Ultrasonic Collagen Extraction
- Collagen is rich in proteins and is widely used in manifold industrial applications, e.g. food, pharma, additives etc.
- Sonication can be easily combined with enzymatic or acid extraction of collagen.
- The implementation of ultrasonics into the collagen extraction process results in higher yields and faster extraction.
Ultrasonic Effects on Collagen Extraction
High-intensity ultrasound is widely used to improve mass transfer in wet processes, e.g. extraction, sonochemistry etc. The extraction (also known as collagen isolation) of collagen can be significantly improved by ultrasonic treatment. Sonication aids during the cleavage of the collagen substrate, opens up the collagen fibrils, thus enzymatic hydrolysis or acid treatment are facilitated.
Ultrasonically Assisted Enzymatic Extraction
Sonication is known for its capability to increase enzyme activity. This effect is based on the ultrasonic dispersion and deagglomeration of the pepsin aggregates. Homogeneously dispersed enzymes offer an increased surface for mass transfer, which is correlated to higher enzyme activity. Furthermore, the powerful ultrasound waves opens up the collagen fibrils so that the collagen is released.
Ultrasonic Pepsin Extraction: Pepsin combined the ultrasonication increases the yield of collagen up to approx. 124% and significantly shortens the extraction time in comparison with the conventional pepsin hydrolysis. Circular dichroism analysis, atomic force microscopy and FTIR proved that the triple helix structure of the extracted collagen was not affected by sonication and remained intact. (Li et al. 2009) This makes the ultrasonically assisted pepsin extraction highly practical for the food industry offering increased protein recovery rates in a significantly shorter processing time.
In a comparative study of ultrasonic vs. non-ultrasonic extraction of collagen from bovine tendon, the ultrasound treatment (20kHz, pulse mode 20/20 sec.) convinced by higher yield and efficiency. The conventional extraction was performed with pepsin in acetic acid for 48 hours. The ultrasonic extraction was performed extraction under the same conditions, but the exposure times to sonication (3 to 24 h) and pepsin (24 to 45 hours) were varied, resulting in a total of 48 hours of treatment. The ultrasonic-pepsin extraction showed superior efficiency of collagen extraction, reaching a yield of 6.2%, when the conventional extraction yield was 2.4%. Best results were achieved at an ultrasonic extraction time using of 18 h. The extracted collagen showed an undamaged continuous helix structure, good solubility and fairly high thermal stability. this means that an ultrasonic-pepsin extraction improved the efficiency of the extraction of natural collagen without damaging the quality of the resulting collagen. (Ran and Wang 2014)
Ultrasonically Assisted Acid Extraction
In a study by Kim et al. (2012), the extraction of acid-soluble collagen from the skin of Japanese sea bass (Lateolabrax japonicus) showed increased yield and reduced extraction time after ultrasonic treatment at a frequency of 20 kHz in 0.5 M acetic acid. Extraction with ultrasound did not alter the major components of the collagen, more specifically the α1, α2 and β chains.
Ultrasonic Extraction of Protein from Egg Shells
Ultrasonically pre-treated enzymatic hydrolysates had better functional properties. For the ultrasonic extraction of functional protein hydrolysates from chicken egg shell, solubility, emulsifying, foaming and water holding properties are improved.
Eggshell membrane is an abundant natural resource and consists of about 64 proteins including Type I, V and X collagen, lysozyme, osteopontin, and sialoprotein. This makes eggshells an interesting raw material for the extraction of proteins. With ultrasonic extraction, the protein release and functionality can be significantly improved resulting in a fast, efficient and economical process.
Ultrasonically Assisted Alkali Extraction
to extract and solubilise these proteins
For the protein extraction from eggshell membrane, the ultrasonic-alkali treatment resulted in a solubilised protein yield close to 100% of the total eggshell membrane protein. Ultrasonic cavitation detached bigger proteins clumps from the eggshell membrane and facilitated the solubilisation of its compounds. The protein structure and properties were not damaged by sonication and remained intact. The antioxidant properties of the proteins were the same for the ultrasonic-assisted alkaline treatment and conventional extraction.
Ultrasonic Gelatin Extraction
Frozen and air-dried pollock skins were treated with cold saline, alkaline and acid solutions to separate collagen tissue and extract gelatin by collagen denaturation at 45°C for four hours with a power ultrasound treatment as a processing aid. Gelatin yield, pH, clarity, gel strength and viscoelastic properties as well as molecular weight distribution determined by PAGE-SDS method were evaluated. Gelatin extracted in a water bath at 45°C for four hours was used as a control. The power ultrasound treatment increased the extraction yield by 11.1% compared with the control while gel strength decreased 7%. Gelation temperature was also lower in ultrasound-extracted gelatin (4.2°C). This behavior is related to differences in molecular weight distribution of polypeptide coils in gelatins. Power ultrasound extraction can be used to increase gelatin extraction from frozen and air-dried fish skins. (Olson et al. 2005)
Industrial Ultrasonic Systems
Hielscher Ultrasonics supplies powerful ultrasonic systems from lab to bench-top and industrial scale. To ensure optimum extraction output, reliable sonication under demanding conditions can be performed continuously. All industrial ultrasonic processors can deliver very high amplitudes. Amplitudes of up to 200µm can be easily continuously run in 24/7 operation. For even higher amplitudes, customized ultrasonic sonotrodes are available. The robustness of Hielscher’s ultrasonic equipment allows for 24/7 operation at heavy duty and in demanding environments.
