GB860305A - Tuberculin active peptide - Google Patents

Abstract

A tuberculin-active peptide of high purity and potency and capable of giving rise to a uniform reaction is obtained by extracting sterilized bacterial cells of Mycobacterium tuberculosis, especially var. hominis H37RV, with a dilute inorganic or organic acid of r H 1 to 3, neutralizing the extract to a r H of 7.0, adding picric acid to the solution to precipitate the peptide as its picrate and subsequently separating the desired peptide from the picrate. The acid may be dilute hydrochloric, sulphuric or phosphoric of 0.1 N, or aqueous citric acetic or propionic acid, extraction being at 37 DEG C. for 72 hours. After filtering and washing with dilute acid, sodium hydroxide is added to r H 7.0, allowed to stand, any precipitate removed and saturated aqueous picric acid added to precipitate the picrate. The picrate is treated with a solution of hydrochloric acid in methanol or ethanol, the insolubles removed and the peptide precipitated by the addition of acetone. Further purification is obtained by repeating the acid-alcohol treatment. The peptide forms pale yellow crystals containing 15.9% nitrogen; it is a single component of low molecular weight, free from sugar and nucleic acid; soluble in water, methanol and ethanol; a neutral aqueous solution becomes opalescent at r H 8.5 and forms a precipitate at r H 9.0; it is stable on heating to 70 DEG C. for 15 minutes as regards the tuberculin activity.