Why The World’s First Drugs Ate Your Butt Cheeks and Turned You Blue

💉 Whether we like it or not, drugs are a fundamental part of our modern world. But what were “early drugs” like, back in the day? At some point, we clearly moved from drilling holes in people’s skulls, to using synthetic molecules as medicines. In this video, we will walk through the history, science and chemistry of some of the early advances.
If you want to fully appreciate the content of upcoming videos on (shocking) history of cancer treatments and modern chemotherapies, you should pay attention!

00:00 Early drugs were CRAZY
00:36 Dubious ancient methods: Mummies?!
01:43 Paul Ehrlich: Stains paving the way of discovery
02:59 Advent of chemotherapy and magic bullets: Methylene blue
03:36 Synthesis of methylene blue
06:45 Malaria in a nutshell, and use of methylene blue
08:29 Mechanism of action of methylene blue
10:27 Provay Blue for treatment of methemoglobinemia
11:33 Methylene blue for treatment of malaria today
12:45 Ehrlich’s work in immunology and syphilis
14:07 Mercury and arsenic as medicine?!
16:10 The birth of medicinal chemistry: Salvarsan/ arsphenamine
17:25 Translation of Salvarsan from clinic to market
18:32 Necrosis and arsenic deposits in butt cheeks
19:15 Chemistry insight #1: Neosalvarsan
19:54 Chemistry insight #2: Synthesis of salvarsan and impurities
21:21 Chemistry insight #3: Salvarsan structure
22:46 Conclusions and THANKS!

https://www.patreon.com/totalsynthesis
🚀 Thanks to all channel supporters!!!
https://www.instagram.com/totalsynthesis_official/
👉 Follow me on IG to learn about interesting news and mechanism quizzes!
https://www.total-synthesis.com

Disclaimer – This channel does not provide medical advice!
No information on this channel is intended to be a substitute for professional medical advice, diagnosis or treatment. Always seek the advice of your physician or other qualified health care provider with any questions you may have regarding a medical condition or treatment and before undertaking a new health care regimen, and never disregard professional medical advice or delay in seeking it because of information on Youtube.

Key references:
– ISIS. 1985, 76, 319: Ancient and medieval chemotherapy for cancer
– Beiträge zur Theorie und Praxis der histologischen Färbung; P. Ehrlich, Leipzig, 1878 (https://www.pei.de/DE/institut/paul-ehrlich/publikationen-von-paul-ehrlich/publikationen-von-paul-ehrlich-node.html)
– Biologisches Zentralblatt 1887, 6, 214 | Ueber die Methylenblaureaktion der lebenden Nervensubstanz
– Berlin Klin Woch 1891, 28, 953 | Ueber die Wirkung des Methylenblau bei Malaria.
– Malaria Journal 2016, 15, 51 | Comparative genome-wide analysis and evolutionary history of haemoglobin-processing and haem detoxification enzymes in malarial parasites
– Lancet Infectious Diseases 2018, 18, 627 | Efficacy and safety of primaquine and methylene blue for prevention of Plasmodium falciparum transmission in Mali: a phase 2, single-blind, randomised controlled trial
– Berlin Klin Woch 1899, 36, 481
– Proc. R. Soc. Lond. B. 1905, 76, 589 | The experimental treatment of trypanosomiasis in animals
– Annales de chimie et de physique 1854, 42,‎ 186 | Sur la nitronaphtaline et la nitrobenzine
– Bull. Hist. Chem. 1999, 23, 28 | Places and chemistry: Strasbourg – A chemical crucible seen through historical personalities
– Wechselmann: The Treatment of Syphilis with Salvarsan (1911), New York: Rebman, 11
– J r Soc Med 2009, 102, 343 | The introduction of ‘chemotherapy’ using arsphenamine – the first magic bullet
– Ber. Dtsch. Chem. Ges. 1912, 45, 756 | Über das salzsaure 3.3′-Diamino-4.4′-dioxy-arsenobenzol und seine nächsten Verwandten
– JACS 1920, 42, 2402 | HYPOPHOSPHOROUS ACID PREPARATION OF ARSPHENAMINE. (3,3-DIAMINO-4,4-DIHYDROXY-ARSENO-BENZENE DIHYDROCHLORIDE)
– ACIE 2005, 44, 829 | The composition of Ehrlich’s salvarsan: resolution of a century-old debate

Did you know that 1 in 3 people will develop cancer throughout their life? As we get older, many of us will rely on modern medicines like chemotherapy. The early days of drug discovery saw patients getting super-sized injections of drugs with cursed structures, with unabsorbed arsenic often living rent-free in their butt cheeks, eating away at their tissues for weeks after treatment. How about using the color blue as a medicine? If you think this is a joke or an obsolete practice, you are in for a surprise. Let s check out some nice history and science. For most of human history, many diseases, including cancer, were treated with dubious methods like bloodletting, or random herb extracts. Look at this: Ancient Persian medicine utilized asphalt residue as a remedy for wounds. Later, medieval Latin translators misinterpreted the term mumiya to mean literal mummies. This sparked century-long trade and medicinal use of ground mummy powder. It