BONE DEGREASING INFORMATION

Before discussing bone degreasing, it is important to understand the components of bone and fat. Bone is made of a mineral called hydroxyapatite, which is calcium phosphate [Ca3(PO4)2]3Ca(OH)2. Collagen makes up a significant part of the bone matrix. Type 1 makes up to 90-95% of all collagen in bone (Eriksen, et al, 2009 and Shehata and Krap, 2024). Bone is about 70% inorganic, 5% water and 25% organics (90% type 1 collagen, 10% non-collagen proteins and organics) (Lambri et al, 2018 and Pang et al, 2021).

Hydroxyapatite can be thought of as a rigid and hard, yet brittle, scaffolding for the collagen. Collagen provides flexibility and tensile strength. There are four relevant kinds of bone for this document. These are divided into two macroscopic types and two microscopic types. The two macroscopic types are cortical bone and cancellous bone. Cortical bone is dense, often thick, and found on the outside of most bones. Cancellous bone is found on the interior of bones and contains marrow. This is where much of the grease is found.

The two microscopic types are woven bone and lamellar bone. These are separated by the orientation of collagen (Bosch et al., 2011). Woven bone has random collagen fibers and is disorganized. It contains more osteocytes and fewer lamellae. This is common in juveniles and fish. Lamellar bone, found in adult mammal bone, is more organized, contains more lamellae and parallel collagen fibers. It is stronger and more resistant to heat.

Most people understand that compromising the minerals in bone results in a poor product. Altering the collagen will change bone mechanics (Viguet-Carrin et al, 2005) leading to brittle bone (Burton et al, 2014) or possibly flakey, powdery or chipping bone. This can occur chemically, enzymatically or with excess heat. It is therefore imperative to shelter collagen as optimally as possible for the best results.

Collagen is the most common protein in the body and a natural polymer that is essential to bone. Bone can be thought of as a collagenous tissue which happens to be mineralized, rather than a mineral which has collagen in it. There are twenty eight types of collagen (Ricard-Blum, 2011). Bone is overwhelmingly composed of collagen type 1, which exists as a triple helix bound by three types of cross-links (pers. comm., De Jonge*). These cross-links are important for maintaining the structure of the protein. Cross-links in collagen can be modified chemically, adding either permanent or temporary support for the polymer. These can provide additional thermal resistance. Collagen can form co-polymers with surprising materials to give incredible qualities.

Other than a low pH environment, which damages the mineral portion of bone, it is typically collagen which restricts what can and cannot be done to bone. It sets the heat limits, the need for a weak base versus a strong base, the time in maceration, the kinds of enzyme and exposure to them and most anything else. Switching views from bone as a mineral tissue with collagen in it to a collagenous tissue that has mineral support and paying special attention to the collagen will yield better results.

For all its strength, collagen has many weaknesses. These include heat, enzymes, and oxidation. Collagen is unstable at body temperature (Leikina, et al 2002 and Miles and Ghelashivili, 1999) and experiences time dependent failures when exposed to heat (Al-Shaer and Forde, 2025). This is on a logarithmic scale, meaning acceptable exposure time dramatically decreases as temperature increases. As a result, by the time simmering temperatures are encountered, the time before collagen begins to suffer is minimal. This may also provide challenges for preparing various types of cold bodied animal bones, and those with less compact or lamellar bones (such as juveniles and birds).

Despite challenges from bone type and body temperature, the Polymer-in-a-Box mechanism provides some thermal stability for bone-bound collagen (Miles and Ghelashvili, 1999) and gives some explanation for stability slightly beyond body temperature while degreasing bone. Polymer-in-a-Box mechanism describes that physical confinement thermally stabilizes polymers, such as collagen surrounded by bone. Hypothetically, in the case of degreasing in solution, this outward pressure may have limited ability to expand before encountering bone confinement, which in turn provides support.

During maceration, simmering or any activity to remove flesh, it is important to consider that most flesh is also made of collagen type 1. Any activity that would modify it may also modify the collagen in bone. Macerating bacteria should have some trouble physically getting to the bone bound collagen, and because it is a “target rich environment” simply devour easier accessed flesh. A great example of materials or techniques damaging both flesh and bone is the use of lye to dissolve flesh. While mass-oxidation from hydroxide oxidizes the flesh and dissolves it, it also oxidizes the collagen in the bone, damaging it.

*Ally De Jonge, Bare Bones Solutions Biochemistry consultant. August 10, 2026.

When heating collagen, the molecule “unzips” and “rezips” back together like a zipper during cooling. Tangling is more likely during rapid cooling. A temperature of approximately 4 degrees Celsius seems ideal for several days after heating and slow cooling. This temperature is commonly used in papers preparing collagen and Bare Bones Solutions has subjective evidence to suggest some flexibility was recovered using a cooling period at this temperature in demineralized bone. While bound in bone, the terminals of the collagen polymer cannot split, and therefore it “bows out” in the middle. This action makes it more susceptible to insertions of chemical groups such as hydroxides, enzyme activity or other modifying activity that would normally find it difficult to damage collagen.

At temperatures above 53 degrees Celsius, collagen fibers shrink by 75% (Maximo and Cunha, 2010). These smaller collagen fibers are more elastic and are likely still bound on either end by bone. As a result, instead of providing tensile support the collagen begins to tug and pull on the mineral matrix. Therefore, overheating causes collagen to literally rip a skull apart over time. It may be the case that some treatments of collagen which provide extra cross-links tension the polymer and have a similar effect (pers. comm., De Jonge`).

Between 80 and 90 degrees Celsius, which are in ideal simmering range for bone preparation, collagen forms into chaotic gelatins. These gelatins can absorb water and may emulsify fats (Asghar and Henrickson, 1982). Gelatins derived from collagen are often sticky, and in a complex system like bone with wicking capacity they, and any emulsified fats and oxidized fats, may be difficult to dislodge. This may provide an explanation for “fats moving deeper into bone” or “fats cooking into bone” when overheating bone. Collagen may be altered as low as 45 degrees Celsius (Maximo and Cunha, 2010) which suggests ideal degreasing temperatures for adult mammals should be 44 degrees Celsius.

