Skip to main content

Sequential Extraction and Characterisation of Lipids, Proteins, and Chitin from Black Soldier Fly (Hermetia illucens) Larvae, Prepupae, and Pupae

Waste and Biomass Valorization volume 11, pages 6455–6466 (2020)Cite this article

Abstract

Over the past years, several insect species have gained increased attention as feedstock for food, feed, and industrial applications. One such species is Hermetia illucens, whose larvae can convert low-value organic waste into valuable fat- and protein-rich biomass. Previous research on extracting their lipids, proteins, and chitin has repeatedly focused on one life stage, while in practice different life stages coexist in the same rearing batch. In this study, the feasibility of the sequential extraction of said components from the larval, prepupal, and pupal stage of H. illucens was investigated. Additionally, the chemical composition of the life stages and their extracts was analysed. Following the lipid extraction with petroleum ether, insect proteins were extracted via solubilisation at pH 11.0 and precipitation at pH 4.0. This procedure delivered protein recoveries ranging between 27 and 57% for the three life stages, with the extracts having high protein contents (85–98%). After protein extraction, the residual impure chitin was treated sequentially with HCl and NaOH for further purification. No residual amino acids were detected by UPLC analysis of the purified chitin, which showed acetylation degrees of ± 90%. Overall, it was concluded that the extraction procedure is indeed suitable for all investigated life stages of H. illucens, allowing for the extraction high-value biomolecules for use in industrial applications.

Graphic Abstract

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

USD 39.95

Price includes VAT (USA)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3

References

  1. Gold, M., Tomberlin, J.K., Diener, S., Zurbrügg, C., Mathys, A.: Decomposition of biowaste macronutrients, microbes, and chemicals in black soldier fly larval treatment: a review. Waste Manag. 82, 302–318 (2018). https://doi.org/10.1016/j.wasman.2018.10.022

    Article  Google Scholar 

  2. Newton, L., Sheppard, C., Watson, D.W., Burtle, G., Dove, R.: Using the black soldier fly, Hermetia illucens, as a value-added tool for the management of swine manure (2005).

  3. Liu, X., Chen, X., Wang, H., Yang, Q., ur Rehman, K., Li, W., Cai, M., Li, Q., Li, L., Mazza, J., Zhang, Z., Yu, L.: Dynamic changes of nutrient composition throughout the entire life cycle of black soldier fly. PLoS ONE 12, 1–21 (2017). https://doi.org/10.1371/journal.pone.0182601

    Article  Google Scholar 

  4. Finke, M.D.: Complete nutrient composition of commercially raised invertebrates used as food for insectivores. Zoo Biol. 21, 269–285 (2002). https://doi.org/10.1002/zoo.10031

    Article  Google Scholar 

  5. Arango Gutiérrez, G.P., Vergara Ruiz, R.A., Mejía Vélez, H.: Compositional, microbiological and protein digestibility analysis of larval meal of Hermetia illucens (Diptera:Stratiomyiidae) at Angelópolis-Antioquia. Colombia. Rev. Fac. Nac. Agron. Medellín. 57, 2491–2499 (2004)

    Google Scholar 

  6. Barry, T.: Evaluation of the economic, social, and biological feasibility of bioconverting food wastes with the black soldier fly (Hermetia illucens). University of North Texas, Denton (2004)

    Google Scholar 

  7. Dordević, M., Brana, R.D., Marijana, V., Baltić, M., Radislava, T., Ljiljana, J., Marija, V., Rajković, M.: Effects of substitution of fish meal with fresh and dehydrated larvae of the house fly (Musca domestica L.) on productive performance and health of broilers. Acta Vet. Brno. 58, 357–368 (2008). https://doi.org/10.2298/AVB0804357D

    Article  Google Scholar 

  8. Shumo, M., Osuga, I.M., Khamis, F.M., Tanga, C.M., Fiaboe, K.K.M., Subramanian, S., Ekesi, S., van Huis, A., Borgemeister, C.: The nutritive value of black soldier fly larvae reared on common organic waste streams in Kenya. Sci. Rep. 9, 1–13 (2019). https://doi.org/10.1038/s41598-019-46603-z

    Article  Google Scholar 

  9. Spranghers, T., Ottoboni, M., Klootwijk, C., Ovyn, A., Deboosere, S., De Meulenaer, B., Michiels, J., Eeckhout, M., De Clercq, P., De Smet, S.: Nutritional composition of black soldier fly (Hermetia illucens) prepupae reared on different organic waste substrates. J. Sci. Food Agric. 97, 2594–2600 (2017). https://doi.org/10.1002/jsfa.8081

    Article  Google Scholar 

  10. Tschirner, M., Simon, A.: Influence of different growing substrates and processing on the nutrient composition of black soldier fly larvae destined for animal feed. J. Insects as Food Feed. 1, 249–259 (2015). https://doi.org/10.3920/JIFF2014.0008

