In this study, colchicine, 2-demethylcolchicine, 3-demethylcolchicine, 3-demethyl-N-formyl-N-deacetylcolchicine, colchifoline, 2-demethylcolchifoline, demecolcine
, 2-demethyldemecolcine and 2-demethyl-[??]-lumicolchicine 12 were isolated from all the separated parts of C.
Briefly, the matured eggs with the first polar body were cultured in medium supplemented with 0.4 mg/mL demecolcine
and 0.05 mol/L sucrose for 1 h.
To block the mitotic process of the cells at the metaphase stage, Demecolcine
(Sigma-Aldrich) was added for the last 2 h of culture at a final concentration of 10 [micro]g/ml.
sibthorpioides, including (A) 2-ethylacridine (20.4%), (B) 2-methyl-3-O-tolyl-6-hydroxy-4(3H)-quinazolinone (18.0%), (C) 3-(4-hydroxy-methylphenyl)-2-methyl-4(3 H)-quinazolinone (23.1 %), (D) 9,10-dihydro-9,9,10-trimethyl-anthracene (19.8%), and (E) demecolcine
This possible difference could be probed by microinjecting a microtubule poison, for example demecolcine
, directly into the vegetal ooplasm.
Moreover, three microtubule poisons - colchicine, demecolcine, and nocodazole - inhibit the normal movement of oil droplets toward the vegetal pole, eliminate the saltatory motion of small inclusions, and slow the growth of the blastodisc: eggs treated with [Beta]-lumicolchicine, an inactive derivative of colchicine, segregated normally (Abraham et al., 1993a).
Working solutions of demecolcine (N-deacetyl-N-methyl-colchicine; Sigma) were prepared by diluting (at least 100X) an aqueous stock solution of 0.35 mM demecolcine.
In some experiments, we treated eggs with both CCD (10 [[micro]gram] [ml.sup.-1]) and demecolcine (0.35 [[micro]molar]).
Eggs that were in a solution of demecolcine and being irradiated with UV light were fixed on the stage of the microscope for 5 min before being transferred to a vial of fixative.
Effects of cytochalasin D and demecolcine on ooplasmic segregation and development
The effects of demecolcine on the eggs were those previously described by Abraham et al.