Please contact us today with your process requirements! We will be glad to recommend you a suitable ultrasonic system for your process!
- Álvarez, Carlos; Lélu, Pauline; Lynch, Sarah A.; Tiwari, Brijesh K. (2018): Optimised protein recovery from mackerel whole fish by using sequential acid/alkaline isoelectric solubilization precipitation (ISP) extraction assisted by ultrasound. LWT – Food Science and Technology Vol. 88, February 2018. 210-216.
- Jain, Surangna; Kumar Anal, Anil (2016): Optimization of extraction of functional protein hydrolysates from chicken egg shell membrane (ESM) by ultrasonic assisted extraction (UAE) and enzymatic hydrolysis. LWT – Food Science and Technology Vol. 69, June 2016. 295-302.
- Kim, H.K.; Kim, Y.H.; Kim, Y.J.; Park, H.J.; Lee, N.H. (2012): Effects of ultrasonic treatment on collagen extraction from skins of the sea bass Lateolabrax japonicus. Fisheries Science Volume 78, Issue 78; 2013. 485-490.
- Li, Defu; Mu, Changdao; Cai, Sumei; Lin, Wei (2016): Ultrasonic irradiation in the enzymatic extraction of collagen. Ultrasonics Sonochemistry Volume 16, Issue 5; 2009. 605-609.
- Olson, D.A., Avena Bustillos, R.D., Olsen, C.W., Chiou, B., Yee, E., Bower, C.K., Bechtel, P.J., Pan, Z., Mc Hugh, T.H. (2005): Evaluation of power ultrasound as a processing aid for fish gelatin extraction. Meeting Abstract No. 71C-26. IFT Annual Meeting. July 2005. New Orleans, LA.
- Ran, X.G.; Wang, L.Y. (2014): Use of ultrasonic and pepsin treatment in tandem for collagen extraction from meat industry by-products. Journal of the Science of Food and Agriculture 94(3), 2014. 585-590.
- Schmidt, M.M.; Dornelles, R.C.P.; Mello, R.O.; Kubota, E.H.; Mazutti, M.A.; Kempka, A.P.; Demiate, I.M. (2016): Collagen extraction process. International Food Research Journal 23(3), 2016. 913-922.
- Siritientong, Tippawan; Bonani, Walter; Motta, Antonella; Migliaresi, Claudio; Aramwit, Pornanong (2016): The effects of Bombyx mori silk strain and extraction time on the molecular and biological characteristics of sericin. Bioscience, Biotechnology, and Biochemistry Vol. 80 , Iss. 2, 2016. 241-249.
- Zeng, J.N.; Jiang, B.Q.; Xiao, Z.Q., Li, S.H. (2011): Extraction of Collagen from Fish Scales with Papain under Ultrasonic Pretreatment. Advanced Materials Research, Volume 366, 2011. 421-424.
Facts Worth Knowing
Collagen is the main structural protein in the extracellular space in the various connective tissues in animal bodies. As the main component of connective tissue, it is the most abundant protein in mammals, making up from 25% to 35% of the whole-body protein content. Collagen consists of amino acids wound together to form triple-helices to form of elongated fibrils. The highest amounts of collagen are present in fibrous tissues such as tendons, ligaments and skin. There are three types of collagen to be distinguished:
Type I collagen: provides 90% of protein in skin, hair, nails, organs, bone, ligaments
Type II collagen: provides 50-60% of protein in cartilage, 85-90% of collagen in articular cartilage
Type III collagen: provides proteins to fibrous protein in bone, cartilage, dentin, tendon, and other connective tissues
Collagen in the Body
Each of the three collagen types is composed from different proteins which fulfil different purposes in the body. The collagen types I and III both are main components of skin, muscles, bone, hair and nail. They are required for their health, growth and rebuilding. The collagen type II is mostly found in the cartilage and joints.
Collagen of type I and III both contain 19 amino acids which are considered as essential amino acids. They are produced by fibroblasts (cells in connective tissues) and osteoblasts (cells that make bones).The most important proteins in collagen type I and III include glycine, proline, alanine, and hydroxyproline. Type III is a fibrous scleroprotein.
Glycine is the amino acid with the highest amount in collagen. Proline is a non-essential amino acid, which can be synthesized from glycine and contributes to joints and tendons. Hydroxyproline is an amino acid that contributes to the stability of collagen. Alanine is an amino acid important for the biosynthesis of proteins.
Like type I and III, type II collagen does form fibrils. This fibrillar network of collagen is important in cartilage since is enables for the entrapment of proteoglycans. Furthermore, it provides tensile strength to the tissue.
Sources and Uses
Collagen is a fibrous protein which is abundantly present in the connective tissue of mammals, e.g. bovine, pig. Most collagen is extracted
from porcine skins and bones and from bovine sources. An alternative source for collagen extraction are fish and fowl. Collagen is widely used in food, dietary supplements, pharmaceuticals/medicals, and cosmetics among other products. The collagen extraction is a growing business since this protein can replace synthetic agents in various industrial processes.