    Article  Google Scholar 

  11. St-Hilaire, S., Sheppard, C., Tomberlin, J.K., Irving, S., Newton, L., McGuire, M.A., Mosley, E.E., Hardy, R.W., Sealey, W.: Fly prepupae as a feedstuff for rainbow trout, Oncorhynchus mykiss. J. World Aquac. Soc. 38, 59–67 (2007). https://doi.org/10.1111/j.1749-7345.2006.00073.x

    Article  Google Scholar 

  12. Kawasaki, K., Hashimoto, Y., Hori, A., Kawasaki, T., Hirayasu, H., Iwase, S., Hashizume, A., Ido, A., Miura, C., Miura, T., Nakamura, S., Seyama, T., Matsumoto, Y., Kasai, K., Fujitani, Y.: Evaluation of black soldier fly (Hermetia illucens) larvae and pre-pupae raised on household organic waste, as potential ingredients for poultry feed. Animals. 9, 98 (2019). https://doi.org/10.3390/ani9030098

    Article  Google Scholar 

  13. Belghit, I., Liland, N.S., Gjesdal, P., Biancarosa, I., Menchetti, E., Li, Y., Waagbø, R., Krogdahl, Å., Lock, E.J.: Black soldier fly larvae meal can replace fish meal in diets of sea-water phase Atlantic salmon (Salmo salar). Aquaculture 503, 609–619 (2019). https://doi.org/10.1016/j.aquaculture.2018.12.032

    Article  Google Scholar 

  14. Wang, Y.-S., Shelomi, M.: Review of black soldier fly (Hermetia illucens) as animal feed and human food. Foods. 6, 91 (2017). https://doi.org/10.3390/foods6100091

    Article  Google Scholar 

  15. Caligiani, A., Marseglia, A., Leni, G., Baldassarre, S., Maistrello, L., Dossena, A., Sforza, S.: Composition of black soldier fly prepupae and systematic approaches for extraction and fractionation of proteins, lipids and chitin. Food Res. Int. 105, 812–820 (2018). https://doi.org/10.1016/j.foodres.2017.12.012

    Article  Google Scholar 

  16. Li, Q., Zheng, L., Qiu, N., Cai, H., Tomberlin, J.K., Yu, Z.: Bioconversion of dairy manure by black soldier fly (Diptera: Stratiomyidae) for biodiesel and sugar production. Waste Manag. 31, 1316–1320 (2011). https://doi.org/10.1016/j.wasman.2011.01.005

    Article  Google Scholar 

  17. Verheyen, G.R., Ooms, T., Vogels, L., Vreysen, S., Bovy, A., Van Miert, S., Meersman, F.: Insects as an alternative source for the production of fats for cosmetics. J. Cosmet. Sci. 69, 187–202 (2018)

    Google Scholar 

  18. Kumar, M.N.V.: A review of chitin and chitosan applications. React. Funct. Polym. 46, 1–27 (2000)

    Article  Google Scholar 

  19. Aranaz, I., Mengibar, M., Harris, R., Panos, I., Miralles, B., Acosta, N., Galed, G., Heras, A.: Functional characterization of chitin and chitosan. Curr. Chem. Biol. 3, 203–230 (2009). https://doi.org/10.2174/187231309788166415

    Article  Google Scholar 

  20. Kaya, M., Baran, T., Asan-Ozusaglam, M., Cakmak, Y.S., Tozak, K.O., Mol, A., Mentes, A., Sezen, G.: Extraction and characterization of chitin and chitosan with antimicrobial and antioxidant activities from cosmopolitan Orthoptera species (Insecta). Biotechnol. Bioprocess Eng. 20, 168–179 (2015). https://doi.org/10.1007/s12257-014-0391-z

    Article  Google Scholar 

  21. Hayes, M.: Chitin, Chitosan and their derivatives from marine rest raw materials: potential food and pharmaceutical applications. In: Marine Bioactive Compounds, pp. 1–229. https://doi.org/10.1007/978-1-4614-1247-2 (2012).