was only a matter of time until sneaky apothecaries started creating fake mummies by drying random corpses in the sun or in ovens. This is some next level stuff. Some ancient methods might have been quite legit though. For instance, already two thousand years ago, the Greek scientist Dioscorides used wine extracts of the autumn crocus plant to treat tumors. The synthesis enthusiasts listening will know that this plant contains the natural product colchicine, which, as was discovered in 1938, does have anti-cancer activity. Nevertheless, most treatments were indeed dubious but things started to change with the German scientist Paul Ehrlich in the late 19th century. Ehrlich studied medicine, and earned his doctorate for his work on histological staining of tissues. You might have done this yourself during your studies. Very unusual for a dissertation in medicine and this tells you about scientific character he went deep into the chemistry of the early aniline stains. Funnily enough, Ehrlich s cousin Karl Weigert was the first person to stain bacteria, and introduce aniline dyes. These guys surely had some special scientist DNA: During his studies, his aniline stains highlighted a specific granulate in cells that he initially believed to be known plasma cells. I m not showing you all the details of the hard-to-translate texts, but after more investigations he realized that he was looking at a completely new cell type. Due to their large granules, he believed that these calls nourished the surrounding tissues, dubbing them mast cells. This is incorrect, as we know today that this type of white blood cell is part of the immune system, and the granules contain different chemical mediators instead of nutrients. Well, even legends can have wrong hypotheses. What does have to do with chemotherapy? Well, another insight by Ehrlich was in vivo staining in this case, nerve cells of frogs with the dye methylene blue. This confirmed that chemicals could interact with specific parts of cells in living organisms as well. You can see that he was eager to advance the field, wondering about the mechanisms of these differential interactions. After more research, he postulated that chemicals and antibodies can selectively target pathogens or disease, rather than healthy cells. He later coined the term of magic bullets which is he quite famous for. Given we will talk about methylene blue, we should check out its synthesis. This was one of the first key products of the emerging dye industry, synthesized by the German Chemist Caro at BASF. This is still the largest chemical producer in the world. There would be quite a lot of history to unpack, but let s just check out the original synthesis in 1876. From dimethyl-aniline, it s just 3 steps so it should be an easy task for you to predict the mechanism. Right?? The first conditions might remind you of diazonium synthesis. While our starting material nucleophile is different as it s not a primary amine, the electrophile is the same. Under acidic conditions, the nitrous acid can be further protonated and decompose into the highly electrophilic nitrosyl cation. Because we have a tertiary amine, we don t have a diazo synthesis but rather, an electrophilic aromatic substitution. This installs a nitroso group. Next, reduction with metals or other reductants reduced the nitroso group to the amine. The third step is the crux. Ferric chloride is iron 3+, so it can function as a single electron oxidant. The primary amine is more reactive than the tertiary, so we form a radical cation that is stabilized into the ring. In parallel, the excess ferric chloride also reduces the hydrogen sulfide. These two radicals can combine, forming one of the carbon-sulfur bonds present in methylene blue. Unsurprisingly, the imine isomerizes to restore the more stable aromatic system. There is a true excess of ferric chloride checking Caro s 1876 US patent tells us he used 200 hundred litres of solution for roughly kilograms of starting material. This means the primary amine can be oxidized again. This time, the stabilized radical lands next to the sulfur, so through yet another oxidation, a thioketone is formed. This intermediate can t isomerize to restore aromaticity so rather, the electrophilic imine is attacked by unreacted starting material. This brings in the second half of methylene blue and through elimination of ammonia, one part of the bridging ring is completed. The sulfur remains pretty electron rich due to the conjugated electron-donating group. So, it’s activated for another oxidation. This creates a sulfur-based radical that can react with the second half, forming the second carbon-sulfur bond. Finally, I don t know if we would have another oxidation and deprotonation, or simply a hydrogen radical loss. In any case, this last step is clearly favored as it forms the aromatic ring in methylene