`Ally De Jonge, Bare Bones Solutions Biochemistry consultant, spring 2026.

Collagen requires hydration (pers. comm., De Jonge-). Hydrated bone plays a role in the collagen-cross links, which contribute to keeping the collagen tensed. Tense collagen lessens brittleness in bone. It is possible that over tensioning the molecule may cause problems for the mineral aspect of bone (pers comm, De Jonge$). Nyman et al (2006) outline that dehydration in almost all cases results in weaker bone.

One of the most accessible ways to hydrate collagen is with glycerol, which is a natural sugar and product of saponification. It works as a humectant, a water-attracting substance, bringing water to the polymer. It helps stabilize the triple-helix by increasing thermal stability and hydration (Venkitesan et al, 2026). It may be an important substance in rehydrating dry or damaged bone. It is miscible in water, and if being used to hydrate bone it should be diluted with water. Glycerol and other sugars are useful in sheltering collagen from heat. Given its polar nature, it doesn’t seem likely to interact with non-polar fats if used during degreasing. In properly hydrated bone, glycerol may compete with water for the hydroxide slots on prolines, and may not be the best option in all circumstances.

-Ally De Jonge, Bare Bones Solutions Biochemistry consultant, summer, 2026.

$Ally De Jonge, Bare Bones Solutions Biochemistry consultant, spring 2026.

Mobilized grease moves through bone via diffusion and wicking (capillary action). Saponification, dissolution, heat, water movement and grease rendering all impact the speed at which this works. In the case of diffusion, the degreasing mixture, carried by water, moves towards the hypertonic grease through the bone until it reaches equilibrium. Once there, it breaks down the grease through its typical means and the products are exported through diffusion gradients, moving from areas of greater to areas of lesser concentration outside the bone. Grease itself may move through the bone too, especially when heated in soapy water, to areas of lesser concentration. To expedite degreasing, using fresh water with degreaser in it is important to keep diffusion gradients high. For these reasons, and because it is less likely to harm collagen, it may be reasonable to use lower percentages of degreasing material to save money and increased water change frequencies. 

Capillary action uses intermolecular forces or cohesion from surface tension and adhesion between the liquid and surface of the bone to draw liquid into the narrow spaces of the bone. Heat causes the cohesive force to weaken and the adhesive for to strengthen. The net result is an increase in capillary activity, or in the case of water and grease, an increased wicking movement into the bones confined spaces. A heat threshold may exist which causes a critical movement of grease in bone, leading to increased difficulty of degreasing. It is possible that chemically mobilized grease may more successfully wick into bone.

In areas of stubborn grease, it is likely that the grease itself isn’t much more resilient but that there is an increased volume of grease and dense bone is preventing degreasing agents from entering or products from leaving. This may not be the case if the skull has a thermal history altering the triglycerides. It is possible that, with sufficient time and degreasing agents, the grease becomes entirely saponified. To test this, diabetes needles were used on a marten skull to access the dentary via the mental foramen and injected with degreaser. The maxillae were injected on the dorsal surface of the maxillary zygomatic process, directly into a pocket of “stubborn grease”. A tricky area around the basisphenoid was also injected. This was successful in rapidly breaking up these tough spots. It is unclear if this simply provided an entrance/exit, or if the direct application of degreasing agent made it successful. It seems reasonable to assume that it was probably a combination. This can be tested by using a needle to puncture a skull with no injection. Unfortunately, larger gauges of needle will be required in order to get through thicker bone on larger mammals which will leave a noticeable hole if either the tooth sockets or foramen aren’t used. This may be cosmetically repairable with minimal effort.

Drilling small holes in tooth sockets, in particular in the jaws and near the zygomatic, can aid in letting grease out and degreaser in to the denser dentary bone. This is potentially favorable because any damage by the drill is covered by replacing the tooth in the socket.

Water is the most ubiquitous ingredient in degreasing. It is a polar covalent solvent. Almost all applications of degreasing will include water. It serves as a host or diluter for most degreasing agents, soaps, oxidizers and as a carrier of lifted or saponified grease. Typically grease is discarded with water changes as the grease carrying capacity of water reaches its limit, the degreasing agent runs out or both. This can vary in timeframe depending on the amount of grease, heat of solution and strength of degreaser.

Many skull enthusiasts do not realize that long term water exposure can harm bone. This happens through several mechanisms which includes, among other things, bacterial action rotting the collagen, chelation of minerals from the skull, and carbon dioxide interacting with the water to form localized carbonic acid which then dissolves the bone. It is therefore useful to find ways to limit the time a skull is submerged.

It is therefore important to be aware that drying bone improperly or using solvents that may dehydrate bone can be harmful. These improper methods may include drying bone in a fume hood or in excessively dry conditions or using solvents such as ethanol or even acetone. Acetone remains a popular and effective choice for degreasing because it has superior wicking qualities. Differential moisture in layers of the bone may contribute to cracking as bone dries or absorbs water. This is true for teeth like canines. Being mindful of the effects of water on extremely wicking materials like bone can make a difference.

Most skull enthusiasts understand that heat mobilizes or renders grease and therefore heat greatly impacts the answer to a common question: How long to degrease a skull? Favorable temperatures vary based on animals, but typically are around 30 degrees Celsius or less for fish, 35 degrees Celsius or less for bird and reptile and 44 Celsius or less for mammals.

Maximo and Cunha, (2010) describes negative effects on collagen beginning at 45 degrees Celsius. Lambri et al (2018) suggests a temperature of 46 degrees Celsius may be ideal. Shrinking in human pelvic elements occurs at 61 degrees Celsius (Danielsen et al, 1994). Collagen denatures as low as body temperature (Leikina et al, 2002). Many skull cleaners use 48 degrees Celsius. Collagen in bone is more resilient than collagen found in other tissue, in part because of Polymer-in-a-Box mechanics (Miles and Ghelashvili, 1999). Fats for many mammals melt around 40 degrees Celsius with various culinary references on the internet suggesting bovid fat renders at 55-60 degrees Celsius. Soap, a product of saponification, melts at around 55 degrees Celsius.

