Eternal youthful beauty - with Botox, the miracle injection, this dream seems to come true. Wrinkle smoothing with the neurotoxin botulinum toxin is very fashionable worldwide. Within a few years, Botox has become a real blockbuster. And the trend is still rising rapidly. Botox is the trade name of one of the botulinum toxin preparations. Apart from the question, why people have a highly toxic substance injected into their face in search of the eternal fountain of youth, this trend also causes a serious animal welfare problem.

Every single production unit (batch) is tested for toxicity. The test substance is injected into the abdominal cavity of mice. The animals’ agony includes cramps, paralysis, and shortness of breath, and can last three to four days. The animals eventually die from respiratory failure. Estimates suggest that at least 350,000 - 400,000 mice are tortured to death in Europe alone - worldwide the number is likely to be many times higher. The trend is rising. Non-animal test methods have already been approved by the authorities, but not all manufacturers use them.

What is Botox?

"Botox" is a trade name for a product made from the bacterial toxin botulinum toxin. The name “Botox” has become established, pushed by mass media, as a generic term for wrinkle smoothing through injections with the bacterial toxin. Just as “Scotch tape” has become established for adhesive tape or “Kleenex” for facial tissues.

Botulinum toxin is produced by the bacterium Clostridium botulinum, which can be found in spoiled canned meat. Botulism used to be a feared, often fatal food poisoning. Affected people died from respiratory paralysis in terrible agony.

Botulinum toxin is the most powerful poison currently known. Even unimaginably tiny amounts, a hundred-thousandth of a milligram, can kill a person. If one were to pour a tablespoon of the substance into Lake Constance, the lake would be completely poisoned (1).

How does botulinum toxin work?

Botulinum toxin blocks the signal transmission from the nerves to the muscles. Consequently, this leads to muscle paralysis. In the event of poisoning, the respiratory musculature is paralysed and the person suffocates in agony. When highly diluted, this property is used to treat a wide variety of diseases. Muscle cramps, dystonia (certain movement disorders), torticollis, writer's cramp, eyelid twitching, excessive sweating, squinting or migraines can be treated by this procedure.

By injecting Botox into the facial muscles, they are paralyzed and the wrinkles above them are smoothed out. The effect wears off after about two to six months. This is how long it takes the body to recreate new nerve endings.

The neurotoxin is also used as a biological warfare agent. Because it breaks down quickly when exposed to air, an area contaminated with Botox can be safely re-entered after just a few days.

Which animal experiments are carried out?

Because botulinum toxin is so incredibly dangerous, it must be heavily diluted for human use. There may be fluctuations in production. Each production unit (batch) is therefore tested for safety before it can be sold.

The toxin is injected into the abdominal cavity of groups of mice. Each group receives a different dilution of Botox. The amount at which exactly half of the animals die is determined. This is called the LD50 test (LD50 = lethal dose in 50% of animals).

This is associated with terrible suffering for the animals. Muscle paralysis, visual disturbances, and respiratory distress occur. The agony can last three or four days. The rodents eventually suffocate while fully conscious.

At least 100 mice are used for each production unit of the bacterial toxin (2). In the field of chemical testing, modified LD50 tests have been accepted for years, in which fewer animals are used and the animals suffer slightly less. Not even these minimal improvements are allowed in Botox batch testing. The classic LD50 test is carried out, in which death is the end point, i.e. one waits until the mice have died a gruesome death.

In addition to batch tests, so-called stabilization tests are carried out for the approval of a new preparation or if something changes in the manufacturing process. This is to guarantee that the manufacturing process is stable and always executed in the same way. These stabilization tests are carried out in the first five years after a product has been approved and involve a particularly large number of animal experiments. If the manufacturing process is then running smoothly, “only” each production unit is tested from there on. This reduces the number of mice tortured to death but it increases again when more of the newly approved preparation is sold.

How many mice have to suffer and die for Botox?

