Riboflavin is one of several vitamins which may be repurposed as a viral inhibitor of SARS-CoV-2, the virus that causes Covid-19. So far, fifteen molecular docking studies have found that riboflavin and/or the related compounds FMN and FAD, are effective inhibitors of multiple viral targets. Flavin mononucleotide (FMN) called riboflavin-5′-phosphate, and Flavin Adenine Dinucleotide (FAD) are also discussed.
Riboflavin, Vitamin B2, Flavin mononucleotide (FMN), riboflavin-5′-phosphate, Flavitan, Flavin Adenine Dinucleotide (FAD), molecular docking, natural inhibitors, SARS-CoV-2, Covid-19.
The purpose of this article is to inform readers on a specific topic, and to encourage research, more generally, into Covid-19. Any discussion of a supplement or medication does NOT suggest or imply they should be used to prevent or treat Covid-19. The research on this topic is preliminary; clinical trials are needed to investigate these supplements and medications further. Please do NOT take any supplements or medications discussed or mentioned on this website, unless prescribed by your physician or other healthcare provider. If you have Covid-19 or any illness, see your healthcare providers and follow their advice. This website is not offer medical advice or treatments, and the information and discussions here are not a substitute for care by medical professionals.
What people call “the Coronavirus” is officially termed SARS-CoV-2, which is the virus that causes Covid-19. And SARS-CoV-1 (or simply SARS-CoV) is the virus that causes SARS.
A viral inhibitor is a compound which binds to a viral component in order to prevent that component from working. SARS-CoV-2, the virus that causes Covid-19, has Spike proteins all around its exterior. These Spikes must dock with a natural protein, called ACE2, found on the surface of cells in the lungs and other organs. An inhibitor could bind to the Spike, to prevent this docking of the Spike with the ACE2 — thereby preventing the infection of the cell by the virus. Or the inhibitor could bind with the ACE2, also preventing infection.
Inhibition is never 100% effective, so some cells will be infected. Other types of inhibitors can work inside an infected cell, to shut down the various processes the virus uses to replicate inside the cell. Ideally, a set of viral inhibitors would bind to numerous viral “targets”, and in this way reduce the amount of virus in the body. Inhibitors do not cure an illness. But they can slow down the viral infection, so that the immune system can overtake the disease and eventually clear the body of the virus.
Many of the drugs used to treat Covid-19 are inhibitors, including lopinavir, nelfinavir, azithromycin, and remdesivir. Hydroxychloroquine works in a few different ways, including by inhibition. An inhibitor can also be a natural compound. In this article, the evidence is reviewed that vitamin B2, also called riboflavin, may be a viral inhibitor of Covid-19. It is possible for a drug to have more than one mode of action, so that a compound which is a vitamin might also act as a natural inhibitor of a virus.
The studies we will review on riboflavin are all molecular docking studies. In this type of study, computer simulations examine the three-dimensional structure of a viral protein, paying particular attention to the active site on that protein — the part of the protein that provides its functionality. And then, by evaluating the shape and atomic structure of possible inhibitors, the software tests thousand, sometimes millions, of compounds for their binding affinity.
An effective inhibitor will have a high binding affinity; this is expressed as a negative number (termed a docking score). Inhibitors will tend to settle into a location which reduces the energy of the compound. So the lower the negative number, the higher the binding affinity, and the more effective the inhibitor will be in shutting down the viral protein. The typical range of docking scores in kcal/mol is -6.0 to -11.0 or lower. (Remember that in negative numbers -7 is lower than -6, and 8 is lower than -7, etc.) Any docking score of -6 or lower may possibly be effective against the virus, with the better scores being -7 or less.
Molecular docking studies can test vast numbers of compounds very quickly. But this is not the final word on whether a drug or natural compound will actually work against a disease. The information from molecular docking studies is very preliminary. Much more work remains to be done.