Observations by Bare Bones Solutions suggest that 46 and 48 degrees Celsius are too high and encourages warping in the zygomatic joints in adult mammals, despite the popularity of 48 degrees Celsius. We conclude that 44 degrees Celsius is ideal for adult mammals, allowing for overshooting by heat controllers for those preferring ranges up to 48 degree Celsius. For those using gear with tighter control, 44 degrees Celsius respects observations by Maximo and Cunha (2010) and Lambri et al (2018).

Heat increases the speed at which chemical reactions occur. While there are critical temperatures for some reactions that aren’t reasonably achievable because of the collagen heat limit, appropriate application of heat is critical in speeding up the chemical reaction and limiting exposure to water and other potentially harmful chemicals.

Efforts have been made to accumulate fat rendering temperatures by several amateur authors. These temperatures are unreferenced and may not be accurate. As organ fat, back fat and other body-fat rendering temperatures may differ, these may not be laterally comparable to marrow-fat temperatures. This effort may also be futile because finding an uppermost temperature that doesn’t harm collagen is likely the best answer. This is because collagen presents a hard limit for heat, and using the highest possible heat maximally mobilizes the grease.

After testing on both dermestid and simmered skulls, cleaning with very low simmering (sub 60C) appears to provide superior degreasing times because significant degreasing occurs during simmering. It also is far more likely to cause damage to collagen because the margin for error is much tighter. Further excessive heat, such as 100C or greater, will cause loss of protein and “smoothing and softening of the bone” (Bosch et al, 2011). As the collagen is destroyed by heat, it may release CO2, HCO3 or Hydrogen ions. These harm the hydroxyapatite, causing damage as seen in Bosch et al, 2011 and noted in White and Hannus, 1983.

Saponification is the process by which fats are turned into glycerol and fatty acids. These fatty acids react with hydroxides to form fatty acid salts, also known as soap. This is achieved by combining a base, such as ammonia, with triglycerides like animal fat. There are different strengths and kinds of saponifiers out there, each with advantages and disadvantages. These may be the most cost effective way of dealing with triglycerides but are also problematic for collagen.

Weak bases such as ammonia and soda ash are popular skull degreasers. These bases don’t completely disassociate in water and therefore may throttle the amount of hydroxide in the water at any given time. Unfortunately, other than soda ash, metasilicate and baking soda, other common weak bases smell horrible including ammonia. No matter what type of base is used, hydroxides will be released and saponify. It is possible that controlling the concentration of stronger bases such as lye can produce favorable results like ammonia, though mitigating other processes like oxidation and chelation is a consideration when ions are involved.

One has to consider cost, strength, availability and negative side effects. Because it is covalent and does not oxidize beyond the hydroxides, ammonia is heavily favored in the skull cleaning community despite being a weak saponifier.

Hydroxides (OH) form from every chemical with a pH higher than 7. This includes ammonia (NH3), lye (NaOH), and potash (KOH). The relative concentration of OH is how pH is measured “basic”. Hydroxides react with, or hydrolyze, grease (fats) breaking the ester bonds in the grease, turning them into soap and glycerol. This is called saponification. Glycerols are technically a solvent, but because they are polar and fats are non-polar, fats don’t readily dissolve in them. Hydroxides are known to damage collagen, so should be controlled. It is possible that throttling the concentration of stronger bases, simulating the trickling effect of hydroxides from weaker bases such as ammonia, may make them more viable as degreasers of bone.

Ammonia (NH3) Overwhelmingly the most popular saponifying degreaser of bone. Weak base. It is cheap and readily available. Despite these advantages, it is a poor degreaser. Costs escalate over time and longer times in solution may cause water to damage bone. It is very noxious. Ammonia is a gas dissolved in water. It will off-gas as a result. Prices of ammonia are currently increasing as are regulatory considerations.

Ammonia works by the following equilibrium reaction (Turner-Walker, 2012):

NH3 + H2O  ↔  NH4+ + OH-

The pH of ammonia is approximately 12.5, and about 1% is hydrolyzed at any given time. The hydroxide ion and ammonium bind to some of the fatty acids to form ammonium salts and water. As this happens, more ammonia is hydrolyzed into ammonium and hydroxide, continuing the process until ammonia is used up or off-gassed completely. Importantly, hydroxide ions are known to cause damage to protein (collagen). As a result, ammonia does have bone damaging potential. This is usually negligible, unless the ammonia is overheated, the bone is overexposed, or the concentration is too high.

A common misconception persists in the skull cleaning community that ammonia and soap “cancel out”. Ammonia and soap function in two different and complimentary ways, meaning it is likely that it is more effective to combine them. Many commercially available soaps contain ammonia and cloudy ammonia contains soap. Ammonia breaks down soap scum, which is a deposit of calcium and soap.

Borax Creates a high pH environment and therefore likely saponifies somewhat. It breaks down into boric acid and sodium hydroxide and therefore isn’t ideal.

Potash and Lye Extremely strong degreasers. Easy to source, extreme handling concerns, storage and disposal concerns and noxious. Strong oxidizing agent that will destroy protein including collagen. Not recommended for cleaning bone. Common additives in industrial commercial degreasers in amounts of 0.5-1%.

Soda Ash is a popular hydroxide forming weak base.

Trisodium Phosphate (TSP) Cheap, powerful degreaser that comes in a powder for cheaper shipping. Do not mix with ammonia because it forms ammonia gas on contact. This was the first substance tested by what would become Bare Bones Solutions. It produces an alkaline solution of pH greater than 11. It is available commercially in both chlorinated and unchlorinated forms. It is versatile, finding use as an anti-bacterial food additive, in stain removal, grease removal, anodizing and more. It is cheap, easy to source and handle, efficient, lacks noxious smell and shelf stable. This gives TSP great advantages as a bone degreaser, though it is too powerful and damages bone even at miniscule concentrations.

It is banned in a dozen states for causing eutrophication in waterways, damages bone over time, and has a low thermal decomposition temperature (73.5C) which means degreasing must be done at a lower temperature. When heated, it can release toxic fumes. It is therefore ill-suited for bone degreasing.

The phosphate in TSP reacts negatively with proteins, including collagen. This is because it is an oxidizing agent. TSP will also react negatively with calcium in bone over time in a process called chelation. In this process, phosphate will bond with surface calcium from the bone to form aqueous apatite in solution.