blue. Pretty wild ride, but this is characteristic of much of 19th century chemistry which was pretty random and often based on redox chemistry-madness. To understand methylene blue story, we need to refresh our understanding of malaria. This disease is caused by parasites of the plasmodium family, transmitted by the Anopheles mosquito as a vector. After a bite of an infected mosquito, injected spores multiply in the liver, and subsequently invade red blood cells. They continue multiplying until the cells burst, causing fever, and circulate in the blood stream. A new mosquito taking a sip of this blood can close the cycle again as the plasmodium reproduce and form new oocysts inside the mosquito guts. What does this have to do with Ehrlich? Well, he realized that methylene blue the same dye that he used for the staining of nerve cells also stained the plasmodium parasite. Ehrlich s next step was very bold, even for 1891. What effects would methylene blue have against malaria in humans? They assessed just two patients, but the benefits were rather remarkable: within a few days, fevers calmed down and pathogens cleared from the blood. This was the very first synthetic, rather than natural compound ever used in clinical therapy, marking the advent of chemotherapy. Of course, as Ehrlich put it, this means that the urine, full of excess methylene blue, turned blue as well. In feces, the compound is excreted in its reduced state. This leuco form is colourless due to the absence of the conjugated donor-acceptor pi-system. However, exposure to air can oxidize it again, regenerating the dark blue color. But how can a simple dye affect mighty parasites? Well, this redox activity is essential for two mechanisms. The first one operates with the oxidized methylene blue. Because it likes to get reduced again, it can compete at an enzyme of plasmodium that is supposed reduce the sulfur group of oxidized glutathione to a thiol in normal glutathione. This un-natural competition essentially inhibits this enzyme, leading to accumulation of oxidized glutathione that the parasite can t get rid of, increasing harmful oxidative stress. The second mechanism relates to hemoglobin, the protein in our red blood cells, responsible for carrying oxygen. You will likely know that it features an iron 2+ ion which is where the oxygen molecules attach to. The parasite digests this protein but is not a real fan, because both the individual iron and heme groups cause oxidative damage – similar to what we’ve just seen with glutathione. The sneaky fellas have evolved food vacuoles and enzyme machinery that convert the toxic heme to an insoluble crystal polymer, so called hemozoin. This degradation prevents the parasite s death but importantly, starts from the oxidized form of hemoglobin with an iron 3+, so called met-hemoglobin. This is where methylene blue kicks in. Remember the colourless form? This reduced system can easily give away its newly found electrons to reduce the iron 3+ in methemoglobin to iron 2+. This regenerates hemoglobin and prevents the parasite s sneaky plan. It can t avoid the oxidative damage from excess iron 2+ and the heme ligands, and its replication is suppressed. Using the color blue to treat disease maybe this reminds you of the ancient practices of ground-up mummy powder. But check this out. Methylene blue is a legit medicine as it s approved by the FDA for treatment of acquired methemoglobinemia. This condition can be caused by other drugs that increase oxidative stress, leading to oxidation of normal hemoglobin and thus high levels of met-hemoglobin the blood. The oxidized form of iron 3+ has a lower affinity to O2, so met-hemoglobin has a lower oxygen-carrying capacity. In severe cases, effects can even be lethal. Methylene blue works the same way again it s reduced to its leuco form, which can reduce met-hemoglobin to hemoglobin. This time, instead of messing with the malaria parasite, it allows for recovery of the patient s oxygen transport. It works pretty well for this case, and I think it s pretty insane that the first synthetic compound ever tested in the clinic, is still used in humans today. On the other hand, use in malaria did not take off, because other compounds like quinine proved more effective. However, research has continued to show that combinations with methylene blue are still effective. On this chart, you can see the proportion of malaria patients that are infectious after 0, 2 or 7 days. The third set of columns is a combination of two drugs, which on their own are not too effective. The patients that received methylene blue on top of these two drugs were non-infectious after day 2. That s pretty remarkable. By the way? How do you think the researchers were able to tell if a patient was infectious or not? They simply offered their blood as a meal to new mosquitoes. Remember the