In 2008, 74,000 mice suffered and died in Botox tests for the Ipsen company, and 34,000 mice for Merz in the same year. An estimated 600,000 mice were used worldwide in 2008 by the three major companies Allergan, Merz and Ipsen (3). According to recent research, in 2015, 350,000 - 400,000 mice had to suffer and die for Botox injections in Europe alone (14) - worldwide the figure is likely to be many times higher, as the huge markets in North America and Asia are not taken into account in these numbers. Because of the continually increasing demand, the number of mice killed will continue to rise. According to the German Federal Government's AnimalTestInfo database, LD50 tests on 150,000 mice were approved in 2014. Of these, 90,000 mice were accounted for by the company Merz and 60,000 mice by the company Eisai. Both tested their products at the Hamburg contract laboratory LPT until its closure.

The Japanese company Eisai, with a branch in Frankfurt, “used” for its botulinum toxin B preparation Neurobloc® 60,000 mice in 2014 and further 30,000 in 2015. In 2018, the product was sold to Sloan Pharma, which has offices in Luxembourg and Switzerland. Sloan Pharma had permission to perform LD50 testing on 46,800 mice in 2019. This information originates from the German national database AnimalTestInfo, in which approved animal testing projects are recorded. In December 2022, Sloan Pharma discontinued its Neurobloc® product and withdrew its approval. According to the company, a follow-up product is not planned.

However, in 2021, according to AnimalTestInfo, Botox tests on 22,440 mice were approved in Germany again. Unfortunately, it was not possible to determine which company was responsible for this.

Ireland is the largest Botox tester in the EU. In 2021, over 80,000 mice were killed there for such purpose, while the year before, over 100,000 mice were killed. 

What suffering is inflicted on the mice?

The British organisation Cruelty Free International conducted undercover investigation at the animal testing laboratory Wickham Laboratories, Hampshire, UK. The footage, recorded between February and October 2009, documented mice twisting in agony. The animals had the botulinum toxin Dysport® injected into their abdominal cavity.

For Dysport® alone, at least 74,000 mice had to undergo the cruel LD50 test in 2008. According to Wickham's own data, 989 mice were killed in the LD50 test in one day in July 2009.

The undercover footage showed mice with paralysis and respiratory distress. Mice that survived the test had their necks broken with a pen or were gassed.

The ugly truth about botox >>

What non-animal tests are available?

Market leader Allergan and the Frankfurt-based company Merz received regulatory approvals for cell culture methods for testing their Botox products in 2011 and 2015, respectively. These are tests developed and validated by the companies themselves using human cell cultures. However, both companies have so far only replaced a large part of their animal testing, the so-called batch testing. Allergan assumes an 80% replacement of its Botox animal tests. 20% are still carried out for other types of testing (bulk testing) and due to legislation in other countries.

Merz has reduced its animal testing from 90,000 in 2014 to around 11,400 in 2016. In 2022, the company informed us that the number had been reduced to zero in Germany. However, animal testing would still be done for other countries.

According to its own information, the French manufacturer Ipsen received approval in the EU and Switzerland for a cell-based test in August 2018. The company does not disclose how many animals are saved as a result.

What is the legal situation?

The European Pharmacopoeia regulates the manufacture, designation, testing, and storage of pharmaceutical products at international level. The rules are drawn up by commissions of the EU authority, the European Directorate for the Quality of Medicines & Health Care (EDQM) in Strasbourg. In Germany, the Federal Institute for Drugs and Medical Devices (BfArM) in Berlin is responsible for the implementation.

The European Pharmacopoeia demands the LD50 test in mice for each production unit (batch) of botulinum toxin (6). In addition, a reference substance with a known toxicity level must be tested for each batch. Since 2005, three alternative methods have been approved, but only if they provide the same results as the LD50 test.

In this so-called validation, non-animal test methods are compared with the animal test to be replaced. The alternative method is approved only if the results of both methods are the same. The problem is that animal testing is an unreliable, poor test method. New, better systems hardly have a chance of being approved because they do not deliver the same results than the animal test, but much better ones. Individual manufacturers develop and validate "alternative tests" for their botulinum toxin products. These are only approved for the respective company.

Why are animal tests carried out for Botox, although animal testing for cosmetic products is prohibited under the German Animal Welfare Act and the EU Cosmetics Regulation?