Surfactants serve several roles, including as wetting agents, emulsifiers and solubizers. Wetting agents reduce the surface tension of water, allowing it to spread more easily and lift grease. Emulsifiers bind liquids that normally do not mix, creating an emulsification. This is achieved by dispersing one liquid as tiny droplets into the other. Non-polar solvents can be emulsified in water, which is highly polar, helping to bend the generalized rule that “like dissolves like”, or non-polar dissolves non-polar (as in triglycerides with non-polar solvents). Emulsifying a non-polar substance in water may have limited success, but some is better than none. Solubizers are used to make grease and other water-insolubles dissolve in water. This process is important, because it aids in mixing of water and grease which normally do not mix. It is accomplished by coating small parts of grease in hydrophilic micelles which then disperse and mix. In many cases, a surfactant will have several roles.

There are two primary types of surfactants available. These are ionic and non-ionic. Ionic surfactants are typically weaker but gentler on skin and eyes. Negatively charged, or anionic, surfactants may denature collagen in small amounts (Nandi et al, 1985). Positively charged, or cationic, surfactants do not appear to damage collagen. Non-ionic surfactants are stronger but harsher on skin and do not appear to damage collagen. They are found in many laundry detergents. Unfortunately, chlorine producing agents are also in many laundry detergents, so care must be taken when selecting a laundry detergent for use. Surfactants add an important element to the grease removal process, especially when used together with saponifiers.

Sodium Lauryl Sulfate and Laureth Sulfate Extremely successful and popular in bone degreasing dish soaps. Used in shampoos and dish soaps because they foam well. Anionic (negatively) charged surfactants that may denature collagen, even in small amounts (Nandi et al, 1985) but that may not apply to collagen bound in bone, as indicated by the widespread success of their use on bone. Sulfates are very weak oxidizing agents.

Powder Degreaser- Created as a stronger alternative to ammonia. Add to water. No noxious odor, no ammonia, lye or bleach. Biodegradable. Highly cost effective, some whitening abilities, works best when heated consistently.

Spot degreaser- Powerful citrus based bone degreaser. Useful for brute forcing stubborn grease spots, including clear grease, or speeding up projects.

Demineralizer- Removers calcium from bone. Originally made as part of a multi step collagen modification protocol. Allows bone to bend, shrink and act more like rawhide. Bone tattooing is possible because of this material.

Organic solvents appear to have the most success (Guilmnot et al, 2014), though they are volatile with low vapour pressures, expensive and are rapidly absorbed through skin (Boman and Maibach, 2000). Polar solvents will be miscible in water and may be less likely to dissolve triglycerides, but not always. Non-polar solvents will not be miscible in water and tend to dissolve triglycerides readily. Choosing the right solvent for the job can be difficult based on cost, disposal, volatility and even danger level.

According to Turner-Walker (2012), the most effective solvent cleaning was a combination of acetone, ethanol and water in a ratio of 1:1:1 with a few drops of a strong surfactant. They added some ammonia as well, which was said to greatly improve the test. Though effective, this may not be the most practical approach due to the volatility and cost of materials used. The following is a list of solvents that is not exhaustive. Many were excluded for lack of availability, extreme toxicity or other unreasonablities.

Acetone Fantastic degreaser. Moderately polar solvent. Grease can be skimmed off the top to reuse acetone. Wicking capabilities make acetone an excellent tool in bone degreasing. Drawbacks include volatility, explosiveness, availability, disposal concerns and higher cost.

Benzyl Alcohol Low toxicity, very polar in water but quite expensive.

Cyclohexane Fantastic degreaser. Non-polar. Expensive, Explosive, flammable and volatile. Do not heat. Products with this are likely available.

Ethanol Excellent degreaser of bone, highly polar solvent, but may cause staining and evaporates quickly.

Gasoline Strong degreaser that is readily available. Has some non-polar solvents. Drawbacks include volatility, high cost, flammability and explosiveness, lingering smell on the skull and disposal concerns.

Glycol Ethers Group of strong solvent degreasers which was tested by Bare Bones Solutions. Can be polar or non-polar. Some disposal and handling concerns because many are toxic. When considering use, propylene glycol is favorable for toxicity and ecological reasons. Propylene glycol is common in laundry detergents and is highly polar, making it easily miscible in water. Effective at miniscule concentrations of less than 0.5-1% and compatible with many surfactants and emulsifiers. They are known to be effective at cleaning bone (Stemmer and Kehagias, 2018). Propylene glycol may induce warping and other damage in bone. In an experimental mix which included glycol, ostrich skull elements warped and a pig skull became powdery.

Glycols remain a very interesting group that interact with collagen in useful ways. They are diverse and generally accessible.

Heptane Non-polar. Much success has been found using Heptane, having removed 30% of fats in just 21 days (Guilminot et al, 2014). Expensive, volatile, not miscible in water and toxic. Not recommended outside of proper laboratories. 

Hexane Non-polar. Very hard to source, very expensive with disposal issues. Recommended only for the highest priority and rarest specimens. Not recommended for casual use outside of a museum or laboratory.

T-butyl Alcohol Non-polar. Probably a decent degreaser of bone. Miscible in water but expensive and flammable.

Toluene Non-polar. Decent degreaser of bone. Available in lacquer thinner. Very flammable, expensive and has health risks. Not recommended.

Turpentine Low polarity. Moderate degreaser. Easy to source, price escalates fast, noxious. Not the best choice, but could be useful for smaller skulls if nothing else is available.

Several oxidizers have been popular in bone cleaning. They work by accepting electrons from another substance, causing the substance donating the electron to become oxidized. These include chlorine (bleach), peroxide and hydroxide (ammonia). Phosphates and sulphates can also be used. Along with the damage to proteins, some also accept calcium ions from the apatite of the bone in a process called chelation. Oxidizers damage proteins, including collagen in the bone.

Chlorine (Bleach) demineralizes bone (Kerbl et al, 2012) in a process that can continue for years. Oxidizes collagen.