Linus Pauling Institute: “Riboflavin is a water-soluble B vitamin, also known as vitamin B2. In the body, riboflavin is primarily found as an integral component of the coenzymes, flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN).”  FMN is also called riboflavin-5′-phosphate.
As a water-soluble vitamin, B2 has a short half-life of only several hours. For supplementation then, the daily dose of riboflavin should be divided into multiple smaller doses spread throughout the day. In the cells and tissues of the body, riboflavin exists mainly as Flavin mononucleotide (FMN). So supplementation increases both riboflavin itself and FMN.
The recommended daily allowance for riboflavin for most adults is 1.3 mg/day; there is no tolerable upper limit as the amount of riboflavin in the human body seems to be self-limiting. As a water-soluble B-vitamin, riboflavin is eliminated from the body through the urine quite readily. In fact, the only common side effect of taking large doses of B2 is that the urine takes on a bright yellow color.
Doses as high as 400 mg/day of riboflavin are sometimes taken as a treatment for migraines. This dosage is safe for most health adults, with side effects being mild and low risk. As with many supplements, vitamin B2 may cause stomach irritation or diarrhea.
Riboflavin has been found to possibly protect against lung cancer in smokers as well as in non-smokers or former smokers [10, 11]. However, an in vitro study of a non‐small cell lung cancer cell line found that riboflavin, only at high doses, made existing lung cancer worse, increasing “growth, invasion, and migration” of the cancer cells . Therefore, certain persons should not take riboflavin supplements, especially at high doses.
The same study found that very high doses of riboflavin, in vitro, “significantly increased the release of interleukin‐6, tumor necrosis factor‐alpha, and vascular endothelial growth factor” . This increase in the pro-inflammatory cytokines IL-6 and TNF-a could conceivably promote or worsen a cytokine storm in severe cases of Covid-19. Vascular endothelial growth factor can make cancers of various kinds worse. So high doses of riboflavin should not be taken by anyone with cancer, or in remission from cancer, or at high risk of cancer. Also, high doses of riboflavin should not be taken by persons with severe Covid-19, due to the stimulation of pro-inflammatory cytokines IL-6 and TNF-a.
On the other hand, riboflavin has been shown to ameliorate acute lung injury and to reduce vascular leakage and lung hemorrhage . The study concluded that “riboflavin can mount a significant protection against oxidant-mediated inflammatory organ injury” . So, in this way, riboflavin might lessen the odds or the severity of a cytokine storm.
But why consider riboflavin as a possible inhibitor of Covid-19, at all, if there are downsides? All medications and supplements have possible detriments. Have you ever read the information packet that comes with medications prescribed by your doctor? There are many different possible side effects. But compared to the current medications used for Covid-19, and even compared to medications used for almost any other illness, riboflavin is low risk. Natural inhibitors in general are low risk. That is one reason they are being investigated to treat Covid-19.
Riboflavin could, in theory, make a cytokine storm worse. But as a viral inhibitor, riboflavin may also prevent Covid-19 from becoming severe enough that a cytokine storm would even be possible. Also, other supplements can be taken with riboflavin, supplements that downregulate IL-6 and TNF-a, counteracting that detriment of riboflavin. For example, while only a high dose of riboflavin increases pro-inflammatory cytokines (IL-6 and TNF-a), even a low dose of quercetin (another possible Covid-19 inhibitor) downregulates the release of TNF-a , and moderate doses of vitamin C and E downregulate IL-6 .
Riboflavin supplementation may make malaria worse, as a mild riboflavin deficiency may be protective against malaria . Therefore, persons with malaria should not take any supplement with riboflavin, not even at low doses. They should obtain their riboflavin from food only, unless otherwise directed by their doctor.
