The Production And Use Of Gelatin

Fruit slices using gelatin for the gel structure.
Image by _Alicja_ from Pixabay

Gelatin has been an ingredient for general product development for many years. It is a protein prepared from collagen which is found in ligaments and tendons, skin and in other types of connective tissue.

Gelatin is prepared by boiling up bones, skin and other connective tissues from mainly animals such as pigs and cows. It is one of the main byproducts from animal meat processing.

Whilst the principal sources are still animal skins fish gelatin is increasingly seen as a competing source material. There are no vegetarian versions of collagen or indeed gelatin that have any of the critical properties possessed by gelatin.

Insect derived gelatins might be a possible alternative source. The desert locust, Aspongopus viduatus (melon bug) and Agonoscelis pubescens (sorghum bug) are all potential sources.

Considerable effort is being expended to create a plant based derivative that mimics this protein’s properties but research has not yet succeeded in this endeavour.

There are a number of web-site that conflate the term gelatin with other ingredients that create gels. It should never be confused with agar agar, carrageenan or even xanthan gum.

General Uses

Gelatin forms strong, clear and transparent films as well as gels. Whilst there are attempts to look for alternatives gelatin still retains its commercial power which few other large polymers are able to match.

As a material it is biocompatible with many other human tissues, is commercially available in large quantities and is widely used in the pharmaceutical industry and medical fields as well as in food. Notable examples in the medical field include vascular prosthesis (Marois et al., 1995), as carriers for drug delivery and as dressings for wound healing.

It is dispersible and highly soluble in water and makes a good composite material both in photography, food processing and range of other processing industries.

Structure Of Gelatin

Gelatin is a mix of large complex polypeptides but all have the same amino acid composition as the parent material collagen. It has a broad weight distribution range. The similarities between gelatin as with collagen from different species is highly conserved at the primary structure level.

The main differences occur in the secondary structure between collagen from different species.

In collagen, the proteins are arranged in a rod with a triple-helix structure of two identical chains called α1 and a slightly different chain called α2. In the gelatin manufacturing process the chains are partly separated and hydrolysed which reduces them in size. The average molecular weight ranges from 20 kDaltons through to 250 kDaltons.

Gelatin is not polydispersed completely. It has a definite molecular weight distribution pattern, which corresponds to the α-chain and its oligomers (Buice et al., 1995).

The Conversion Of Collagen To Gelatin

Collagen is converted to gelatin during its manufacture. Collagen starts off as a highly organized mass of fibres which are wholly insoluble in water. Processing gradually reduce the size of the collagen chains until it so depolymerized that the material now gelatin is soluble in water. Most conversion processes rely on acid or alkali catalysed conversion.

Enzyme hydrolysis has been used using collagenases but this more expensive and less controllable.

Thermal denaturation requires heating the collagen found in skin or bone at around 40ºC.

Manufacture Of Gelatin

Pretreatment And Preparative Steps

The basic process of manufacture is still largely unchanged since it was developed in the 1920s. in the post-war years improvements in separation technology involving cross-flow membrane processing have helped as has ion-exchange technology.  There are two types of process which yield either type A or type B gelatin.

The source material for gelatin is usually bones and skins from animals. 

Generally, dilute mineral acid solutions are used to solubilise and extract calcium  so that the bones and skin are rendered pliable and easier to handle.

In some cases just hot water and solvents are needed to solubilise fat. Fat is a major contaminant of gelatin preparation and needs to be reduced to less than 1 per cent before it proceeds to the main extraction step. 

Manufacture Of Type A Gelatin

Type A gelatin is manufactured from pig (pork) skin with grease as a major byproduct. In Europe, 80% of the gelatin is type A and is prepared from pork skin and this method is now more accepted. The remainder for this process will come from sheepskins and mature cattle skins. The age of the animal is critical in the production of any type of gelatin but more so in the acid process recommended for type A manufacture.  For cattle skin, animals of 2 to 3 years old are preferred. Pigskins from animals less than 18 months old produce higher gelatin yields than skins from 30 month old animals.

The process is preferred for the presence of less cross-linked gelatin. It is preferred too because it is much shorter a time of processing.