Animal testing for cosmetics has been banned in the EU since 2004. In Germany, such a ban has been in place since 1998, at least for cosmetic end products. Since March 2013, the marketing of both products and their ingredients that were tested on animals has been banned in the EU.

Botulinum toxin products are approved as medicines, which is why the ban on animal testing for cosmetics does not apply. Furthermore, they are not considered cosmetics because they are injected and not just applied topically (12).

In this way, the existing ban on animal testing for cosmetics in the EU and Germany is circumvented. The majority of consumers who are against cosmetic animal testing are being deceived.

When it comes to wrinkle-free skin, doctors make use of the so-called "off-label use", i.e. drugs approved for certain medical indications are rededicated. Patients have to sign an agreement that they are being treated with a drug not approved for treatment of facial lines at their own risk. In Germany, Vistabel®, Azzalure®, and Bocouture® are approved for the treatment of the glabella fold between the eyebrows. All other cosmetic applications are "off-label".

There are tendencies to declare wrinkles as medical problems in order to legalise aesthetic Botox applications.

Due to the presumably widespread "off-label use", it is not possible to determine what percentage is actually used for cosmetic or medical indication.

How was Botox discovered?

Botulinum toxin is the poison of spoiled sausages. It was already known in the 18th century that eating spoiled blood sausages can have fatal consequences. In 1895, the Belgian microbiologist Emile van Ermengen succeeded in identifying the often fatal "sausage poison". He coined the term "botulinum toxin", from Latin "botulus = sausage" and "toxin = poison".

In 1946, botulinum toxin A was produced in pure form for the first time. The targeted paralysis of individual muscles was tested on monkeys. The potential of the nerve toxin was already recognised, but a suitable human disease still had to be found for it to be used. From 1980, the American ophthalmologist Allen Scott used the poison to treat strabismus by injecting it into the outer eye muscles. In the 1980s, further indications were tested on volunteers. In 1989, the first botulinum toxin A preparation from the company Allergan was approved in the USA. It was initially called Oculinum and was used for strabismus, blepharospasm (eyelid spasms), and other muscle disorders. Later, the drug was renamed Botox. The range of therapeutic applications increased dramatically. Today, botulinum toxin is used for over 50 nerve and muscle disorders (2).

In the late 1980s, the American physicians Jean and Alistair Carruthers, discovered that the toxin also smoothed facial lines. In 1992, they published an article about their observations in a scientific journal. Botox was now increasingly used for cosmetic purposes, although it was only approved for this purpose in the USA in 2002 (2). Promoted by reports in the tabloid media, the bacterial toxin soon became a best-seller as a lifestyle drug.

Who produces botulinum toxin?

There are 9 types of botulinum toxin, which are labelled with letters A, B, C. etc.... Mainly botulinum toxin A is used for medical and aesthetic purposes. Botulinum toxin B is also available as a medicine. In the following list, only the company Eisai offers botulinum toxin B, all others offer A.

The following companies operating in Europe offer botulinum toxin preparations:

Botulinum toxin A

Allergan Inc., Pforzheimer Str. 160, 76275 Ettlingen: BOTOX®, BOTOX® COSMETICS, Vistabel®

All three products are identical. BOTOX® is the original product. In 2002, BOTOX® COSMETICS was approved in the USA for cosmetic purposes. It is marketed in smaller dosages than BOTOX® for medical indications. BOTOX® COSMETICS is now approved in numerous countries. In Germany and other European countries, it is marketed under the name Vistabel®. Vistabel® has been approved in Germany since 2006 for the treatment of the line between the eyebrows (glabella fold). Production and animal testing for the European market were conducted in Ireland. At the beginning of 2012, an EU-wide approval of an animal-free cell test developed by Allergan was granted.  

Ipsen Pharma GmbH, Einsteinstraße 30, 76275 Ettlingen: Dysport®

Dysport® is only approved for medicinal purposes. The animal tests were carried out around 2009 at the contract laboratory Wickham Laboratories in England, later presumably in Ireland. According to Ipsen, a cell test was approved by the competent authorities in the EU and Switzerland in August 2018. 