Riboflavin, FMN, and FAD
The human body converts much of the riboflavin from diet or supplements into flavin mononucleotide (FMN), also called riboflavin-5′-phosphate. Some riboflavin is also converted into flavin adenine dinucleotide (FAD). Fortunately, all three compounds have shown potential as viral inhibitors of SARS-CoV-2. Therefore, taking one OTC supplement of ordinary vitamin B2 (riboflavin) provides three different possible inhibitors of the virus that causes Covid-19. Now let’s review the studies on these compounds as inhibitors.
1) Alabboud 
The study by Alabboud and Javadmanesh reviewed 16 vitamins, along with many other compounds, as potential inhibitors of the main protease (Mpro, also called 3CLpro) of SARS-CoV-2 . The study was published in Dysona Life Science: DLS 1 (2020) 44-63 DOI: 10.30493/DLS.2020.225404
Several vitamins scored well enough to be possible inhibitors of SARS-CoV-2. The algorithm used to gauge the binding affinity is MolDock , which has values for effective inhibitors in the range of -100 to -150. The vitamins with the best scores, i.e. the best binding affinities, were B9 (Folic Acid), Reintol (vitamin A), gamma-tocopherol (a type of vitamin E), vitamin K1, Thiamine (B1), and Riboflavin (B2). Some other vitamins had mild binding affinities: B3, B5, B6, B7, a couple of forms of vitamin D, and Ascorbic acid (vitamin C) .
Riboflavin had the strongest hydrogen bond score (-14.73), along with some other advantages. Riboflavin is used in doses as high as 400 mg/day to treat migraines [19, 20]. There is no tolerable upper limit , and no known toxic dose . Side effects are mild and unusual, other than the effect of changing the color of the urine to a bright yellow . Riboflavin has a moderate solubility in water of 84.7 mcg/ml .
Folic acid scored better than riboflavin as an inhibitor of Mpro. However, it has a tolerable upper limit of only 1000 micrograms (1 mg) , which might not provide enough of a concentration to act as an inhibitor in vivo. Prescription viral inhibitors usually have doses in the range of 100 to 1,000 mg/day or more .
2) Choudhury 
This molecular docking study compared many natural compounds for their ability to inhibit PLpro (the lesser of the two proteases used by SARS-CoV-2), using the same software as in the previous study, Molegro Virtual Docker. Compared to Ascorbic acid, Glutathione, Aspartame, Chloroquine, Doxycycline, and Hydroxychloroquine, Riboflavin had the best reranking score (-100.85), but a mediocre score for hydrogen bonds (-6.83).
3) Garabato 
Riboflavin was compared, in this molecular docking study, to compounds from an FDA database of medications . Riboflavin compared favorably to lopinavir and nelfinavir, among other compounds, based on the estimated binding energy (kcal/mol) and the time (ns) at the binding site, which was Mpro. The study did not use the typical docking scores, nor the newer MolDock score, used by other studies.
4) Joshi 
These study authors “created a phytochemical library of 318 phytochemicals from 11 plants which have been reported as antiviral, antibacterial and antifungal activity.”  These compounds were screened for their ability to inhibit two targets: Mpro and ACE2. Riboflavin scored well against Mpro with a docking score of -7.6; the value for riboflavin against ACE2 was not reported.
5) Pendyala 
“In this study we used COVID-19 Docking Server to predict potential food bioactive compounds to inhibit Mpro and RdRp. The results showed that Phycocyanobilin, Riboflavin, Cyanidin, Daidzein, Genistein are potent inhibitor bioactive compounds to Mpro and RdRp in comparison to antiviral drugs.”  The SARS-CoV-2 viral protein RdRp is also called replicase. With three other proteins (Nsp7, 8, and 9), replicase makes copies of the viral RNA. Mpro is the main protease used by SARS-CoV-2 to make shorter functional proteins out of longer non-functional proteins. Here is a list of the compounds from this study, and the docking scores for inhibition of Mpro and RdRp .