Pigskin is usually received frozen. The only pretreatment needed is to thaw it, then finely chop and macerate the pieces further.  There is rarely any need to remove fat at this stage because separation will occur during the acid extraction step. The skin is washed to remove extraneous matter such as hair.

Alkali pretreatment is not an option because of the potential for soap formation. 

In some instances the skin is degreased and demineralised before any pretreatment takes place.

Acid conditioning and extraction produces a gelatin with an isoelectric point well above pH 5. The proporotion of glutamine and asparagine residues remains in their amide form. Titration curves have shown that peptide bond hydrolysis using acid treatment is more significant than alkali treatment (discussed below).

The skin is fully hydrated for between 10 and 48 hours in very dilute acid solutions such as 1 to 5 per cent hydrochloric, phosphoric or sulphuric acid. The pH varies from 3.5 to 4.5 and usually starts off at a low temperature of about 15ºC. This acid treatment is halted once the raw material has reached maximum swelling or has become fully acidified.

-Acid Extraction Process

Acid extraction is usually performed on material which has not been alkali treated but will have probably received some simple acid conditioning.

Most extraction this way requires a pH of 3.0 to 3.5 although lower pHs have been tried. The objective in any extraction is to recover the gelatin as rapidly as possible from the ossein or other material so that it is not hydrolysed further by thermal degradation.

Most extractions have moved from batch to semi-continuous processing. In all cases hot water supplied at a defined temperature and flow-rate is poured onto the animal material. 

At least 4 to 5 extractions are needed with increasing temperature from 55 to 65ºC in the first extraction to a final temperature close to boiling water of 95 to 100ºC.

Each extraction takes between 4 and 5 hours.

The grease (fat) and any fine fibres are removed at each step and the gelatin solution filtered and then deionised.

The next step is concentration which we discuss below. 

The Manufacture Of Type B Gelatin

The main source of gelatin here is bone but also pork and cow skin. About 15% of Europe’s edible gelatin comes from what is known as split. Split is the thin collagen-containing layer of the cattle hide that lies between the epidermis and the subcutaneous layer.

In this process, bones are crushed and degreased at the abbatoir or a rendering facility. the rendered bone pieces have a size between 0.5 and 4cm.  These fragments are treated with 4 to 7 per cent hydrochloric acid in a steeping process lasting between 4 and 14 days to remove calcium as described above in the general pretreatment step.

One of the byproducts of this type of treatment is the release of calcium in the form of dibasic calcium phosphate. This is precipitated and removed from the spent liquor.

– Degreasing Process

Degreasing is a necessary step in the pretreatment of bone before gelatin extraction. Bone itself is 35% water and 15% fat. Fat removal is critical because its presence causes problems later on in processing. Liming for example leads to the production of soaps which then clog capillary pores and spaces when bone is demineralised to produce ossein. During extraction, fat will emulsify which clouds the extraction liquor and reduces filtration especially when cross-flow filtration is used. Fat presence in gelatin also produces non-uniform wettability especially in hard-gel capsules.

Degreasing relies on mechanical agitation of fragmented bone with hot water. It is largely a continuous processes with bone and water entering at one end and treated bone with a dat emulsion in water leaving at the other. Bone fragments and fat are separated.

The bone is then further treated with hot water to remove soft tissue followed by hot air drying.

The fat content is then less than 3% after this phase.

Demineralisation

 Demineralization is key pretreatment following degreasing.  The bone needs to be treated so that collagen can be released for conversion to gelatin. Dilute hydrochloric acid is the treatment of choice using a counter-current method.

– The Liming Process

Liming is a conditioning process to help convert collagen to gelatin. The main purpose is to destroy as many cross-links as possible so that gelatin polymers can be created and released.

The amide groups of glutamine and asparagine amino acids are converted to carboxyl groups and large quantities of ammonia are released. The isoelectric point of collagen also alters. Gelatin produced by this method has an isoelectric point of pH 4.8 to 5.2. It also means that Type B gelatin has a higher carboxylic acid content than Type A gelatin.

