The invention particularly relates to a method for separating and purifying cobra-
venom neurotoxin by combining
ion exchange and hydrophobic
chromatography. The method comprises the following steps: (1) weighing 4-5 parts by weight of cobra-
venom crude product, adding the cobra-
venom crude product into 0.04-0.05 part by volume of water or 0.01-0.05M of
phosphate buffer solution with the pH of 5.0-7.0, stirring to dissolve the cobra-venom crude product, centrifuging the solution at the temperature of 0-8 DEG C, discarding precipitates, and filtrating supernatant so as to obtain a
test sample solution A for later use; (2) purifying the
test sample solution A through
ion-exchange
chromatography, carrying out
gradient elution, carrying out HPLC (High-Performance Liquid
Chromatography) detection, and carrying out collecting so as to obtain a
test sample solution B for later use; and (3) purifying the test sample solution B through hydrophobic
chromatography, carrying out
gradient elution, carrying out HPLC detection, carrying out collecting, carrying out concentrating, and carrying out
drying thereby obtaining a pure product of cobra-venom
neurotoxin, wherein when the unit of parts by weight is g, the unit of parts by volume is correspondingly L. According to the method, operating steps are simplified, the separation and purification cycle is shortened, the yield is relatively high, the purity is relatively high, and thus the method is applicable to the industrial production of cobra-venom
neurotoxin.