Compound Mpro RdRp
Astaxanthin -7.00 -8.20
Catechin -7.30 -8.40
Curcumin -7.00 -8.10
Cyanidin -7.90 -8.80
Daidzein -7.80 -8.40
Genistein -7.60 -8.60
Phycocyanobilin -8.60 -9.30
Resveratrol -7.00 -7.30
Riboflavin -7.90 -9.00
Remdesivir -8.10 -9.00
Nelfinavir -7.90 -9.30
Lopinavir -7.90 -9.70
Astaxanthin is a carotenoid found in seafood and algae; it is available as a supplement. Catechin is found in Green tea extract. Curcumin is found in turmeric, and is also a supplement. Resveratrol is a supplement taken for good effects on the heart and on cholesterol levels. Daidzein and Genistein are found in Soy foods.
Phycocyanobilin (PCB) is a compound found in Spirulina. “A heaping tablespoon (about 15 g) of Spirulina can be expected to provide about 100 mg of PCB” . Spirulina is high in copper, so a zinc supplement would complement the Spirulina well. (Copper and zinc are antagonists; more of one leads to less of the other.) However, few studies have examined PCB in its possible effects on Covid-19. More research is needed on this compound.
Remdesivir, nelfinavir, and lopinavir are medications used to treat Covid-19.
Riboflavin is vitamin B2. Notice that riboflavin inhibits RdRp as well as Remdesivir, and almost as well as the other two Covid-19 drugs, nelfinavir and lopinavir. Then riboflavin’s scores for inhibition of Mpro are also competitive with the Covid-19 medications. The advantages offered by riboflavin are lower side effects, better safety profile, greater availability, and no prescription is required.
In addition, riboflavin can be taken safely on a daily basis by persons who are well, so that if they become ill, they will already be taking a viral inhibitor. The average length of time from infection to first symptoms is 5.1 days . If many persons were to take a set of supplements while well, they would in effect also be taking those supplements while they are sick and do not know it. And this might lessen the severity of Covid-19. However, these studies on riboflavin are very preliminary. More studies and especially clinical studies are needed before riboflavin can be recommended against Covid-19.
6) Verma 
The study title explains its research succinctly: “Potential inhibitors of SARS-CoV-2 Main protease (Mpro) identified from the library of FDA approved drugs using molecular docking studies”. These compounds were narrowed down to 51 top-scoring medications, including vitamins B2 and Pantethine, a derivative of vitamin B5. Both are also FDA-approved as medications.
“Considering bioavailability, lesser toxicity, route of administration some of the top-ranked drugs including … riboflavin (vitamin B2) and pantethine (vitamin B5) may be taken forward for further in vitro and in vivo experiments to investigate their therapeutic potential.” 
Riboflavin inhibited Mpro with a docking score of -7.219. That score is competitive with some Covid-19 drugs. Pantethine inhibited Mpro with a docking score of -6.365. That is a mediocre score, but pantethine has a good safety profile, and might be useful as part of a set of inhibitors.
7) Wu 
This study by 13 different authors examined a wide range of viral targets and natural as well as artificial compounds. Riboflavin was the 9th strongest inhibitor of PLpro from the ZINC drug database, behind doxycycline as the 6th and aspartame as the 4th . The study did not use common docking scores, but other data to rank the compounds. This study also found that ascorbic acid (vitamin C) and glutathione were inhibitors of PLpro, though with limited effectiveness.
8) Anwar 
The above seven studies found that riboflavin itself was an inhibitor of various viral targets. This study by Anwar et al. found that two compounds which the human body derives from riboflavin are also inhibitors:
(1) riboflavin 5’-monophosphate, also called flavin mononucleotide (FMN)
(2) flavin adenine dinucleotide (FAD), also called flavitan.