Arginine is degaunidated to form ornithine and urea. This is a slower reaction which explains the reason for the long steeping process in lime solution. There is also some reduction in the protein polymer lengths caused by alkaline catalysed hydrolysis of peptide bonds adjacent to glycine, serine, threonine, aspartic acid ad glutamic acid.

In terms of cross-links, some but not all are destroyed in the liming process otherwise gelatin would not exist and the less destructive a process, the higher the number of high molecular weight chains. The impact is that the less the impact of liming the greater the likelihood of generating gelatin for higher strength gels.

In some cases, acidification of collagen with say 9% hydrochloric acid helps reduce the liming period. This step may well have been introduced as referenced to earlier with type A gelatin production.

Liming also helps remove protein contaminants which become soluble at alkaline pHs. The other change occurring to collagen is swelling. The internal adhesion of the fibrils is reduced by rupture of certain intermolecular links. Swelling occurs in just two phases: an initial and rapid equilibrium swell is completed after three days and then a slower swelling which suggests structures are breaking down.   

The process is as follows: demineralised bones known as ossein are washed before transference to a large steeping tanks or pits where they are treated in a lime slurry. A slaked lime concentration of between 2 and 5% is needed.

Gentle agitation helps release the gelatin over a 3 to 16 week period.

The type B gelatin is washed to remove all lime and takes 15 to 30 hours. Washing is usually conducted in a log-washer or paddle-washer with water. Much of the lime on the surface of the tank is mechanically removed and may be reused if it is still viable. The material which is washed will still have a pH of between 9 and 10.

-Neutral Extraction

The ossien as it is known or other raw material  is acidified as part of a neutralization step to between pH 5 and 7 using mineral acid (hydrochloric, phosphoric and sulphuric acids). There are instances where it is over acidified and then rewashed to bring the pH back up. Over-acidification can excessively swell the collagen and make it too fragile. The pH of the solution needs to be in the range 5.0 to 6.5.

 Here a series of extractions starting with an initial temperature of 50 to 60ºC and rising to boiling point is needed. As with acid extraction, 4 or 5 steps are needed.

Similar to the previous process, the first fraction of gelatin is obtained through thermal degradation of the hydrogen bonds between collagen molecules. All later extracts are a culmination of collagen breakdown.

Neutral extraction produces gelatin of a lower gel strength than those obtained from acid extraction. The highest viscosity gelatin is obtained from neutral extraction processes of collagen that had previously been limed.

Concentration Of Gelatin

Common to both processes. Any thermal input produces degradation of the gelatin so keeping evaporation to a minimum is paramount.

A concentration step to between 20 and 40 % w/w is needed using continuous vacuum evaporation. A variety of continuous evaporators are available from tubular types to centrifugal versions. Plate and frame types are still the most popular.

It is feasible to concentrate the liquor to a level of between 20 and 25% gelatin below 55ºC in the final effect.

Localised overheating is an issue for high viscosity gelatin solutions.

Sterilization

No preservatives other than some sulphur dioxide is permitted.

The final viscous solution is cooled and then extruded into fine strings passing through successive temperature changes. Air is used to cool the mass.

The gelatin is ground up and blended to complete the specification.

Extrusion Of Gelatin

Extrusion is a widely used technology in all forms of processing from snack manufacture through to meat processing.

Gelatin manufacture has widely improved since the introduction of this technology to produce widely uniform pieces and films for further processing.

The extrusion relies mostly on a form of twin-screw, co-rotating extruders. The process usually operates in this situation at screw speeds between 100 and 400 rpm  with a processing temperature of between 90 and 120ºC.

One or two studies have been used to characterise the process of extrusion. As the screw speed is increased up to 300 rpm, the tensile strength (TS) increased and the water vapour permeability (WVP) reduced (Nur Hanani et al., 2012). In this study, the WVP in all the gelatin types examined dropped at screw speeds above 300 rpm.

Increasing the screw speed for extrusion improved the solubility of the films when dissolved in water.

