Malate: Difference between revisions
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'''M: Malate''' (L-Malic acid, C<sub>4</sub>H<sub>6</sub>0<sub>5</sub>); Sigma M 1000, 100 g, store at RT; FW = 134.1. | '''M: Malate''' (L-Malic acid, C<sub>4</sub>H<sub>6</sub>0<sub>5</sub>); Sigma M 1000, 100 g, store at RT; FW = 134.1. | ||
'''Preparation of 400 | '''Preparation of 800 (or 400) mM stock solution''' (dissolved in H<sub>2</sub>O): | ||
::1) Weigh 268.2 | ::1) Weigh 536.4 (or 268.2) mg of L-malic acid. | ||
::2) Add 3 ml H<sub>2</sub>O. | ::2) Add 3 ml H<sub>2</sub>O. | ||
::3) Neutralize with 10 N KOH (approx. 350 µl). | ::3) Neutralize with 10 N KOH (approx. 350 µl). | ||
| Line 25: | Line 25: | ||
::6) Store frozen at -20 °C. | ::6) Store frozen at -20 °C. | ||
Comment: 800 mM stock until 2013-11-20. | Comment: 800 mM stock until 2013-11-20 (for 2 mM final concentration); 400 mM stock (for 0.5 mM final concentration) in discussion since then. | ||
'''Oxygraph-2k manual titrations''' >> [[MiPNet09.12 O2k-Titrations]] | '''Oxygraph-2k manual titrations''' >> [[MiPNet09.12 O2k-Titrations]] | ||
::* Titration volume: | ::* Titration volume: 5 µl 800 mM (or 2.5 µl 400 mM) using a 25 µl Hamilton syringe (2 ml O2k-chamber). | ||
::* Final concenteration: 0.5 | ::* Final concenteration: 2.0 mM (or 0.5 mM). | ||
[[Image:BB-Bioblast.jpg|left|30px|link=Bioblast:About|Bioblast wiki]] | [[Image:BB-Bioblast.jpg|left|30px|link=Bioblast:About|Bioblast wiki]] | ||
Revision as of 20:32, 5 March 2015
- high-resolution terminology - matching measurements at high-resolution
Malate
Description
Malic acid, C4H6O5, occurs under physiological conditions as the anion malate2-, M, with pKa1 = 3.40 and pKa2 = 5.20. L-Malate is formed from fumarate in the TCA cycle in the mitochondrial matrix, where it is the substrate of malate dehydrogenase oxidized to oxaloacetate. Malate is also formed in the cytosol. It cannot permeate through the lipid bilayer of membranes and hence requires a carrier (dicarboxylate carrier, tricarboxylate carrier and 2-oxoglutarate carrier). Malate alone cannot support respiration of mt-preparations from most tissues, since oxaloacetate accumulates in the absence of pyruvate or glutamate.
Abbreviation: M
Reference: Gnaiger 2014 MitoPathways
MitoPedia topics: Substrate and metabolite
Application in HRR
M: Malate (L-Malic acid, C4H605); Sigma M 1000, 100 g, store at RT; FW = 134.1.
Preparation of 800 (or 400) mM stock solution (dissolved in H2O):
- 1) Weigh 536.4 (or 268.2) mg of L-malic acid.
- 2) Add 3 ml H2O.
- 3) Neutralize with 10 N KOH (approx. 350 µl).
- 4) Adjust final volume to 5 ml (in 5 ml volumetric glass flask).
- 5) Divide into 0.5 ml portions.
- 6) Store frozen at -20 °C.
Comment: 800 mM stock until 2013-11-20 (for 2 mM final concentration); 400 mM stock (for 0.5 mM final concentration) in discussion since then.
Oxygraph-2k manual titrations >> MiPNet09.12 O2k-Titrations
- Titration volume: 5 µl 800 mM (or 2.5 µl 400 mM) using a 25 µl Hamilton syringe (2 ml O2k-chamber).
- Final concenteration: 2.0 mM (or 0.5 mM).
Malate: 0.5 mM versus 2 mM in HRR
In mitochondrial preparations obtained from a diversity of tissues and organisms malate at concentrations >0.5 mM exerted an inhibitory effect on CII-linked and CI&II-linked respiration, whereas 0.5 mM malate was saturating for CI linked respiration. Selection of an optimum malate concentration for SUIT protocols is a compromise. Current investigations point towards 2 mM malate, but a final result is not yet available.
- Further details: »Discussion.