Galderma Laboratories GmbH, Georg-Glock-Str. 8, 40474 Düsseldorf: Azzalure®

Galderma was originally a joint venture between Nestlé and L'Oreal and was fully acquired by Nestlé in 2014. Azzalure® is Ipsen's Dysport® preparation prepared for cosmetic use. Azzalure® is marketed for cosmetic purposes only. In Germany, Azzalure® is approved for the smoothing of the glabella fold between the eyebrows and the "crow's feet" on the sides of the eyes.

Merz-Pharma GmbH & Co. KGaA, Eckenheimer Landstraße 100, 60318 Frankfurt am Main: Xeomin®, Bocouture®

Xeomin® is approved in Germany for only two purposes, torticollis and blepharospasm. Merz has also had Bocouture® on the market since September 2009, which is approved for cosmetic applications (glabella fold between the eyebrows). Animal testing was carried out at the contract laboratory LPT in Hamburg until its closure in February 2020. In 2015, the company received EU-wide approval for an animal-free cell-based test, so that the number of animal tests conducted annually could be greatly reduced. According to the company's own information, animal testing was reduced to zero in 2021, at least in Germany. 

Croma Pharma GmbH, Industriestraße 6, A-2100 Leobendorf

The Austrian company acquired a licence in 2014 to distribute botulinum toxin A products from the Korean company Hugel in Europe and North America. The distribution partner in Germany is Stada AG. The product was launched under the name “Botulinumtoxin” in eleven European countries in 2021. According to its own information, the company conducts animal experiments for the approval of its product, but not for batch testing. A test with mouse cells is being developed for this purpose. 

Nestlé Suisse S.A, Postfach 2222, 1800 Vevey, Switzerland: Dysport®, Azzalure®

The food giant Nestlé entered the Botox business in 2015. The Swiss company distributes the botulinum toxin products Dysport® and Azzalure® from the manufacturer Ipsen through its newly established Nestlé Skin Care division. Through the complete takeover of Nestlé-L'Oreal's subsidiary Galderma and the purchase of distribution rights for Dysport® in the USA from the American company Valeant Pharmaceuticals - the main competitor of the market leader Allergan - Nestlé has tapped into a hugely lucrative and rapidly growing market. 

Botulinum toxin B

The effect of botulinum toxin B sets in faster, but lasts for a shorter period of time. Off-label use in the cosmetic field is therefore not very effective. Some patients develop neutralising antibodies through excessive use of Botox, which means that the injection no longer works. In these cases, botulinum toxin B can be used (2).

Sloan Pharma Sarl, Bertrange Cham Branch, Alte Steinhauserstr. 21, 6330 Cham, Switzerland

Sloan Pharma distributes the botulinum toxin B preparation Myobloc of the US company WorldMeds under the name Neurobloc® in Europe. Neurobloc® is approved for the treatment of torticollis. In 2018, Sloan Pharma bought Neurobloc® from the Japanese company Eisai for distribution in Europe. Eisai Germany was responsible for 90,000 LD50 tests at the Hamburg laboratory LPT in 2014/2015. After the transfer to Sloan Pharma, this company got LD50 tests on 46,800 mice approved at the LPT in 2019. In December 2022, Sloan Pharma withdrew the approval for Neurobloc. 

There are many other producers of Botox products: Acorda Therapeutics, Aldeyra Therapeutics, Azidus Brasil, Biolab Sanus Farmaceutica, Canbex Therapeutics, DAEWOONG, Emcure Pharmaceuticals, Endo Pharmaceuticals International, Evolus, Fresenius Kabi, GlaxoSmithKline, INSYS THERAPEUTICS, Johnson & Johnson, Lannett, Lanzhou Institute of Biological Products, LEO Pharma, Mylan, Par Pharmaceuticals, Pfizer, PhytoTech Therapeutics, Revance Therapeutics, Sun Pharmaceuticals, Takeda Pharmaceutical, and UroGen Pharma (13).

How is the market for botulinum toxin products developing?

After three to six months, the human body has rebuilt the paralysed nerve endings, the anti-wrinkle effect wears off, and the treatment has to be repeated. Beauty patients become permanent customers. An injection costs between 200 and 600 euros. Six- to 20-fold profit margins are possible for the physician administering the treatment (2).