Types Of Commercial Gelatin

Grades of gelatin exist to help the food producer with particular types of quality for various applications. In the food product development world, the grades are usually quoted as bronze, silver, gold and platinum. These grades are linked to their ‘bloom strength’ which relates to their ability to form and set a gel. On a weight basis, the more expensive the ‘metal’ description the higher the gel strength. The “bloom strength” for each grade is:-

  • Titanium: less than 125
  • Bronze: 125-135
  • Silver: 160
  • Gold: 190-220
  • Platinum: 235-265

Professional chefs have at their disposal sheets, granules or powder. The best dispersability is obtained with powders although wettability is an important characteristic and sheets very often preserve their popularity because they store more effectively. Gelatin powders have a tendency to pick up water especially if stored poorly in a high humidity environment.

Sheet weights usually reflect the equivalent gel strength. The following illustrates the differences on a weight basis per sheet:-

1 Platinum sheet = 1.75g

1 Gold = 2g

1 Silver = 2.5g

1 Bronze = 3.5g

1 Titanium = 5g

Most sources such as pig provide types described as pork gelatin 240 Bloom. This is a type A where a 240 Bloom is much stronger or has a higher tensile strength than those sold commercially in grocers. It is often used in dessert products or as a stabilizer and thickener.

Gelatin powder is usually soaked in three to four times its own weight of cold water for 30 minutes. It is then heated to just below boiling. Boiling however destroys the ability of the gelatin to form a gel.

Gelatin Gelling

 Gelatin forms gels in a manner similar to carbohydrates. It relies on a microstructural network. A concentration as low as 1.0% is all that is needed to form a thermoreversible gel. 

Gelatin is readily soluble in water because it can form hydrogen bonds with water molecules at its many exposed polar regions. On binding water, it swells and absorbs more water. the level of swelling is between 5 and 10 times its own volume of water.  Heating readily improves the dispersion process.

Gel forming begins when the swollen gelatin is heated above its melting temperature in solution and starts forming a basic gel structure on cooling. This is a sol-gel conversion which is reversible and in most cases is easily repeatable. Having a gel melting temperature around 30ºC means it is ideal for capsules and food. Gels at this temperature for example melt in the mouth which is highly desirable from a sensory texture perspective.

Gel formation is endothermic and its formation is a slower process but at roughly the same rate as pectin gel formation. Increasing the amount of gelatin in solution not only increases the rate of gelling but also the level of firmness.

The mechanism of gelation is called random coiled helix conversion. The imino acid-rich regions from different parts of the gelatin polypeptide chain form potential junction zones. Above gelling temperature, these zones are free to move without annealing but on cooling, they take up a helical conformation which is stabilized by hydrogen-bonding. This produces a three-dimensional gel (Nishimoto et al., 2005). This structure is most likely a form of the triple collagen‐like helix. It resembles a partial reformation as in collagen where the reformed parts act as junction zones in the gel.

Gel Strength Or Bloom Strength

The Bloom strength is a measure of gel rigidity and was devised by its eponymous inventor in 1925. It is an arbitrary test method that has little bearing on material required for say capsule manufacture. It relies on piston penetration of a defined gel.

Bloom strength It is determined by preparing a standard gel (6.66% w/v) and maturing it at 10 °C. It is defined as the load in grams required to push a standard flat-bottomed plunger just 4 mm into the gel. The gelatin used in hard capsule manufacture is of a higher Bloom strength (200–250 g) than that used for soft capsules (150 g) because a more rigid film is required for the manufacturing process.

The AOAC uses a different style of plunger to that identified in the British Standard. the former style plunger from the AOAC is now the accepted version.

Gelatin Gel Colour

The colour of gelatin gels is mostly influenced by the source and processing method. Colour is usually assessed visually and compared with standard gels of defined colour or artificial samples depicting the gel. Colour is expressed in Hellige units and varies from 1.5 which is pale yellow to 14 (brown).

Opaqueness can be assessed using turbidity methods as well as colourimeters to measure transmittance.

Turbidity

Turbidity is measured as for other food materials such as beer in nephelometric turbidity units (NTUs). The clarity of a gelatin solution depends on extraction and the processing occuring afterwards. 