Hydrogen Peroxide is the most common oxidizer and likely the only devoted oxidizer encountered in skull cleaning. Used ubiquitously to whiten bone in the last stage of cleaning and often to quickly (and poorly) degrease skulls in European mounts. Known to damage bone by forming oxygen radicals in the water-peroxide reaction which then oxidize collagen. It comes in many forms including liquid developer, various percentages of hydrogen peroxide as well as oxyclean. It is often debated which product is superior, but ultimately what matters is percentage, heat applied and time of exposure. As with any chemical, too much heat or exposure has increasingly deleterious effects on the bone.

Enzymes are catalysts, often proteins that speed up chemical reactions in organisms. They are successful in some bone preparation (Boyde and Firth, 2004, Bromage et al, 2011 and Irlicht et al, 2002). However, “using enzymatic detergent could be considered risky to collagen in bone” (pers. comm., Wallace, J.*). Detergents contain lipases, amylases and proteases which last approximately 18 hours in solution (pers. comm., detergent company^). A commonly used protease is subtilisin (Maurer, 2004). “Nearly ten” from this family of enzymes are known to damage type 1 collagen (Ran et al 2013). Piazza and Garcia (2014) and Fujimoto (1968) point out that the triple helix of collagen prevents or slow proteolysis. The rate of proteolysis was greater in subtilisins than trypsin (Piazza and Garcia, 2014). Their tests were just 48 hours and despite some of the collagen being derived from bone it was ground in meal and not in situ, creating a discrepancy between their results and degreasing whole bone which requires much longer exposure. Most other enzymes have great difficulty interacting with collagen.

Pronase, a non-specific protease used in some detergents, may also damage collagen rich in hydroxyproline, such as type 1 (Fujimoto, 1968) at thermal thresholds around 35 degrees Celsius. Therefore, if using an enzymatic detergent, a lower temperature may be ideal.

Trypsin is an enzyme used in diaphanization. Thomas and Seymour-Jones (1923) demonstrated that it causes “measurable hydrolysis of the collagen” and Liu et al (2018) claim that “collagen fibres were partially degraded into collagen fibrils by trypsin”. Burns and Meadow (2013) report success preparing bone with trypsin when paired with enzyme detergents.

Bromelin, Papain and other fruit derived enzymes, have also been used. These are added often during maceration. Papain degrades collagen (Wihastyoko et al, 2021) though it may have difficulty with hydroxyapatite-coated collagen.

Bare Bones Solutions has tested bone degreasing with pure lipase. Though some results were promising, cost comparisons and mixed results made these tests scale poorly.

*Joseph Wallace. Bone Biology and Mechanics Lab, Weldon School of Biomedical Engineering, Purdue University. Email to author, February 15, 2025.
^Detergent company. Phone call with author, April 2024.

Maceration is a useful flesh removal technique. Typically at Bare Bones this is done in 30-35 degrees Celsius water, with water changes occurring every 48-72 hours to limit the smell. Cold water applications are successful as well and reportedly much less odorous, but take significantly longer. This should be done ahead of chemical application. Ingredients common in most detergents, even enzymatic ones, interfere with bacterial growth (Moore, et al 2006 and Mosiichuk et al, 2025). If the goal is to form a bacterial culture to eat flesh off bone, this should be a consideration.

Light has had some success in degreasing (Horak et al, 2022). Their study found that macerated bone achieved a desired condition of degreasing in 21 days in sunlight. Several other forms of light were used, and all appear to have had some success.

Formation This waxy substance has been studied for over two centuries, and presents novel taphonomy and serious obstacles when skeletonizing dead animals. This material is stable and resilient, often more than collagen and fat, and therefore requires special consideration. It consists of saponified saturated fatty acids (O’Brien and Kuehner, 2007 and Ubelaker and Zarenko, 2011) which may require high pH environments under specific conditions to form (Vass, 2001). In particular, these are myristic acid, palmitic acid, and stearic acid. The ideal pH for formation may be lower than high end degreasers, but still mildly alkaline (Ubelaker, 2023), and require anaerobic conditions at 21-45 degrees Celsius for optimal enzymatic release and bacterial growth (Forbes et al, 2011). Schoenen and Schoenen (2013) report that a lack of both oxygen and “bacterial degradation” is key to adipocere formation. Excellent summaries can be found on google scholar in Forbes et al, 2004, Takatori, 2001, Ubelaker and Zerenko, 2011 and others.

Magni et al (2021) makes a distinction between saponification and adipocere formation, and describes the chemical process in detail. They also indicate Bacillus subtilis, Micrococcus luteus, Staphylococcus aureus, some Pseudomonas species and Clostridium perfringens may be involved. B. subtilis generates subtilisin enzymes for cleaners, and C. perfringens can survive in higher pH environments and has a varied diet but prefers glucose. It may be possible to limit adipocere growth by controlling bacterial food sources or growth.

Glucose may be important in adipocere formation because of its role as bacterial food and presence in bone marrow. There may be a different glucose level in marrow than ambient blood levels (Suchacki et al, 2020) due to varying “insulin responsiveness” in different kinds of marrow tissues. Varying diets, such as those in canids and ursids compared to others in the carnivora, should cause the composition of bone marrow tissues to differ (Dzubanova et al, 2024). This may explain the observed tendency of canids and ursids to develop adipocere compared to other carnovores.

Removal There have been successful reports of some products removing adipocere (Obermeyer et al, 2011). It seems likely that any sufficiently powerful degreasing agent may remove adipocere well. Adipocere naturally forms in high pH environments through saponification. Ammonia is indicated as a remover of soap scum, and may function as a final wash to clean up adipocere residue formed during the process.

The melting point of adipocere is reported as 71 degrees Celsius by Ruttan and Marshall (1917). Melting it without damaging collagen may not be possible because the uppermost heat breakpoint of collagen may only be as high, at maximum, as approximately 61 degrees Celsius (Danielsen et al, 1994, reported shrinking in human pelvises at that temperature). It is well known in art communities that oil melts wax. With a very small amount of olive oil (triglyceride) and a stiff bristle brush, the majority of adipocere was successfully removed from a canid skull in a test. This miniscule amount of oil (less than half of one teaspoon) mostly washed away with soapy water, and degreased away with ease.

Aggressive mechanical means of removal such as sandpaper are discouraged.