Compound, Target, Docking Score
Cholecalciferol (Vitamin D3), Membrane protein, -7.2
Flavin mononucleotide (FMN), PLpro, -7.0
Flavin adenine dinucleotide (FAD), Spike protein, -12.9
FAD, Nucleocapsid protein (N-protein), -13.3
FAD, Uridylate-specific Endoribonuclease (Nsp15), -12.3
FAD, Helicase (Nsp13), -11.2
FAD, RdRp (Nsp12), -12.8
FAD, Nsp2, -11.8
FAD, Guanine-N7 methyltransferase (Nsp16), -13.6
From the above data, it is clear that FAD is a strong inhibitor of multiple important targets of SARS-CoV-2. The N-protein wraps the viral RNA to protect it, hence its name, nucleocapsid protein. But also has a role in turning off the RNA silencing system naturally found in human cells, so that the virus can make viral proteins at full speed from its viral RNA. Inhibiting the N-protein is particularly important if we wish to use inhibitors to slow down the progress of Covid-19.
Helicase and replicase are the two most important polymerases of SARS-CoV-2. Inhibiting both of those proteins at such a high level (values like -11, -12, -13 all indicate very high binding affinity) is very beneficial. Then the Spike protein (or “surface glycoprotein” as the study terms it) is inhibited at a high level also, thereby reducing the infectivity of the virus.
It is interesting that vitamin D3 inhibits the membrane protein on the surface of SARS-CoV-2. Levels of D3 in the blood are in nanograms per ml, and are converted to other forms of vitamin D by the skin, liver, and kidney. So this inhibition is not particularly useful. But vitamin D has been shown by multiple studies to effectively limit the severity of Covid-19. It just does this by other means than inhibition.
The problem with FAD is that it is not used as a supplement. This means its safety profile is unknown, and approval of FAD as a medication for Covid-19 would be a long regulatory-fraught journey. Do not take FAD for Covid-19. Though it is a natural compound made by the body, we don’t know what the effects might be if it were taken as a supplement. The best way to obtain FAD in your body is to take Riboflavin, and allow the body to convert the riboflavin to FMN (mainly) and also FAD.
9) Mittal 
This study examined a wide range of natural and artificial compounds for their ability to inhibit the main protease of SARS-CoV-2 (Mpro).
Compound, Docking Score for Mpro
Flavitan -10.237 (FAD)
Baicalein -8.169 (not Baicalin)
Isorhamnetin -6.901 (a natural flavonoid)
Some of the highest scores were from natural inhibitors. Hesperidin is a strong inhibitor, and has been mentioned by over a dozen different molecular docking studies. The relationship to riboflavin is that “Flavitan” is actually FAD, a compound made in the body from riboflavin.
10) Shankar 
The unusual viral target considered in this study is non-structural protein 16 (Nsp16), which “performs the 2’-O-methyltransferase activity and puts a 5’ cap on the viral RNA molecules” . SARS-CoV-2 has 16 non-structural proteins, any of which might be targeted for inhibition. Nsp16 is necessary for the virus to make copies of itself, especially to make copies of its own RNA. Few studies have sought inhibitors for this target.
The study found that FAD, a compound related to Riboflavin, inhibits Nsp15 with a docking score of -9.05. By comparison, ritonavir’s score was a little better at -10.07, and the score for the natural carotenoid astaxanthin was a little worse at -8.63.
Using a different inhibitor scoring system, LibDock, the study also found effective natural inhibitors, each with a LibDock score of greater than 146. These scores are expressed as a positive number, but the idea is essentially the same, to find compounds with the strongest binding affinity to a viral target, so as to inactivate that component of the virus.
Compound LibDock score
Flavin Adenine Dinucleotide (FAD) 190.744
Folic Acid 152.859
Glutathione Disulfide 199.367
Phylloquinone (Vitamin K1) 151.703
Lutein is a carotenoid found in dark leafy green vegetables, and is important for good health. Phylloquinone is vitamin K1, also found in dark leafy greens. Hesperidin is a flavonoid from citrus, also available as a supplement. Folic acid is vitamin B9, which has a tolerable upper limit of 1 mg (1,000 mcg) per day; this limit makes folic acid less useful as an inhibitor. Diosmin is a flavonoid often taken with hesperidin for venous health. And FAD is made in the body from Riboflavin.
