Difference between revisions of "Malate"
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|description=[[File:Malic_acid.jpg|left|100px|Malic acid]] | |description=[[File:Malic_acid.jpg|left|100px|Malic acid]] | ||
'''Malic acid''', C<sub>4</sub>H<sub>6</sub>O<sub>5</sub>, occurs under physiological conditions as the anion '''malate<sup>2-</sup>, M''', with p''K''<sub>a1</sub> = 3.40 and p''K''<sub>a2</sub> = 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 [[Mitochondrial preparations|mt-preparations]] from most tissues, since oxaloacetate accumulates in the absence of [[pyruvate]] or [[glutamate]]. | '''Malic acid''', C<sub>4</sub>H<sub>6</sub>O<sub>5</sub>, occurs under physiological conditions as the anion '''malate<sup>2-</sup>, M''', with p''K''<sub>a1</sub> = 3.40 and p''K''<sub>a2</sub> = 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 [[Mitochondrial preparations|mt-preparations]] from most tissues, since oxaloacetate accumulates in the absence of [[pyruvate]] or [[glutamate]]. | ||
Malate is a [[NADH Electron transfer-pathway state |type N substrate]] (N) required for the [[Fatty acid oxidation pathway control state| FAO-pathway]]. In the presence of [[Malate anaplerotic pathway control state|anaplerotic pathways]] (''e.g.'', [[Malic enzyme|mitochondrial malic enzyme, mtME]]) FAO-pathway could be overestimated due to a contribution of NADH-linked respiration, F(N) (see [[SUIT-002]]). | |||
|info=[[Gnaiger 2014 MitoPathways]] | |info=[[Gnaiger 2014 MitoPathways]] | ||
|type=Respiration | |type=Respiration | ||
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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 mM | '''Preparation of 400 mM stock solution''' (dissolved in H<sub>2</sub>O): | ||
::1) Weigh 268.2 mg of L-malic acid. | ::1) Weigh 268.2 mg of L-malic acid. | ||
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::* Titration volume: 10 µL 400 mM using a 25 µL Hamilton syringe (2 mL O2k-Chamber). | ::* Titration volume: 10 µL 400 mM using a 25 µL Hamilton syringe (2 mL O2k-Chamber). | ||
::* Final | ::* Final concentration: 2.0 mM. | ||
'''Preparation of 50 mM solution''' | '''Preparation of 50 mM solution''' | ||
::1) dilute 0.625ml of 400 mM stock solution with 4.375 ml of H<sub>2</sub>O ( final volume | ::1) dilute 0.625ml of 400 mM stock solution with 4.375 ml of H<sub>2</sub>O (final volume 5 mL) | ||
::2) Divide into 0.250 mL portions. | ::2) Divide into 0.250 mL portions. | ||
::3) Store frozen at -20 °C. | ::3) Store frozen at -20 °C. | ||
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::* Titration volume: 4 µL 50 mM using a 10 µL Hamilton syringe (2 mL O2k-Chamber). | ::* Titration volume: 4 µL 50 mM using a 10 µL Hamilton syringe (2 mL O2k-Chamber). | ||
::* Final | ::* Final concentration: 0.1 mM. | ||
Revision as of 09:46, 12 March 2019
- 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. Malate is a type N substrate (N) required for the FAO-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) FAO-pathway could be overestimated due to a contribution of NADH-linked respiration, F(N) (see SUIT-002).
Abbreviation: M
Reference: Gnaiger 2014 MitoPathways
MitoPedia topics: Substrate and metabolite
Application in HRFR
M: Malate (L-Malic acid, C4H605); Sigma M 1000, 100 g, store at RT; FW = 134.1.
Preparation of 400 mM stock solution (dissolved in H2O):
- 1) Weigh 268.2 mg of L-malic acid.
- 2) Add 3 mL H2O.
- 3) Neutralize with 5 M 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.
O2k manual titrations >> MiPNet09.12 O2k-Titrations
- Titration volume: 10 µL 400 mM using a 25 µL Hamilton syringe (2 mL O2k-Chamber).
- Final concentration: 2.0 mM.
Preparation of 50 mM solution
- 1) dilute 0.625ml of 400 mM stock solution with 4.375 ml of H2O (final volume 5 mL)
- 2) Divide into 0.250 mL portions.
- 3) Store frozen at -20 °C.
O2k manual titrations
- Titration volume: 4 µL 50 mM using a 10 µL Hamilton syringe (2 mL O2k-Chamber).
- Final concentration: 0.1 mM.
Malate: 0.5 mM versus 2 mM in HRFR
In mitochondrial preparations obtained from a diversity of tissues and organisms malate at concentrations >0.5 mM exerted an inhibitory effect on succinate-pathway (S-pathway) and NADH and S (NS-)pathway, whereas 0.5 mM malate was saturating for NADH (N-) linked respiration. Selection of an optimum malate concentration for SUIT protocols is a compromise. Current investigations point towards 2 mM malate. Sumbalova_2016a_Abstract_MitoFit_Science_Camp_2016
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.
- Further details: »Discussion.