There is a belief that heating grease will cause it to move “deeper into the bone” and make it harder to degrease. That grease can be moved through dense cortical bone indicates this hypothesis may be incorrect because “deeper grease” should still be movable through bone. This ability may be interfered with by capillary action. An alternative hypothesis is that the triglycerides are crystalizing post-heating. According to Gunjan et al., 2024, slow-cooled triglycerides required ten times the amount of time to degrease successfully with a surfactant. Bone is a decent insulator, and any grease inside it may cool slower as a result. When they cool slower, the crystals are bigger and the lattice is more organized, making it harder to degrease.

Tristearin, a long-chain (saturated) triglyceride, has three or four plateaus as heat changes where crystallization can occur (Hoerr and Harwood, 1956). The lowermost melting point of this triglyceride is 55 degrees Celsius, which exceeds collagen’s comfort temperature. When excessive heat is applied, these crystallization plateaus may be reached. In highly polar solvents, solubility of tristearin limited. In non-polar solvents, solubility is considerably higher. These observations are consistent with grease behavior reported by skull preparators when dealing with skulls with an excessive thermal history. This author hypothesizes that non-polar solvents will likely be more successful than surfactants in handling most crystallized grease.

Triglycerides oxidize in heat. This can also happen during rot in a process called rancidification. There are several relevant products from this process. One is fatty acids, some of which may contribute to grave wax production. Free radicals which interact negatively with collagen may occur (Geng et al, 2023). Oxidized fats can turn dark, and drop in pH as they form carboxylic acid. Other products include aldehydes and carbonyls.

In bone it may be possible for grease to wick and then oxidize, creating a complicated degreasing scenario. Some soaps may find it difficult to dislodge oxidized greases. Oxidizers such as peroxide may also have issues moving these materials because they are already oxidized.  Oxidized greases are typically more difficult to move because their polarity has increased, altering their attraction to some surfaces. They may still react to saponifiers. The typical method of removing epoxies or oxidized greases is solvents.

Rarely, a polymerization of the fats can occur, creating a type of monomeric oxidized epoxy (Khor et al, 2019). These epoxies are likely uncommon in bone cleaning. Epoxies are generally considered one of the better glues for highly porous substrates. Triglyceride epoxies are known to form at or above 160 degrees Celsius (Khor et al, 2019) and provide another scenario for difficult to remove grease post-heat exposure. In a boiling or simmering pot, where bones are at the bottom closest to the heating element, it may be the case that bones are exposed to heat in excess of what is measured. Some epoxification may occur prior to 160 degrees Celsius.

Recommended action is soaking in a non-polar solvent and heat not in excess of recommended guidelines for the animal in question. Unheated acetone may be a great option because of superior wicking and an altered charge on the oxidized triglyceride. Isopropyl alcohol may be useful on oxidized grease.

See “heating collagen” section for interactions with gelatin formation.

Skulls can be compromised in a number of ways. Most typical are high heat, ultraviolet or chemical exposures. In all three cases, collagen has been damaged. In chemical exposure, the bone mineral may be damaged. In this case, pitting, cracking or fraying may occur. This is not typical in bone prep, as common knowledge is to avoid low pH environments which are damaging to bone. Instances where blood stains persist do occasionally require the use of products which use low pH ingredients.

In cases of compromised collagen, flaking, powdery texture, cracking and eventual disintegration are not uncommon. Heat above 44-48 degrees Celsius for adult mammals will produce reliable results of bone shrinkage, warpage, and previously listed damage. Ultraviolet and other environment exposures gives a classic “field texture” to bone which can have its own charm.

When a bone is failing, consolidating the hydroxyapatite with reversible methods is required. The standard is to use acrylic resins such a Paraloid b-72, or other polymers which can bond the brittle bone together. In the past the Archaeology Survey of Alberta has used water diluted white glue to consolidate bone. There are many other polymers or other materials which may be effective at consolidating bone such as epoxy.

Sealing a skull with an acrylic spray is not effective. The bone continues to decay under the spray, and will still fall apart. It should be consolidated first. A skull bleached with chlorine may require that the chlorine be neutralized somehow to stop the progression of the chlorine ions.

Bone degreasing has many options and there is no silver bullet or universal technique. Online resources are disparate, discombobulated, amateur and largely unreferenced. It is the hope that this information may remedy some of these issues and begin to organize some of the information in a more formal and accessible way. Please reach out if there are errors.

Al-Shaer, A., and Forde, N. (2025). Decoding collagen’s thermally induced unfolding and refolding. Biochemistry, Biophysics and Computational Biology. 122(20). doi: 10.1073/pnas2420308122

Alers, J.C., Krijtenburg, P.J., Vissers, K.J. and van Dekken, H. (1999). Effect of bone decalcification procedures on DNA in situ hybridization and comparative genomic hybridization. EDTA is highly preferable to a routinely used acid decalcifier. Journal of Histochemistry and Cytochemistry. 47(5), 703-10. doi: 10.1177/002215549904700512

Asghar, A, and Henrickson, R.L. (1982) Chemical, biochemical, functional, and nutritional characteristics of collagen in food systems. Advances in Food Research. 28, 231-372. doi: 10.1016/s0065-2628(08)60113-5. PMID: 6762058.

Boman, A. and Maibach, H.I. (2000). Percutaneous absorption of organic solvents. Int J Occup Environ Health, 6(2), 93-5. doi: 10.1179/oeh.2000.6.2.93

Bosch, P., Aleman, A., Moreno-Castilla, C., and Botella, M. (2011). Boiled versus unboiled: a study on neolithic and contemporary human bones. Journal of Archaeological Science. 38(10), 2561-2570. doi: 10.1016/j.jas.2011.04.019

Boyde, A. and Firth, E.C. (2004). Articular calcified cartilage canals in the third metacarpal bone of 2-year-old thoroughbred racehorses. Journal of Anatomy, 205(6), 491-500. doi: 10.1111/j.0021-8782.2004.00354

Bromage, T.G., Juwayeyi, Y.M., Smolyar, I, Hu, B, Gomez, S, Chisi, J. (2011). Enamel-calibrated lamellar bone reveals long period growth rate variability in humans. Cells Tissues Organs, 194(2-4), 124-30. doi: 10.1159/000324216

