I don't know if you can buy pure FADH2 but you can purchase FADH2-disodium salt hydrate from Sigma: http://www.sigmaaldrich.com/catalog/product/sial/46360

FADH2 is produced when complex 2 of the electron transport chain breaks down succinate and 2 hybrides and 2 electrons are transferred to FAD.

Hi Martin

I understand that  you want to isolate it from a natural source (?)

Remember that FAD/FADH2 is mainly a prosthetic group. I wonder whether would be possible to isolate FADH2 outside a living cell or a functional mitochondria, or if it in the process of isolation all FADH2 is converted into FAD.

Are you also asking for a company selling FADH? Perhaps you need FAD to enter into the protein structure and then use a protocol to convert it into FADH2.

FADH2 is probably too readily oxidized in air to be stable. Perhaps you can prepare it in situ from FAD by treating with sodium dithionite.

Dear Martin, I do not remeber having used FADH2 ever. I assume you may be preparing an enzymatic assay that requires it in saturating conditions. This is very difficult. I would propose to couple your assay to an upstream NAD/NADH source or try a classicañl cycling approach regenerating repeatidly FADH2 from FAD (commercial).

NADH is surely available from Oriental Yeast Co., Tokyo, Japan.   FADH may be produced as suggested by Dr. Sebastián Cerdán.   In place of NADH, ascorbate-PMS may be used (see file;  Alanine transport, please rotate the attached article counter-clock wise by 90 degrees). 

Please, see the URL

http://www.oyc-bio.jp/pages/info/english

Further, dithonite (sodium hydrosulfite) is too strong reducing reagent to use in biochemical study.   Thus, I have used it to reduce purified cytochrome c (from Candida krusei; please see file Red-cyt c Hg).  

Dear

Flavin adenine dinucleotide (FAD) is a redox cofactor, more specifically a prosthetic group, involved in several important reactions in metabolism. FAD, in its fully oxidized form, or quinone form, accepts two electrons and two protons to become FADH2 (hydroquinone form). The semiquinone (FADH·) can be formed by either reduction of FAD or oxidation of FADH2 by accepting or donating one electron and one proton, respectively.

Please to have information according order of  FADH2 check the following link: https://pubchem.ncbi.nlm.nih.gov/compound/446013

Thank

Marius

Thanks Daniel, but I think that the form of SIGMA's reference is oxidied. See, the structure of the reduced form in Wikipedia:

https://en.wikipedia.org/wiki/Flavin_adenine_dinucleotide

thanks, Marius, we need in theory the purified product

Thank you Rafael, Sebastian and Kou. Yes, I could couple FAD to any reaction requiring this cofactor, but in theory I will need to use later purify FADH2. Hence I am not sure if I could couple it to any reaction. I have to think longer about it.

thank you Adam for the idea of using sodium dithionite. Do you know if it will originate only FADH2? I am afraid that using a reducing salt as that one could in fact reduces the whole molecule and not only the 2 nitrogen atoms.

I think that obtaining FADH2 is impossible if not inside a protein forming a flavoprotein that can be reduced. In this case we need to figure out another approach for the experiment we were planning.

All the best and I will be happy if you have additional comments.

Hi Martin

I still wonder about the need to purify FADH2  as it will be oxidized quite easily.

Anyways if you need to purify FADH2 the first step, as mentioned in the earlier answers, would be the conversion of FAD to FADH2 adding the co-substrate of FAD in your specific system, and an appropraite enzyme and substrate to regenerate NADH  and keep it always in excess. Subsequently, during purification you may need to keep strong reducing conditions (apart from the NADH regenerating system) during the purification. And at the end you would need to keep it free from oxygen, i.e. in nitrogen atmosphere. A real challenge!.

Hi Rafael,

we wanted FADH2 for mass spectometry to be sure that we correctly distingush the peaks of FAD and FADH2. But I agree that it is going to be very difficult for this reaon we are looking for alternatives ways.

Thank you for your help.

Amended on 08 November 2016

Immortal fetal-normal Hc (derived from fetal livers of several Americans) has three flavoproteins of Neutrophil NADPH oxidase factor 4 /Neutrophil cytosol factor 4 at 14.6 μg/mg of cell protein, Metalloreductase STEAP2/Protein up-regulated in metastatic prostate cancer at 5.4 μg/mg of cell protein,  and Prenylcysteine oxidase 1 at 3.9 μg/mg of cell protein.   Flavoproteins becomes 23.9  μg/mg of cell protein or 2.4% among cell proteins.   Neutrophil cytosol factor 4 binds FAD loosely, and detaches easily by non-ionic detergent.

Immortal healed HepG2 (with fucoidan; three days at 0.104 mg/mL) has one flavoprotein of Acyl-coenzyme A oxidase 2, peroxisomal at 1.3 μg/mg of cell protein.

Interestingly, immortal cancer HepG2 (without fucoidan) has no flavoprotein at all. Although, it has L-Lactate dehydrogenase A-like 6A at 2.5 μg/mg of cell protein and Alcohol dehydrogenase 6 at 0.64 μg/mg of cell protein, but these are NAD-linked fermentation enzymes.　　　

Therefore, possible preparation of FADH should be performed from fetal Hc cells and/or ES cells, which get the energy from oxidative respiration.

By the way, I have come to an important conclusion about the additive of dithionite due thanks to Dr. Martin Villalba (French Institute of Health and Medical Research, Paris, France).    The dangerous dithionite should not be added to the Wine, since dithionite helps growth of cancer cells via inhibition of oxidative respiration in surrounding normal non-cancer cells.   Therefore, I like "Sake (Japanese rice wine)"  very much than "Wine".   

Thanks Joerg, we need to figure out if we perform the experiments measuring FADH2 in intact cells instead that using extracts to avoid oxidation. Thnaks also for the papers.

Thnak you Kou. I was not aware of these differences. We should carefully choose the cell lines to be used.

best