11) Bank 
This study examined compounds which may inhibit the Spike protein. They used by the open and closed configuration. A subunit of the Spike is capable of pivoting open, like a jaw, so as to bind to the ACE2 receptor and initiate infection of the cell. It is not clear how or when the protein subunit pivots in this manner. This might only occur as it approaches the ACE2 receptor.
Compound and Docking Score (kcal/mol)
Flavin Adenine Dinucleotide (FAD) -6.8
Flavin mononucleotide (FMN) -5.8
Phylloquinone (Vitamin K1) -5.0
The compound FMN is made by the body from riboflavin. Most riboflavin is turned into FMN by the body, so its activity as an inhibitor is important, if we want to consider using riboflavin as a viral inhibitor against Covid-19. FAD is less common, but also made by the body from riboflavin. Glutathione and Vitamin K1 were mild inhibitors of the Spike protein in this study.
12) Kumar 
This study, which focused mainly on folic acid, found that riboflavin and folic acid both inhibit Mpro with the same binding affinity of -7.7 kcal/mol.
13) Zhang 
This study found that riboflavin binds to RdRp (also called replicase) with a docking score of -7.1.
14) Kumar 
For the target PLpro, riboflavin was found to bind with a glide score of -8.11, while riboflavin-5′-phosphate bound with a better score of -9.91. For 3CLpro, riboflavin-5′-phosphate bound with a glide score of -8.05. This study also found targets for other vitamins. Folic acid had glide scores of -11.41 for PLpro, -8.00 for helicase, and -8.03 for Nsp10. Biotin had glide scores of -9.05 for RdRp and -8.54 for PLpro.
15) Prajapat 
Riboflavin was found to be one of the top inhibitors of the Spike protein – ACE2 interaction, blocking the connection between the two that leads to infection of the cell. “Conclusion: We identified 4 already approved drugs (riboflavin, fenoterol, cangrelor and vidarabine) as possible agents for repurposing as inhibitors of S1:ACE2 interaction.” 
The first seven studies above, plus the 12th, 13th, and 14th, found that riboflavin (vitamin B2) is an effective inhibitor of SARS-CoV-2, the virus that causes Covid-19. Then studies 8, 9, 10, and 11 found that FAD is an inhibitor of the same virus, while studies 8 and 11 also found that FMN is an inhibitor. The targets inhibited by these three related compounds include both viral proteases, the Spike protein, helicase, replicase, the N-protein, and Nsp15 and 16. That is a broad range of activity against this virus. And the binding affinity found by these studies ranged from moderate to very strong.
Taking a riboflavin supplement increases the riboflavin and the FMN in the body, and provides plenty of substrate to be converted into FAD. Riboflavin is an essential nutrient, which has proved to be safe in doses up to 400 mg/day. There is sufficient evidence, at this point, to justify in vitro studies and also clinical studies. Ordinarily, research would progress from molecular docking studies to in vitro studies to animal studies to clinical trials. But riboflavin is already FDA-approved, is available without a prescription, and has a known safety profile. Given the severity of the current pandemic and the need for effective treatments, clinical trials testing riboflavin as a prophylaxis or as part of a treatment program for mild to moderate Covid-19 are justifiable.
This article does not recommend riboflavin, nor any other supplement mentioned above, as a prevention or treatment for Covid-19. The purpose of the article is to inform the readership of this website, and to encourage further research into natural inhibitors. More research is needed on riboflavin before it can be recommended for any use beyond ordinary supplementation (the RDA of 1.1 and 1.3 mg for women and men, respectively).
If you are ill and may have Covid-19, consult your healthcare provider. This website does not offer medical advice or treatments.
Ronald L Conte Jr
Note: the author of this article is not a doctor, nurse, or healthcare provider.
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