Burns, P. and Meadow, R.H. (2013). The use of trypsin to prepare skeletal material for comparative collections with a focus on fish. Archaeofauna, 22. 29-36

Burton, B., Gaspar, A., Josey, D., Tupy, J., Grynpas, M. D., And Willett, T. L. (2014). Bone embrittlement and collagen modifications due to high-dose gamma-irradiation sterilization. Bone. 61, 71-81. doi: 10.1016/j.bone.2014.01.006

Danielsen, C.C., Mosekilde, L., Bollerslev, J., and Mosekilde, L. (1994). Thermal stability of cortical bone collagen in relation to age in normal individuals and in individuals with osteopetrosis. Bone, 15(1), 91–96. doi: 10.1016/8756-3282(94)90897-4 

Dzubanova, M., Benova, A., Ferencakova, M., Coupeau, R. and Tencerova, M. (2024). Nutrition and bone marrow adiposity in relation to bone heath. Physiological Research. 73(1). 107-138. doi: 10.33549/physiolres.935293

Eriksen, E.F., Charles, P., Melsen, F., Mosekilde, L., Risteli, L., and Risteli, J. (2009). Serum markers of type I collagen formation and degradation in metabolic bone disease: Correlation with bone histomorphometry. Journal of Bone and Mineral Research, 8(2), 127–132. doi:10.1002/jbmr.5650080202

Forbes, S.L., Stuart, B., Dadour, I., and Dent, B. (2004). A Preliminary Investigation of the Stages of Adipocere Formation. ASTM International. J. Forensic Sci.,49(3). doi: 10.1520/JFS2002230

Forbes, S.L., Wilson, M.E. and Stuart B.H. (2011). Examination of adipocere formation in a cold water environment. Int J Legal Med. 125(5):643-50. doi: 10.1007/s00414-010-0460-6

Fujimoto, D. (1968). Correlation between pronase susceptibility and hydroxy proline content of collagen fiber. Biochimica et Biophyica Acta (BBA) - Protein Structure. 154(1); 22, 183-189

Geng, L., Liu, K., and Zhang, H. (2023). Lipid oxidation in foods and its implication on proteins. Frontiers in Nutrition. 15(10). doi: 10.3389/fnut.2023.1192199

Guilminot, E., Lemoine, G., Pelé, G., Poisson, L., Surbled, M., Louvet, I., Mevellec, J.-Y., and Rémy, L. (2014). Re-treatment of whale bones – How to extract degraded fats from weakened bones? Journal of Cultural Heritage, 15(2), 128-135. doi: 10.1016/j.culher.2013.03.008

Gunjan, T., Torquato, L. M. G., Ahmad, Z., Fong, R., and Cabral, J. T. (2024). How triaclyglycerol thermal history impacts film removal by surfactant solution. Journal of Colloid and Interface Science, 670 (15), 540-549. doi: 10.1016/j/jcis.2024.05.069

Horak, O., Pyszko, M., Paral, M., and Sandor, O. (2022). Degreasing and bleaching bones using light sources as a tool to increase the safety of teaching osteology at the university of veterinary science, Brno. Peer J. doi: 10.7717/peerj.14036

Hoerr, C.W and Harwood, H.J. (1956). The solubility of tristearin in organic solvents. The Journal of Physical Chemistry. 60(9), 1265-1269. doi: 10.1021/j150543a028

Irlicht, R.A, O’Connell, S.L., Thomas, C.D.L. and Clement, J.G. (2002). The effect of Terg-a-zyme on the breaking strength of bone. ANZBMS 12th Annual Scientific Meeting. Stamford Grand, Adelaide, Australia.

Kerbl, F.M., DeVilliers, P., Litaker, M., and Eleazer, P.D. (2012). Physical effects of sodium hypochlorite on bone: an ex vivo study. Journal of Endodontics. 38(5), 357-359. doi: 10.1016/j.joen.2011.12.031

Kohr, Y. P., Khai, S. H., Abas, F., Cheong, L.Z, Nehdi, I. A., Sbihi, M.H, Gewik, M.M., and Tan, C.P. (2019). Oxidation and polymerization of triacylglycerols: in-depth investigations towards the impact of heating profiles. Foods. 8(10), 475. doi: 10.3390/foods8100475

Lambri, L.L., Bozzano, P. B., Giordano, E. D. V., Bonifacich, F. G., Gargicevich, D., Zelada, G. I. and Lambri, O. A. (2018). Denaturation processes of collagen from cow bones as a function of temperature. Materia, Rio J, 20(2)

Leikina, E., Mertts, M.V., Kuznetsova, and Leikin, S. (2002). Type 1 collagen is thermally unstable at body temperature. Biophysics and Computational Biology. 99(3), 1314-1318. doi: 10.1073/pnas.032307099

Liu, Z-q., Tuo, F-y., Song, L., Liu, Y-x., Dong, X-p., Li, D-m., Zhou, D-y. and Shahidi, F. (2018). Action of trypsin on structural changes of collagen fibres from sea cucumber (Stichopus japonicus). Food Chemistry. doi: 10.1016/j.foodchem.2018.02.117

Maurer, K-H. (2004). Detergent proteases. Current Opinion in Biotechnology.15 (4), 330-334. doi: 10.1016/j/copbio.2004.06.005

Magni, P.A., Lawn, J. and Guareshi, E. E. (2021). A practical review of adipocere: key findings, case studies and operational considerations from crime scene autopsy. Journal of Forensic and Legal Medicine. 78. doi: 10.1016/j.jflm.2020.102109

Maximo, G.J. and Cunha, R.L. (2010). Mechanical properties of collagen fiber and powder gels. Journal of Texture Studies. 41. 842-862. doi: 10.1111/j.1745-4603.2010.00258.x

Miles, C., and Ghelashvili, M. (1999). Polymer-in-a-Box mechanism for the thermal stabilization of collagen in fibers. Biophysical Journal. 76(6), 3243-3253. doi: 10.1016/S0006-3495(99)77476-X

Moore, S.L., Denyer, S.P., Hanlon, G.W., Olliff, C.J., Lansley, A.B., Rabone, K. and Jones, M. (2006). Alcohol ethoxylates mediate their bacteriostatic effect by altering the cell membrane of Eschericheria coli NCTC 8196. International Journal of Antimicrobial Agents. 28(6), 503-513. doi: 10.1016/j/ijantimicag.2006..08.023

