Tyrosinase Composition

 Tyrosinase Article

Enzymatic Assay of TYROSINASE (EC 1 . 14. 18. 1) THEORY: L-Tyrosine + O2 L-DOPA Tyrosinase Tyrosinase

> L-DOPA

> L-DOPA-quinone + H20

Abbreviation utilized: L-DOPA sama dengan L-3, 4-Dihydroxyphenylalanine CONDITIONS: METHOD: REAGENTS: A. 50 logistik Potassium Phosphate Buffer, ph level 6. five at 25В° C (Prepare 50 cubic centimeters in deionized water using Potassium Phosphate, Monobasic, Desert, Sigma Push. No . P5379. Adjust to ph level 6. a few at 25В° C with 1 Meters KOH. ) 1 millimeter L-Tyrosine Option (Prepare 95 ml in deionized normal water using L-Tyrosine, Free Basic, Sigma Prod. No . T-3754. ) Tyrosinase Enzyme Solution (Immediately prior to use, prepare a solution that contains 500 - 1, 000 units/ml of Tyrosinase in cold Reagent A. ) T sama dengan 25В° C, pH sama dengan 6. five, A280nm, Light path = 1 centimeter

Continuous Spectrophotometric Rate Dedication

B.

C.

PROCEDURE: Make a reaction tropical drink by pipetting (in milliliters) the following reactants into a appropriate container: Deionized Water Reagent A (Buffer) Reagent B (Tyrosine) on the lookout for. 00 twelve. 00 12. 00

SPTYRO01. 001 Revised: 02/22/94

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Enzymatic Assay of TYROSINASE (EC 1 ) 14. 18. 1) METHOD: (continued) Mix and conform to pH six. 5 at 25В° C with 1 M HCl or one particular M NaOH, if necessary. Immediately before use, oxygenate by simply bubbling 99. 9% real O2 throughout the reaction beverage for 3-5 minutes. Pipette (in milliliters) into appropriate quartz cuvettes: 1 Evaluation Blank Response Cocktail 2 . 90 installment payments on your 90

Equilibrate to 25В° C. Keep an eye on the A280nm until regular, using a very well thermostatted spectrophotometer. Then put: Reagent A (Buffer) Reagent C (Enzyme Solution) -----0. 10 0. 10 ------

Immediately combine by inversion and record the increase in A280nm for about 10 minutes. Obtain the r A280nm/minute using the maximum linear charge for both the Ensure that you Blank. COMPUTATIONS: Units/ml enzyme = (0. 001) (0. 1) df = Dilution factor zero. 001 sama dengan The change in A280nm/minute per unit of Tyrosinase at pH 6. 5 in 25В° C in a a few ml response mix 0. 1 = Volume (in milliliters) of enzyme utilized units/ml enzyme Units/mg...

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