malaria cycle! If the patients are infectious, the mosquitoes would unfortunately see some nasty plasmodium deposits in their guts. Before we get to the butt eating, I wanted to thank all of my channel members for their continued support, and thank you for watching this video. If you re new to the channel or for some strange reason have not subscribed or liked the video yet, please consider doing so! When not turning people blue, Ehrlich experimented with several other dyes and as was in fashion back in the day, swapped out most of his diet for strong cigars. But more importantly, he was busy with pioneering the field of immunology including the discovery of the complement immune system. This was just one of his contributions that ultimately netted him the Nobel Prize in 1908. As many of you will know, this system consists of protein precursors circulating in the blood which support killing of foreign cells. Nowadays, we know that complement is implicated in many diseases that have an immune component, so many modern medicines aim to regulate this system. Ehrlich continued with a breakthrough of drug discovery, but not in immune mediated diseases. Rather, the gruesome STD syphilis was Ehrlich s next quest. Looking at the various effects, syphilis was arguably one of the largest public health burdens between the 16th and 19th century. In 1905, two German scientists identified the coil-shaped treponema bacteria as the culprits behind disease transmission. Ehrlich s concept was again put to the test: would he be able to find a chemical that selectively harms the bacteria, while not compromising the patients health? Not harming patients is obviously important. The old school standard of care was treatment with mercury, through intramuscular injections or other ways. Because doctors were enthusiastic and accurate dosing of things like mercury fumes is impossible, the supposed cure often proved worse than the disease. If mercury can t help, how about arsenic compounds? Said no sane scientist, ever. Nevertheless, the compound atoxyl came into the spotlight of antibiotics. It had shown promising effects for sleeping sickness, which is another disease you might have heard of, transmitted by the infamous tse tse fly. Atoxyl was believed to not be harmful hence its name but larger testing in patients by the german physician Koch caused blindness in 2% of cases. Shouldn t surprise you that this compound might be dangerous. Ehrlich was asked if he could do something with atoxyl to make it mo re benign, while still maintaining its efficacy. The first problem was: what is atoxyl? You see, atoxyl was synthesized by the French chemist B champ in 1854. This guy had just found a pretty pioneering reduction of nitro-benzene to synthesize aniline. Playing around with his product, he thought addition of arsenic acid gave the anilide form, stemming from nucleophilic addition of the nitrogen. However, after Ehrlich s co-worker Bertheim spent enough time working on the compound, he corrected the structure, corresponding to an electrophilic aromatic substitution in B champ s reaction. In these good old days, structures were often determined by assessing reactivity. Amongst other experiments, they found that atoxyl underwent diazo-formation with nitrous acid similar to the reaction we ve seen in methylene blue but this reaction would be impossible for the substituted anilide structure. Having figured out what atoxyl actually is, Bertheim and Ehrlich systematically modified and tested over a thousand compounds. One of them was compound 606, which they assumed contained an arsenic-arsenic double bond. This agent didn t work against sleeping sickness, but some years later, through some serendipity and the involvement of the expert Japanese bacteriologist Sahachiro Hata, they found promising effects against syphilis. Hata diligently tested the compound over and over in mice, then guinea pigs, and then rabbits, confirming its benefit and even prophylactic effects. Obviously, they knew about Paracelsus quote of the dose makes the poison , so they investigated the effective dose versus the maximally tolerated dose. For their time, their pre-clinical research was very systematic and thoughtful. The side effects were better compared to atoxyl, with severe neurological problems like vision loss not occurring. However, direct subcutaneous injections did cause pain and necrosis, so unintended cell death in the skin and other tissues. Keep this in mind. To avoid the mistakes made by Koch the german physician who blinded people with atoxyl Ehrlich arranged carefully recorded clinical studies, giving guidance on dose and selection of patients himself. The human tests proved successful, with much of the medical community deeming this a breakthrough. It was sold as Salvarsan in 1910 but was not the optimal one-time, magic bullet chemotherapy