A comparison of the acid Beta-Glycerophosphatases of the Lutz Hamster Sarcoma and of the Walker 256 Carcinosarcoma
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Abstract
The discovery of tissue acid phosphatase in 1934 by Davies (23) was foreshadowed by the demonstrations of Demuth (24) of an acid phosphatase acitivity in human urine. Bamann and Riedal (8) were among the first inyestigators to demonstrate that animal tissues that hydrolyzed glycerophosphate in the alkaline range (pH-9.4) possessed a second optimum pH in an acid medium (pH-5.5).
Phosphatases from various sources although differing primarily in their pH optimum, activating and inhibiting effect, do overlap in their effect on various substrates.
Using .017 Molar sodium-beta-glycerophosphate as substrate, the pH optima of the Lutz hamster sarcoma and of the Walker 256 carcino-sarcoma were determined. The pH optimum of each tumor type was 5.0. This was determined by preparing a series of solutions from pH 3.5 to pH 6.O.
The two tumor types showed similar degrees of activity. A solution of 0.1 mg.P/ml. of a standard phosphate was used as reference. The maximum activity of the Lutz hamster sarcoma at pH 5.0 using the method of Hoffman (41) was 0.153 mgP/ml.: for the Walker 256 carcinosarcoma at pH 5.0 it was 0 .152 mgP/ml.
Inhibition of acid phosphomonoesterase in the two tumor types by solutions of copper, fluoride, and L-tartrate was very marked and also comparable. The standard phosphate solution used as reference contained 0.1 mgP/mL. The net hydrolysis of the control solution for the Lutz hamster sarcoma was 0.122 mgP/ml. Inhibition with copper resulted in 0.0597 mgP/ml. or 51.1% inhibition; with fluoride, 0.0088 mgP/ml or 92.8% inhibition and with L-tartaric acid, 0.0174 mgP/ml or 85.8% inhibition.
Net hydrolysis of the control solution of the Walker 256 carcino-sarcoma was 0.126 mgP/ml. Inhibition with copper resulted in 0.0691 mgP/ml or 45.2% inhibition; with fluoride, 0.0022 mgP/ml or 93.6% inhibition and with L-tartaric acid, 0.0081 mgP/ml or 93.6% inhibition.
The degree of hydrolysis for the Lutz hamster sarcoma and for the Walker 256 carcinosarcoma enzyme homogenates was obtained at 30, 60, 90, 120, 150, and 180 minute intervals. A plot of hydrolysis versus time was linear in both cases. The activity in Klett units for both tumors was similar.
Since the literature contains little knowledge about blood acid phosphatase values in hamsters, the sera of 30 normal hamsters were analyzed using a semi-micro method. The substrate was p-nitrophenyl phosphate and activity was measured by the liberation of nitrophenol.
The acid phosphatase activity in the sera of the 30 animals tested ranged from 0.8420 to 1.0719 mM (Bessey-Lowry units) with a mean of 0.9126 mM. These results will have greater significance when the activity of acid phosphatase in the sera of tumor bearing hamsters is investigated.
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Thesis (M.A.)--Boston University
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