Mosiichuk, N., Enggren, G., Da Silva, Z.L., Morén, A.K., Hansson, H., Engblom, J. and Ruzgas. T. Antibacterial activity of propylene glycol against Staphylococcus aureus and Staphylococcus epidermidis in neutral and mild acidic conditions. Journal of Antimicrobial Chemotherapy. 80(7), 1947-1950. doi: 10.1093/jac/dkaf153

Nandi, P. K., Grant, M. E. and Robinson, D. R. (1985). Destabilization of collagen structure by amides and detergents in solution. International Journal of Peptide and Protein Research. 25(2), 206-12. doi: 10.1111/j.1399-3011.1985.tb02166.x

Nyman, S., Roy, A., Shen, X., Acuna, R.L., Tyler, J.H., Wang,. X. (2007). The influence of water removal on the strength and toughness of corticol bone. Journal of Biomechanics. 39(5), 931-938. doi: 10.1016/j.jbiomech.2005.01.012

O’Brien, T.G. and Kuehner, A.C. (2007). Waxing grave about adipocere: soft tissue change in an aquatic context. J. Forensic Sci.52, 294–301

Obermeyer, C., Finnegan, F. and Salem. S. (2011). Adipocere Removal from Human Skeletal Remains. Poster, Chemistry Department, Washburn University, Topeka, Kansas

Piazza, G.J., and Garcia, R. A. (2014). Proteolysis of meat and bone meal to increase utilisation. Animal production Science.54, 200-206. doi: 10.1071/AN13041

Ran, L.-Y., Su, H.-N., Zhao, G.-Y., Gao, X., Zhou, M.-Y., Wang, P., Zhao, H.-L., Xie, B.-B., Zhang, X.,Y., Chen, X.-L., Zhou, B.-C., and Zhang, Y.-Z. (2013). Structural and mechanistic insights into collagen degradation by a bacterial collagenolytic serine protease in the substilisin family. Molecular Microbiology. 90(5), 997-110. doi: 10.1111/mmi.12412

Ricard-Blum, S. (2011). The collagen family. Cold Spring Harbor Perspectives in Biology. Eds. R. Hynes and K. Yamada. 1-20. doi: 10.1101/cshperspect.a004978

Ruttan, R.F. and Marshall, M.J. (1917). The composition of adipocere. The Journal of Biological Chemistry. 29(2), 319-327

Schoenen, D., and Schoenen, H. (2013). Adipocere formation- The result of insufficient microbial degradation. Forensic Science International. 226 (1-3). doi: 10.1016/j/forsciint.2013.01.023

Shehata, T.P. and Krap, T. (2024). An overview of the heat-induced changes of the chemical composition of bone from fresh to calcined. International Journal of Legal Medicine, 138, 1039-1053. doi: 10.1007/s00414-024-03160-z

Stemmer, D. and Kehagias, O. (2018). Finding a new solution to an old problem. Biodiversity Information and Science Standards. Sofia. doi: 10.3897/biss2.26392

Suchacki, K.J., Tavares, A.A.S., Mattiucci, D., Scheller, E.L., Papanastasiou, G., Gray, G., Sinton, M.C., Ramage, L.E., McDougald, W.A., Lovdel, A., Sulston, R.J., Thomas, B.J., Nicholson, B.M., Drake, A.J., Alcaide-Corral, C.J., Said, D., Poloni, A., Cinti, S., Macpherson, G.J., Dweck, M.R., Andrews, J.P.M., Williams, M.C., Wallace, R.J., van Beek, E.J.R., MacDougald, O.A., Morton, N.M., Stimson, R.H. and Cawthorn, W.P. (2020). Bone marrow adipose tissue is a unique adipose subtype with distinct roles in glucose homeostasis. Nature Communications. 11(3097). doi: 10.1038/s41467-020-16878-2

Takatori, T. (2001). The mechanism of human adipocere formation. Legal Medicine. 3(4), 193-204. doi: 10.1016/S1344-6223(01)00036-0

Thomas A.W. and Seymour-Jones F.L. (1923). The hydrolysis of collagen by trypsin. Proceedings of the Society for Experimental Biology and Medicine. 20(8), 433-434. doi:10.3181/00379727-20-215

Turner-Walker, G. (2012). The removal of fatty residues from a collection of historic whale skeletons in Bergen: an aqueous approach to degreasing. doi: 10.13140/2.1.1996.8969

Ubelaker, D.H. and Zarenko, K.M. (2011). Adipocere: what is known after over two centuries of research? Forensic Science International. 208(1-3), 167-172. doi: 10.1016/j.forsciint.2010.11.024

Ubelaker, D.H. (2023). Postmortem inteverval.Encyclopedia of forensic sciences, third edition. Ed M. M. Houck. Elsevier. 241-245. doi: 10.1016/B978-0-12-823677-2.00003-9

Van Heiningen, A.R.P., Kang, G.J. and Ni. Y. (1999). Addition of sodium silicate and chelant to the FAS stage to blach recycled fibers. Pupling Conference Proceedings.

Vass, A.A. (2001). Beyond the grave – understanding human decomposition. Microbiology Today. 28, 190-192

Venkitesan, A.C., Muthulakshmi, Y.P, Lobo, N.P, Raja, N.K., Balamurugan, K. and Fathima, N.N. (2026). Synergistic effects of glycerol on the hydration dynamics and rehological properties of collagen. Biophysical Chemistry. 333. doi: 10.1016/j.bpc.2026.107600

Viguet-Carrin, S., Garnero, P., and Delmas, P. D. (2005). The role of collagen in bone strength. Osteoporosis international. 17(3), 319-336. doi: 10.1007/soo198-005-2035-9

White, E.M., and Hannus, L.A. (1983) Chemical weathering of bone in archaeological soils. American Antiquity. 48, 316-322.

Wihastyoko, Y.L., Soeharto, S., Widjajanto, E., Handono, K., and Pardjianto, B. (2021). Does papain enzyme improve collagen degradation? Systematic Reviews in Pharmacy. 12(3), 676-684.