Lab 7 - Bio. oxidation Flashcards Preview

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Flashcards in Lab 7 - Bio. oxidation Deck (15):

What are the examinations?

1.Demonstatrion of bio. oxidation by measuring oxygen consumption. We use 2,6-dichlorophenolindophenol.
2.Detection of cytochrome a+a3 in viro, by using p-phenylenediamine as H-donor.
3.Detection of cytochrome a+a3 in situ, by using Nadi´s reagent.
4.Detection of xanthine oxidase.


What are the methods to measure oxygen consumption of cells and tissues?

1.Manometric method
2.Electrodes showing the changes in the partial pressure of oxygen
3.Redox dyes and redox indicators, that has diff. colour in reduced and oxidized form.


How do we demonstrate the bio. oxidation by using 2,6-dichlorophenolindophenol?

It act´s as a redox dye, by accepting H from reduced flaboproteins (FMNH2, FADH2). In oxidized form it is blue, when reduced it is colourless.


What materials are used in "demonstatrion of bio. oxidation by measuring oxygen consumption"?

1.Heart muscle
2.Phosphate buffer
4.Glucose solution
5.Potassium succinate solution
6.Lactate solution
7.NAD+ in potassium phosphate buffer


What does the time of fading depen on, in "Demonstatrion of bio. oxidation by measuring oxygen consumption"?

How fast and how many reduced NAD+ and FAD are prod. during the oxidation, and how fast they can pass the H2 to the redox dye.


How is energy released in the resp.chain? Who are the H-donors?

By burning hydrogen into water. Reduces coenzymes and prosthetic groups (NADH+H+, FADH2) of dehydrogenases are H-donors.


How do we detect cytochrome a+a3 in vitro?

P-phenylenediamine acts as a H.donor and will be oxidized into violet coloured p-phenylenediimine.


What can inhibit the last enzyme of the resp. chain (cytochrome a+a3) ?



What are the materials in "detection of cytochrome a+a3 in vitro"?

3.Phosphate bugger
4.KCN sol.


Where are the mitochondria found in the n.c. in the brain?

In the cortex


How do we detect cytochrome a+a3 in situ?

Nadi´s reagent diffusing in brain slices reacts with cytochrome oxidase and changes into dark violet colour.


What are the materials in "detection of cytochrome a+a3 in situ"?

1.Pig brain
2.Nadi´s reagent (p-phenylenediamine and alpha-naphtol)
4.Phosphate buffer


What is xanthine oxidase?

Plays a role in degradation of purine bases. An enzyme, aerobe degrydrogenase, that can foward the H carried by it´s prostetic group (FADH2) directly to the molecular oxygen.


Which two reactions are catalysed by xanthine oxidase?

1.Hypoxanthine->xanthine->uric acid
2.Formaldehyde->aldehyde-hidrate-(methylene blue)->formic acid


How is xanthine oxidase indicated? What can inhibit the reaction?

Discolourization. Not KCN, because the enzyme can oxidise H without cytochrome too, but allopurinol.