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a-Galactosidase from germinating Vicia sativa seeds has been extracted with a 0.9O/, NaCl solution. Treatment with manganous sulphate eliminated inactive fractions. The enzyme was precipitated from the supernatant with (NH,),SO, at 80 Oi0 saturation, redissolved in water and dialyzed. Purification was further increased by three successive steps of column chromatography. Only fractions showing u-galactosidase activity have been used for each subsequent step. Successive passage through two hydroxylapatite and one Sephadex column gave an enzyme with a specific activity 1,100 times higher than that of the crude extract, and with a yield of loo/,, from the original material.The enzyme thus obtained behaves as a single protein in electrophoresis on Cellogel. Sedimentation in a sucrose gradient and ultracentrifugation in an analytical ultracentrifuge suggest a molecular weight of about 30,000. The protein is rich in aspartic and glutamic acids and has a low cysteine content. Dansylation and the dinitrofluorobenzene method showed the presence of a single N-terminal residue which was identified as L-alanine. The purified a-galactosidase seems t,herefore to be a single polypeptide chain.The pH optimum of the a-galactosidase is 6.3. The enzyme is not very sensitive to thermal inactivation and is devoid of activity on u-glucosides and /?-galactosides. It hydrolyses phenylu-galactoside rapidly and galactinol very slowly. Unlike the a-galactosidase from coffee beans it does not liberate galactose from the galactomannans of Leguminoseae. Michaelis constants a t pH 6.3 are markedly different for the different substrates: K m = 9.7 x for p-nitrophenyla-galactoside ; 3.8 x for rafhose and 4.1 x lo-' for galactinol. I n the case of the two phenol galactosides a strong inhibition was observed with excess substrate. This inhibition does not take place with raffinose or galacthol as substrate. u-Galactosidase is inhibited by Hg2+, Ag+ and weakly activated by Fes+, Zn2+ and Mn2+. A transferase activity was observed with phenyl-a-galactoside as donor and various acceptors : glucose, mannose, galactose, cellobiose and sucrose. Raffinose also acts as an a-galactosyl donor. With galactinol as acceptor the following compound is formed : Galpal-6Galpal-l'-myoinositol. La germination des graines de legumineuses s'accompagne en g6nBral de la trks nette augmentation de l'activitd a-galactosidasique [6-91. De plus la d6polym6risation des galactomannanes et autres polysaccharides visqueux, au cours de cette germinatjion, facilite les operations de purification. Les graines germ6es de Vicia sativa sont riches en diverses glycosidases. Nous avons rBussi A purifier une a-mannosidase [lo]. Nous avons poursuivi nos essais en vue de purifier l'u-galactosidase et avons obtenu cet enzyme B un degr6 de puret6 trhs avanc6. Nous en d6crivons les principales propri6t6s. MATGRIEL ET M6THODESSubstrats. Phthyl-a-D-galactoside pr6par6 selon la mBthode d'Helferich [l I], p-nitroph6nyl-u-D-galactoside synth6tis6 selon Wakabayashi [ 121.Le galactinol (Galpul-1'-my...

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An epidemiological study of negative middle ear pressure in children made it possible to test its relationship to conductive hearing loss. About 350 children were subjected to a screening procedure recording audiogram and middle ear pressure five times during a 12-month period. Those children who failed to perceive just one tone or who had a middle ear pressure equal to or worse than -150 mmH2O in one or both ears were referred to the Hearing Clinic for conventional audiometry and middle ear pressure measurement each month. By computing the weighted average of the regressions for each child, a straight linear relationship was found between negative pressure and conductive hearing loss. In addition, a frequency dependence was found, the hearing loss being maximal at about 500 Hz. In general, the study shows that tympanometry is of limited value in predicting hearing loss in a child. The threshold for pathology of about -150 mmH2O, being a predisposing factor in secretory otitis media, corresponds to the upper confidence limit of the normal range of hearing loss found in this series. There is no distinct value of negative pressure that clearly distinguishes between normal and pathological condition, but it is concluded that a middle ear pressure worse than -150 mmH2O should be considered a probable hearing handicap.