Early extractions contains the best quality gelatin and produce gels of very high quality and clarity in particular. Later extractions produce gelatin gels of increasing degrees of turbidity with higher levels of colouring intensity.

In some processing, the gelatin must be clarified, bleached and filtered to obtain a better quality gelatin for gel work.

Gelatin In Food Use

Gelatin is used emulsified meats and jellied meat products at levels ranging from 3 to 15%, but more typically from 0.5 to 3%. It is also used in canned meat products such as hams, loaves, frankfurters, and cured, canned pork, to hold juices lost during cooking and to provide a good heat transfer medium during cooking (Sams, 2001).

Characterisation of Gelatin Films

Gelatin films make ideal edible structures for use in pharmaceuticals (Gennadios et al., 1994). Films have also been suggested for coating fresh meat products to extend their shelf-life. gelatin coatings act as barriers to further moisture movement and to oxygen ingress (Krochta and de Mulder-Johnson, 1997; Mendis et al., 2005).

These films are defined by their level of solubility in water. The other key features are tensile strength (TS) and water vapour permeability (WVP).  

Gelatin For Hard Capsules

The pharmaceutical industry requires gelatin for hard capsules of a particular quality because it needs to dissolve in the gut uniformly.

In this technology, a hard capsule is prepared by coating a stainless steel pin with a gelatin solution that forms in a few seconds. The gelatin solution is 30% by weight and the hard capsules is formed on cooling from 50 to 25ºC very rapidly.

Soft-gels are formed in a rotary die process. A concentrated gelatin solution is rapidly gelled on a chilled drum. Plasticizers such as glycerol are added to help produce a flexible structure.

Shelf-life studies on gelatin capsules derived from pig skin that has been processed at different plants have different properties. It reveals that production methods which largely remain confidential have a typical bearing on the quality of material and careful quality control is needed to demonstrate some uniform qualities.

Aging of gelatin in capsules for shelf-life testing is mimicked using high temperature and humidity conditions. Both conditions induce further cross-linking of the gelatin chains and increase not only the hardness but the dissolving rate of the capsules.

Other aspects of gelatin are monitored such as the changes to lipids inherent in the material, dityrosine content, 3,4 dihydroxyphenylalanine (DOPA) and levels of oxidation (Duconseille et al., 2017).

The rate of dissolution is linked to the ability of the gelatin chains to bind water based on near infrared spectroscopy examination. As gelatin ages it also appears to develop more methylene groups (-CH2-) and more aldehydic groups.

The same group explored Raman microspectroscopy before and after aging of gelatin (Duconseille et al., 2018). Here and relying on principal component analysis (PCA) they could see aging caused denaturation of the gelatin triple-helices as well as cross-linking due to dityrosine, glucosyl-galactosylhydroxylysinonorleucine and proteoglycan-like sugar adducts. This study also seemed to confirm that poorer or non-compliant gelatins had a higher lipid content than the compliant ones.

Regulations On Gelatin Use And Manufacture

Europe is highly dependent on importing a vareity of raw materials for the production of gelatin and collagen. It is a requirement of food business operators that they ensure raw materials for all human consumption come from sources that meet stringent public and animal health requirements based on EU legislation.

Any treatments must be clearly specified and comply with relevant residue levels and limits. There are also appropriate analytical methods and validated methods needed to ensure these are accurately monitored.

The production standards for both gelatin and collagen manufacture:

  •  Council Directive 2002/99/EC laying down the animal health rules governing the production, processing, distribution and introduction of products of animal origin for human consumption;
  • Regulation (EC) No 178/2002 laying down the general principles and requirements of food law, establishing the European Food Safety Authority and laying down procedures in matters of food safety;
  • Regulation (EC) No 852/2004 on the hygiene of foodstuffs;
  • Regulation (EC) No 853/2004 laying down specific rules for food of animal origin;
  • Regulation (EU) 2017/625 laying down specific rules for the organisation of official controls on products of animal origin intended for human consumption; and on official controls performed to ensure the verification of compliance with feed and food law, animal health and animal welfare rules.

