Difference between revisions of "Ascorbate"
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{{MitoPedia | {{MitoPedia | ||
|abbr=As | |abbr=As | ||
|description=In respiratory assays for cytochrome ''c'' oxidase activity ([[Complex IV|Complex IV, CIV]]), '''ascorbate''' is added as regenerating system | |description=In respiratory assays for cytochrome ''c'' oxidase activity ([[Complex IV|Complex IV, CIV]]), '''ascorbate''' is added as regenerating system to maintain [[TMPD]] in a reduced state. It has to be titrated into the respiration medium prior to the addition of TMPD, otherwise the [[autoxidation]] reaction velocity is permanently elevated. | ||
|info=[[MiPNet06.06]] | |info=[[MiPNet06.06]] | ||
}} | }} |
Revision as of 13:06, 5 March 2021
Description
In respiratory assays for cytochrome c oxidase activity (Complex IV, CIV), ascorbate is added as regenerating system to maintain TMPD in a reduced state. It has to be titrated into the respiration medium prior to the addition of TMPD, otherwise the autoxidation reaction velocity is permanently elevated.
Abbreviation: As
Reference: MiPNet06.06
Application in HRR
- As: Ascorbate sodium salt (Sodium L-ascorbate); Sigma A 7631, 100 g, store at RT; M = 198.1 g·mol-1.
- Caution: Light sensitive (store solution in a dark glass vial).
- Preparation of 800 mM stock solution (dissolved in H2O) for 2-mL O2k-chamber:
- Weigh 1.584 g of ascorbate sodium salt and dissolve in 10 mL H2O
- To prevent autoxidation, adjust pH to approx. 6 with ascorbic acid (a 137.6 mg.mL-1 solution of pH 2).
- Divide into 0.2 mL portions.
- Store frozen at -20 °C.
- »O2k manual titrations: MiPNet09.12
- Titration volume (2-mL O2k-chamber): 5 µL using a 25 µL Hamilton syringe.
- Final concentration: 2 mM.
- Preparation of 1000 mM stock solution (dissolved in H2O) for 0.5-mL O2k-chamber:
- Weigh 1.980 g of ascorbate sodium salt and dissolve in 10 mL H2O
- To prevent autoxidation, adjust pH to approx. 6 with ascorbic acid (a 137.6 mg.mL-1 solution of pH 2).
- Divide into 0.2 mL portions.
- Store frozen at -20 °C.
- »O2k manual titrations: MiPNet09.12
- Titration volume (0.5-mL O2k-chamber): 1 µL using a 10 µL Hamilton syringe.
- Final concentration: 2 mM.
MitoPedia topics:
Substrate and metabolite