Driven by mass media, the Botox boom is a huge business with enormous growth. In 1993, the Allergan company made worldwide sales of $25 million with BOTOX®. By 2001, sales had increased to $310 million. With approval for cosmetic use in the US in 2002, sales soared to $430 million in just one year. By 2006, Allergan was making $1.2 billion in sales of its Botox products, and by 2017 it was making as much as $3.17 billion (2,8,9,9b). Ipsen's Dysport grew by 14% to €128 million in 2007 (9).

43% of the BOTOX® production in 2005 was used for aesthetic purposes, 57 % in the medical field (8). In terms of the most frequently performed cosmetic surgical procedures worldwide, botulinum injection is the undisputed leader with 4.63 million treatments (9c). In Germany, it ranked second in 2017 with 19% of all cosmetic procedures, just behind tissue relining (20%) (10). In the USA alone, anti-wrinkle injections were administered more than 1.2 billion times in 2017; in Germany, the figure was in that year 220,000, more than twice as many as in 2008 (9a).

Are there any dangers for humans?

In the application of botulinum toxin for both medical and cosmetic purposes, serious side effects have been reported sometimes. Spreading of the toxin to an undesirable location can result in muscle weakness or difficulty in swallowing for weeks and thus to food aspiration. Over 600 cases of severe adverse effects and 28 deaths have been documented internationally (10). In February 2008, the US Food and Drug Administration (FDA) reported severe side effects again. Several children with spastic paralysis had died as a result of the treatment (11).

What is to be done?

The problem must be made public. Every citizen, every cosmetic surgeon or dermatologist must know that there is an infinite amount of animal suffering associated with anti-wrinkle injections. The more botulinum toxin is used, the more animals have to suffer. Pressure must be put on the manufacturers to continue to develop and validate animal-free procedures. Above all, the LD50 test on mice must be deleted from the regulations.

Dr. med. vet. Corina Gericke
Last update: 21st September 2023

References

(1) Dr. Friedrich-Wilhelm von Hesler from the Charité Teaching Hospital in Berlin, quoted in ZDF, Frontal 21, 12.10.2004
(2) K. Botrill: Growing old disgracefully: The cosmetic use of botulinum toxin. 2003, ATLA 31, 381-391
(3) S. Bitz: The Botulinum Neurotoxin LD50 Test – Problems and Solutions. Altex 27, 2/10, 114-116
(4) D.W. Straughan: Progress in applying the three Rs to the potency testing of botulinum toxin type A. 2006, ATLA, 34, 305-313
(5) Potency without Pain - Dr Hadwen Trust Science Review 2006, S. 7-8
(6) European Pharmacopoeia 5.0, Botulinum Toxin Type A for injection, 01/2005:2113
(7) European Directorate for the Quality of Medicines (EDQM), personal communication
(8) Allergan: Annual Report 2005, S. 8
(9) Pharmafirma Merz will in den Botox-Markt, Der Tagesspiegel, 2.2.2008 
(9a) Statista: Anzahl von Botulinumtoxin-Behandlungen ("Botox") in ausgewählten Ländern 2017 (accessed on 4.12.2020)
(9b) Statista: Allergan: Umsatz mit Botox im Jahr 2017 (accessed on 4.12.2020)
(9c) Statista: Infografik: Zahl der Schönheits-OPs nimmt zu (accessed on 4.12.2020)
(10) DGAEPC-Statistik 2016 - Zahlen, Fakten und Trends der ästhetisch-plastischen Chirurgie, S. 5 (PDF)
(11) FDA News: FDA notifies public of adverse reactions linked to botox use, 08.02.2008 (abgerufen am 4.12.2020)
(12) Ruhdel, I.: Über Tierversuche zum faltenfreien Gesicht. ALTEX 21, 1/04, 23-25
(13) Technavio: Global Botulinum Toxin Market 2017-2021 (abgerufen am 4.12.2020)
(14) Katy Taylor, Corina Gericke, Laura Rego Alvarez: Botulinum toxin testing on animals is still a Europe-wide issue. ALTEX 2019; 36(1): 81-90 (accessed on 4.12.2020)