Ascorbate: Difference between revisions
From Bioblast
No edit summary |
No edit summary ย |
||
(38 intermediate revisions by 11 users not shown) | |||
Line 1: | Line 1: | ||
{{MitoPedia | {{MitoPedia | ||
|abbr=As | |abbr=As | ||
|description=Ascorbate | |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.ย 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=[[ | }} | ||
| | __TOC__ | ||
ย | |||
== Application in [[HRR]] == | |||
{{Chemical_description | |||
|abbr=As | |||
|trivial name=Ascorbate | |||
|complete name=(+)-Sodium L-ascorbate | |||
|chem formula=C<sub>6</sub>H<sub>7</sub>NaO<sub>6</sub> | |||
|molar mass=198.11 | |||
|vendor=Sigma-Aldrich | |||
|product number=A7631 | |||
|store at=RT | |||
|sensitivity= | |||
|cas=134-03-2 | |||
|h statements= | |||
|h info= | |||
}}<!--:::: '''As: Ascorbate sodium salt''' (Sodium L-ascorbate); Sigma A 7631, 100 g, store at RT; M = 198.1 gยทmol<sup>-1</sup>.--> | |||
:::: <span style="color:#8B008B"> '''Caution:''' Solutions of ascorbic acid are rapidly oxidized in air and in alkaline media; the compound should be protected from air and light. See 'ascorbid acid' on [https://pubchem.ncbi.nlm.nih.gov/compound/54670067 PubChem]</span> --> | |||
ย | |||
ย | |||
:::: '''Preparation of 800 mM stock solution''' (dissolved in H<sub>2</sub>O) for '''2-mL O2k-chamber''': | |||
ย | |||
::::# Weigh 792.4 mg of ascorbate sodium salt and dissolve in 3 mL H<sub>2</sub>O | |||
::::# To prevent autoxidation, adjust pH to approx. 6 with 1 M HCl (approx. 85 ยตL). | |||
::::# Adjust final volume to 5 mL and divide into 0.2 mL portions in dark vials. | |||
::::# Store frozen at -20 ยฐC. | |||
ย | |||
:::ยป'''O2k manual titrations:''' [[MiPNet09.12]] | |||
ย | |||
::::* Titration volume ('''2-mL O2k-chamber'''): 5 ยตL using a 25 ยตL Hamilton syringe. | |||
::::* Final concentration: 2 mM. | |||
ย | |||
ย | |||
:::: '''Preparation of 1000 mM stock solution''' (dissolved in H<sub>2</sub>O) for '''0.5-mL O2k-chamber''': | |||
ย | |||
::::# Weigh 990.5 mg of ascorbate sodium salt and dissolve in 3 mL H<sub>2</sub>O | |||
::::# To prevent autoxidation, adjust pH to approx. 6 with 1 M HCl. | |||
::::# Adjust final volume to 5 mL and divide into 0.2 mL portions in dark vials. | |||
::::# Store frozen at -20 ยฐC. | |||
ย | |||
:::ยป'''O2k manual titrations:''' [[MiPNet09.12]] | |||
ย | |||
::::* Titration volume ('''0.5-mL O2k-chamber'''): 1 ยตL using a 10 ยตL Hamilton syringe. | |||
::::* Final concentration: 2 mM. | |||
ย | |||
{{MitoPedia topics | |||
|mitopedia topic=Substrate and metabolite | |||
}} | }} | ||
Latest revision as of 09:20, 10 June 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 L-ascorbate; C6H7NaO6), Sigma-Aldrich: A7631, store at RT, CAS: 134-03-2, M = 198.11 gยทmol-1
- Caution: Solutions of ascorbic acid are rapidly oxidized in air and in alkaline media; the compound should be protected from air and light. See 'ascorbid acid' on PubChem -->
- Preparation of 800 mM stock solution (dissolved in H2O) for 2-mL O2k-chamber:
- Weigh 792.4 mg of ascorbate sodium salt and dissolve in 3 mL H2O
- To prevent autoxidation, adjust pH to approx. 6 with 1 M HCl (approx. 85 ยตL).
- Adjust final volume to 5 mL and divide into 0.2 mL portions in dark vials.
- Store frozen at -20 ยฐC.
- ยปO2k manual titrations: MiPNet09.12
- Titration volume (2-mL O2k-chamber): 5 ยตL using a 25 ยตL Hamilton syringe.
- Final concentration: 2 mM.
- Preparation of 1000 mM stock solution (dissolved in H2O) for 0.5-mL O2k-chamber:
- Weigh 990.5 mg of ascorbate sodium salt and dissolve in 3 mL H2O
- To prevent autoxidation, adjust pH to approx. 6 with 1 M HCl.
- Adjust final volume to 5 mL and divide into 0.2 mL portions in dark vials.
- Store frozen at -20 ยฐC.
- ยปO2k manual titrations: MiPNet09.12
- Titration volume (0.5-mL O2k-chamber): 1 ยตL using a 10 ยตL Hamilton syringe.
- Final concentration: 2 mM.
MitoPedia topics:
Substrate and metabolite