David N. Kramer

Lipolysis of fluorescein and eosin esters. Kinetics of hydrolysis

Abstract A series of fluorescein and eosin esters have been prepared for use in the assay of the activity of lipase and acylase: the diacetyl, dipropionyl, dichloropropionyl, dibutyryl, divaleryl, and dicaproyl esters of fluorescein, and the diacetyl, dipropionyl, and dibutyryl esters of eosin. The eosin esters were found to hydrolyze extremely slowly in the presence of enzyme, even at pHs as high as 10.0. The rate of hydrolysis of the fluorescein esters by all enzymes except porcine pancreas lipase was found to decrease in the order: acetyl > propionyl > butyryl > valeryl > caproyl. With porcine pancreas, the divaleryl and dicaproyl esters of fluorescein had a higher rate of hydrolysis than the dibutyryl ester. Steapsin, porcine pancreas, and wheat germ lipase, as well as acylase, catalyze the hydrolysis of fluorescein esters to fluorescein. The order of activity is porcine pancrease > acylase > steapsin > wheat germ lipase. The kinetic and thermodynamic constants ( K′ m , k 3 , Δ E ∗ , Δ H′, Δ F′, Δ S′ ) for the hydrolysis of the various esters by four different enzymes are reported. Sodium taurocholate was found to increase the rate of chymotrypsin hydrolysis of the fluorescein esters. A decrease in Km, Δ E ∗ , ΔH, ΔF, and ΔS and an increase in k3 were observed when this bile salt was added to the chymotrypsin reaction solutions. No effect on lipase was observed, thus indicating that the probable effect of taurocholate on lipolytic hydrolysis is in the solubilization of the substrate.

Fluorometric determination of hyaluronidase and of Cu(II), Fe(II), and cyanide ion inhibitors

Abstract : A fluorometric method is described for the assay of the enzyme hyaluronidase and of cupric, ferrous, and cyanide ions, which inhibit the enzymic activity. The method for hyaluronidase is based upon the hydrolysis of nonfluorescent indoxyl acetate by the enzyme to give the highly fluorescent indigo white. The metal ions can be determined by recording their inhibitory effect on the enzymic activity. By this procedure, from 0.0010 to 0.0330 micrograms/ml of hyaluronidase, 0.10 to 4.0 micrograms/ml of CN(-), and 0.20 to 12 micrograms/ml of Fe(++) or Cu(++) could be determined with standard deviations of plus or minus 1.8% for hyaluronidase and plus or minus 2.3% for the inhibitors. Evidence is offered to show that the activity of hyaluronidase, and not esterase, is measured. There appears to be a good correlation between the fluorometric method and other methods for hyaluronidase, and this technique should be a good one for the rapid, sensitive determination of this enzyme and of cupric, ferrous, and cyanide ions, which selectively inhibit its activity. (Author)

Some theoretical considerations in the microchemical detection of airborne toxic chemical agents

Abstract The increasing use of toxic phosphonates, thiophosphonates, phosphates, and carbamates as pesticides, insecticides, corrosion inhibitors, and antiknock compounds has shown the desirability of developing highly sensitive detector devices for use in industry, agriculture, food distribution centers, and the home. This paper deals with some of the theoretical considerations in the microchemical detection of toxic chemical agents as aerosols or gases in the absence of concentration or sampling devices. It appears that, even if the kinetics of a stoichiometric reaction between toxic gases or aerosols and porous solid detectors is assumed to be adequate, a sufficient amount of reaction to produce visible color in seconds will be possible only in regions of very high air flow velocities.

A SPECIFIC AND SENSITIVE METHOD FOR THE DETERMINATION OF BENZILIC ACID

Abstract A specific, sensitive and relatively simple procedure is described for the quantitative analysis of benzilic acid. The method is based on α-naphthofuchsone formation obtained by condensation of benzilic acid with α-naphthol in the presence of concentrated sulfuric acid and measuring the resultant hydroxytriaryl methyl cation at 558 mμ. No interference was observed with other glycolic acid derivatives or oxygen-bearing compounds. As little as 5 × 10−6 M benzilic acid is determinable.