue was treated with H2O (200 ml). The insoluble DCI-urea was removed and the filtrate was evaporated in vacuo to afford 38 g of crude LHRH. A portion of this (6.47 gm) in 1% AcOH (250 ml) was passed over Bio-Rad AG1-8 (OAc) (1×15 cm) and the filtrate was evaporated in vacuo. The residue, in H2O (15 ml), was chromatographed over CMC (4.5 × 27 cm) and eluted in a stepwise manner with NH4OAc buffers of pH 6.9: (0.005, 0.01, 0.025, 0.075 and 0.1 M). The pooled LHRH fractions, located in the 0.075 M eluate by the Pauly reaction, were lyophilized. The LHRH (3 gm) was further purified by gel filtration over Bio-Gel P-2 (400 gm) (110 cm × 2.5 cm) with 1.3% AcOH. Individual fractions (5 ml) were collected at a flow rate of about 5 ml.min and the pure LHRH was located in fractions 21-31 by the Pauly reaction and by tlc. Fractions showing one spot on tlc were pooled and lyophilized to constant weight affording 2.85 gm. (59%) of pure pGlu--His--Trp--Ser--Tyr--Gly--Leu--Arg--Pro--Gly--NH2 (XXII) (LHRH): [α]D22 -33.9 (c, 0.66, MeOH); [α]D22 -46.6 (c, 0.73, 1% AcOH); Rf1 0.64 and Rf3 0.37, single spot with Pauly, chlorine-tolidine and I2 reagents. Amino acid analysis gave Glu 1.08; His 0.99; Trp 0.87; Ser 0.85; Tyr 0.98; Gly 2.07; Leu 1.04; Arg 1.02; Pro 0.97; NH3 1.08.

1. custom(OAc) (1×15 cm) and the filtrate was evaporated in vacuo
2. customThe residue, in H2O (15 ml), was chromatographed over CMC (4.5 × 27 cm)
3. washeluted in a stepwise manner with NH4OAc buffers of pH 6.9
4. customThe pooled LHRH fractions, located in the 0.075 M eluate by the Pauly reaction
5. filtrationThe LHRH (3 gm) was further purified by gel filtration over Bio-Gel P-2 (400 gm) (110 cm × 2.5 cm) with 1.3% AcOH
6. customIndividual fractions (5 ml) were collected at a flow rate of about 5 ml
7. custommin and the pure LHRH was located in fractions 21-31 by the Pauly reaction and by tlc
8. customaffording 2.85 gm

ORD 기록

ord-329fc832e2714ddc8177644dae9605fc

출처 데이터셋

uspto-grants-1976_04

DOI

10.6084/m9.figshare.5104873.v1

특허 번호

US03953416