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Difference between revisions of "Rotenone"

From Bioblast
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:::: '''Preparation of 1 mM stock solution''' (dissolved in 99.9% ethanol)
:::: '''Preparation of 1 mM stock solution''' (dissolved in 99.9% ethanol) for '''2-mL O2k-chamber''':


::::# Weigh 0.39 mg of rotenone and dissolve in 1 ml of ethanol absolute (99.9%).
::::# Weigh 0.39 mg of rotenone and dissolve in 1 ml of ethanol absolute (99.9%).
::::# Divide into 100 µl aliquots (use dark glass vials).  
::::# Divide into 100 µL aliquots (use dark glass vials).  
::::# Store frozen at -20 °C.
::::# Store frozen at -20 °C.
:::» '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]
::::* Titration volume ('''2-mL O2k-chamber'''): 1 µL using a 10 µL Hamilton syringe.
::::* Final concentration: 0.5 µM.


:::: '''Preparation of 0.25 mM solution''' (dissolved in 99.9% ethanol) for '''0.5-mL O2k-chamber''':
::::# Take 50 µL of the 1 mM stock solution
::::# Add 150 µL of ethanol absolute (99.9%).


:::» '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]
:::» '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]


::::* Titration volume: 1 µl using a 10 µl syringe (2 ml O2k-Chamber).
::::* Titration volume ('''0.5-mL O2k-chamber'''): 1 µL using a 10 µL Hamilton syringe.
::::* Final concentration: 0.5 µM.
::::* Final concentration: 0.5 µM.




:::'''Microalgae applications'''
:::'''Microalgae applications'''
::: For the specific case of microalgal cells it is recommended to prepare higher concentration:
::: For the specific case of microalgal cells it is recommended to prepare higher concentration:
:::: '''Preparation of 5.5 mM stock solution''' (dissolved in 99.9% ethanol)
:::: '''Preparation of 5.5 mM stock solution''' (dissolved in 99.9% ethanol)


Line 33: Line 45:
::::# Divide into 200 µl aliquots (use dark glass vials).  
::::# Divide into 200 µl aliquots (use dark glass vials).  
::::# Store frozen at -20 °C.
::::# Store frozen at -20 °C.


:::» '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]
:::» '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]


::::* Titration volume: 15 µL using a 25 µL syringe (2 mL O2k-Chamber).
<!--::::* Titration volume: 15 µL using a 25 µL syringe (2 mL O2k-Chamber).-->
::::* Titration volume ('''2-mL O2k-chamber'''): 15 µL using a 25 µL Hamilton syringe.
::::* Titration volume ('''0.5-mL O2k-chamber'''): 3,75 µL using a 10 µL Hamilton syringe.
::::* Final concentration: 41 µM.
::::* Final concentration: 41 µM.


== Inhibition of CI by rotenone ==
== Inhibition of CI by rotenone ==

Revision as of 11:24, 22 October 2020


high-resolution terminology - matching measurements at high-resolution


Rotenone

Description

Rotenone is an inhibitor of Complex I (CI) and thus inhibits NADH oxidation. It inhibits the transfer of electrons from iron-sulfur clusters in CI to ubiquinone via binding to the ubiquinone binding site of CI. See also Complex II-linked substrate state, S(Rot).

Abbreviation: Rot

Reference: Gnaiger 2014 MitoPathways, MiPNet09.12, MiPNet03.02

Application in HRR

Rot: Rotenone (C23H22O6); Sigma R 8875, 1 g, store commercial stock at RT; FW = 394.42 (≥ 95%).
Caution: Very toxic and light sensitive (store solution in a dark glass vial).


Preparation of 1 mM stock solution (dissolved in 99.9% ethanol) for 2-mL O2k-chamber:
  1. Weigh 0.39 mg of rotenone and dissolve in 1 ml of ethanol absolute (99.9%).
  2. Divide into 100 µL aliquots (use dark glass vials).
  3. Store frozen at -20 °C.
» O2k manual titrations MiPNet09.12 O2k-Titrations
  • Titration volume (2-mL O2k-chamber): 1 µL using a 10 µL Hamilton syringe.
  • Final concentration: 0.5 µM.


Preparation of 0.25 mM solution (dissolved in 99.9% ethanol) for 0.5-mL O2k-chamber:
  1. Take 50 µL of the 1 mM stock solution
  2. Add 150 µL of ethanol absolute (99.9%).
» O2k manual titrations MiPNet09.12 O2k-Titrations
  • Titration volume (0.5-mL O2k-chamber): 1 µL using a 10 µL Hamilton syringe.
  • Final concentration: 0.5 µM.


Microalgae applications
For the specific case of microalgal cells it is recommended to prepare higher concentration:
Preparation of 5.5 mM stock solution (dissolved in 99.9% ethanol)
  1. Weigh 4.4 mg of rotenone and dissolve in 2 ml of ethanol absolute (99.9%).
  2. Divide into 200 µl aliquots (use dark glass vials).
  3. Store frozen at -20 °C.
» O2k manual titrations MiPNet09.12 O2k-Titrations
  • Titration volume (2-mL O2k-chamber): 15 µL using a 25 µL Hamilton syringe.
  • Titration volume (0.5-mL O2k-chamber): 3,75 µL using a 10 µL Hamilton syringe.
  • Final concentration: 41 µM.


Inhibition of CI by rotenone

  • Fato R, Bergamini C, Bortolus M, Maniero AL, Leoni S, Ohnishi T, Lenaz G (2009) Differential effects of mitochondrial Complex I inhibitors on production of reactive oxygen species. Biochim Biophys Acta 1787:384–92. >>PMC2724837






MitoPedia topics: Inhibitor