that Ehrlich had hoped for. The first issue was that Salvarsan was an unstable solid and prone to oxidation requiring dissolution for IV infusions or subcutaneous injections. As you might tell from its structure, Salvarsan is not particularly hydrophilic, so its solubility in water was very limited. This meant injections were slow and painful the injected volumes were initially 10-times what some of you might know from at home autoinjectors. Doctors tried to find nicer liquids but as you can see, some of them caused even more pain, requiring use of narcotics. Remember the necrosis which Hata identified? Well, that was also an issue. I found it particularly disgusting that the injections often led to deposits that remained un-absorbed basically, arsenic living rent free in their butt cheeks, and greenish-yellow areas. Yikes! General toxicity was also some of a risk. Aside from low tolerability causing issues like diarrhoea, this case report describes a patient becoming deaf. At least this was less common than the blindness triggered by Koch s atoxyl trials. No wonder did Ehrlich famously state the challenge of translation from research to real-world settings. Let s conclude the Ehrlich saga with three interesting chemistry insights. First, just two years later in 1912, neosalvarsan was launched. As you can tell, the sulfinate group increases solubility in water and conveniently also shelf life by reducing unwanted oxidation of salvarsan. It s pretty remarkable that the world s first designed drug also saw the world s first next-generation molecule, trying to make the original better. This is definitely more innovative than deuterium copycats for example, as we have seen in our previous video. The second insight relates to why Salvarsan, despite generally being decently safe, still showed variable toxicity between different batches of product. You see, Ehrlich s cooked up a precursor featuring a nitro and arsonic acid together with dithionite as a reducing agent. This simultaneously reduced the nitro group and arsenic and gave the condensed product. This synthesis was not always reproducible, and resulted in sulfur-containing impurities that were likely not always removed to the same degree. The isolation was just a conversion to the dihydrochloride salt, followed by immediate filling into inert vials or ampules due to the low stability. So, depending on the chemist s rigor and luck, the drug product injected into patients might not have been the cleanest. As the field was exploring modified syntheses, it was found that a dithionite reduction at low temperatures instead of 60 C selectively reduced just the nitro group. This allowed an additional purification step with charcoal and crystallization. Then, in a second step, reduction with hypophosphorous acid gave Salvarsan. The product obtained in this manner was much more pure and accordingly less toxic. I don t know what they used for large scale manufacturing down the line, but I hope not too many people lost their hearing. Our third insight to wrap up the video relates to the drug s structure. We ve drawn the arsenic-arsenic double bond several times now, but this structural element proposed by Ehrlich was actually debated for long. Due to its properties, Salvarsan proved too tricky to be characterized with old school methods was not possible due to its properties. Crystals were not suited for X-ray analysis and traditional mass spectrometry did not work due to its very low volatility. Only in 2005, chemists managed to solve the mystery by using electrospray ionization mass spec. Check this out: The profile does not look like something you would want to inject into humans. It turns out that Salvarsan actually consists of different cyclic species in solution, with three and five-membered cycloarsines being preferred. I think it s hilarious that the first synthetic drug discovered by humanity looks devilish as heck. But even funnier is the following: Despite the systematic modification and medicinal chemistry work from Ehrlich that we ve covered, Salvarsan is actually not the active agent in the body. Instead, it is oxidized and serves as a delayed-release source of the monomeric arsenic acid. Exposure to air and water converts most of the different-sized rings to the active drug after some time. The cursed arsenic drug is not used against syphilis today, because the discovery of penicillin in the mid-20th century made it obsolete. Mummy corpses are also not regularly violated anymore. But as you know now, our modern arsenal of medicines also still features methylene blue. In one of our next videos, we will take a look at other random, even crazier compounds were used as drugs. I can tell you, will not want to miss that one. If you haven t seen all my other videos you should! Thanks again for supporting this channel, and I will see you in the next one.