The European Commission (EC) issued a Commission Regulation (EU) 2016/355 which is titled ‘Specific Requirements for Gelatine, Collagen and Highly Refined Products of Animal Origin intended for Human Consumption’. This is an amendment of Annex III of the EU regulation No. 853/2004 which covers revised toxicity levels for contaminants especially heavy metals. It was published on the 11th March and became effects on 1st April, 2016.

The amended annex III means the following: 

  • raw materials which had not previously been preserved save for being frozen, chilled or quick-frozen had to come from registered or approved establishments according to Regulation (EC) No. 852/2004, on the hygiene of foodstuffs or in accordance with this regulation.
  • The following treated raw materials of the production of gelatin and collagen such as bones, hides and skins of farmed ruminant animals, pig skins, poultry skins, wild game hides and skins coming from establishments under the control of and listed by the competent authority may be used the specific treatment in accordance with this regulation

  • Raw material for the production of gelatin and collagen for human consumption for which animal health certification is required, the raw materials must be transported directly to the establishment at the place of destination

  • The production process for collagen, all ruminant bone material derived from animals born, reared or slaughtered in countries or regions with a controlled or undetermined Bovine Spongiform Encephalopathy (BSE) risk, must ensure that all bone material is finely crushed and degreased with hot water, washed and treated with dilute hydrochloric acid. If it’s over a period of at least two days, the treatment must be followed by pH adjustment using acid or alkali followed by one or more rinses, filtration and extrusion

  • Moreover, food business operators manufacturing highly refined products of animal origin such as chondroitin sulfate, hyaluronic acid, other hydrolyzed cartilage products, chitosan, glucosamine, rennet, isinglass and certain amino acids, must ensure the treatment of the raw material used eliminates any animal or public health risk.

The UK’s DEFRA through its Animal & Plant Health Agency also prepared a document entitled ‘Import of Gelatine and Collagen for Human Consumption from Third Countries Import Information Note (IIN) BLGC/1‘ in February 2020 which is explicit in the requirements on the importing of gelatin products from countries outside the EU.

References

Buice R.G. Jr., Gold T.B., Lodder R.A., Digenis G.A. (1995). Determination of moisture in intact gelatin capsules by Near-Infrared Spectroscopy. J. Pharm. Res. 12 (1), pp. 161-163

Duconseille, A., Andueza, D., Picard, F., Santé-Lhoutellier, V., & Astruc, T. (2017). Variability in pig skin gelatin properties related to production site: A near infrared and fluorescence spectroscopy study. Food Hydrocolloids63, pp. 108-119 (Article)

Duconseille, A., Gaillard, C., Santé-Lhoutellier, V., & Astruc, T. (2018). Molecular and structural changes in gelatin evidenced by Raman microspectroscopy. Food Hydrocolloids77, pp. 777-786.

Gennadios AMcHugh THWeller CLKrochta JM1994Edible coating and films based on proteins. In: Krochta JMBaldwin EA Nisperos‐ Carriedo M, editors. Edible coatings to improve food quality. Lancaster , Pa .: Technomic Pub. Co. Inc. p 210278.  

Hanani, Z. N., Beatty, E., Roos, Y. H., Morris, M. A., & Kerry, J. P. (2012). Manufacture and characterization of gelatin films derived from beef, pork and fish sources using twin screw extrusion. Journal of Food Engineering113(4), pp. 606-614 (Article)

Krochta JM, De Mulder‐Johnson C. (1997)Edible and biodegradable polymer films: challenges and opportunitiesFood Technol. 51(2) pp. 6174 (Article).

Marois YChakfé NLDeng XMarois MHow TKing MWGuidoin R. (1995) Carbodiimide cross‐linked gelatin: a new coating for porous polyester arterial prosthesesBiomaterials. 16: pp. 11311139.

Nishimoto M., Sakamoto R., Mizuta S., Yoshinaka R., (2005) Identification and characterization of molecular species of collagen in ordinary muscle and skin of the Japanese flounder (Paralichthys olivaceus). J. Food Chem. 90, pp. 151-156 .

Sams A.R., (2001). Poultry Meat Processing. CRC Press Taylor and Francis Boca Raton, USA . 

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