  22. Synowiecki, J., Al-Khateeb, N.A.: Production, properties, and some new applications of chitin and its derivatives. Crit. Rev. Food Sci. Nutr. 43, 145–171 (2003). https://doi.org/10.1080/10408690390826473

    Article  Google Scholar 

  23. Lim, S.H., Hudson, S.M.: Review of chitosan and its derivatives as antimicrobial agents and their uses as textile chemicals. J. Macromol. Sci. 43, 223–269 (2003). https://doi.org/10.1081/MC-120020161

    Article  Google Scholar 

  24. Arbia, W., Arbia, L., Adour, L., Amrane, A.: Chitin extraction from crustacean shells using biological methods—a review. Food Technol. Biotechnol. 51, 12–25 (2013). https://doi.org/10.1002/biot.200800027

    Article  Google Scholar 

  25. Hamed, I., Özogul, F., Regenstein, J.M.: Industrial applications of crustacean by-products (chitin, chitosan, and chitooligosaccharides): a review. Trends Food Sci. Technol. 48, 40–50 (2016). https://doi.org/10.1016/j.tifs.2015.11.007

    Article  Google Scholar 

  26. Liu, S., Sun, J., Yu, L., Zhang, C., Bi, J., Zhu, F., Qu, M., Jiang, C., Yang, Q.: Extraction and characterization of chitin from the beetle Holotrichia parallela motschulsky. Molecules 17, 4604–4611 (2012). https://doi.org/10.3390/molecules17044604

    Article  Google Scholar 

  27. Kaya, M., Baublys, V., Can, E., Satkauskiene, I., Bitim, B., Tubelyte, V., Baran, T.: Comparison of physicochemical properties of chitins isolated from an insect (Melolontha melolontha) and a crustacean species (Oniscus asellus). Zoomorphology 133, 285–293 (2014). https://doi.org/10.1007/s00435-014-0227-6

    Article  Google Scholar 

  28. Kaya, M., Sofi, K., Sargin, I., Mujtaba, M.: Changes in physicochemical properties of chitin at developmental stages (larvae, pupa and adult) of Vespa crabro (wasp). Carbohydr. Polym. 145, 64–70 (2016). https://doi.org/10.1016/j.carbpol.2016.03.010

    Article  Google Scholar 

  29. Sheppard, D.C., Tomberlin, J.K., Joyce, J.A., Kiser, B.C., Sumner, S.M.: Rearing methods for the black soldier fly (Diptera: Stratiomyidae): table 1. J. Med. Entomol. 39, 695–698 (2002). https://doi.org/10.1603/0022-2585-39.4.695

    Article  Google Scholar 

  30. Tomberlin, J.K., Sheppard, D.C., Joyce, J.A.: Selected life-history traits of black soldier flies (Diptera: Stratiomyidae) reared on three artificial diets. Ann. Entomol. Soc. Am. 95, 379–386 (2006). https://doi.org/10.1603/0013-8746(2002)095[0379:slhtob]2.0.co;2

    Article  Google Scholar 

  31. Harnden, L.M., Tomberlin, J.K.: Effects of temperature and diet on black soldier fly, Hermetia illucens (L.) (Diptera: Stratiomyidae), development. Forensic Sci. Int. 266, 109–116 (2016). https://doi.org/10.1016/j.forsciint.2016.05.007

    Article  Google Scholar 

  32. Gao, Q., Deng, W., Gao, Z., Li, M., Liu, W., Wang, X., Zhu, F.: Effect of sulfonamide pollution on the growth of manure management candidate Hermetia illucens. PLoS ONE 14, e0216086 (2019). https://doi.org/10.1371/journal.pone.0216086

    Article  Google Scholar 

  33. Myers, H.M., Tomberlin, J.K., Lambert, B.D., Kattes, D.: Development of black soldier fly (Diptera: Stratiomyidae) larvae fed dairy manure. Environ. Entomol. 37, 11–15 (2008). https://doi.org/10.1093/ee/37.1.11

    Article  Google Scholar 

  34. Barros, L.M., Gutjahr, A.L.N., Ferreira-Keppler, R.L., Martins, R.T.: Morphological description of the immature stages of Hermetia illucens (Linnaeus, 1758) (Diptera: Stratiomyidae). Microsc. Res. Tech. 82, 178–189 (2019). https://doi.org/10.1002/jemt.23127

    Article  Google Scholar 

  35. Liao, C., Upadhyay, A., Liang, J., Han, Q., Li, J.: 3,4-Dihydroxyphenylacetaldehyde synthase and cuticle formation in insects. Dev. Comp. Immunol. 83, 44–50 (2018). https://doi.org/10.1016/j.dci.2017.11.007

    Article  Google Scholar 

  36. Merritt, M.E., Christensen, A.M., Kramer, K.J., Hopkins, T.L., Schaefer, J.: Detection of intercatechol cross-links in insect cuticle by solid-state carbon-13 and nitrogen-15 NMR. J. Am. Chem. Soc. 118, 11278–11282 (1996). https://doi.org/10.1021/ja961621o

    Article  Google Scholar 

  37. Vandeweyer, D., Lenaerts, S., Callens, A., Van Campenhout, L.: Effect of blanching followed by refrigerated storage or industrial microwave drying on the microbial load of yellow mealworm larvae (Tenebrio molitor). Food Control 71, 311–314 (2017). https://doi.org/10.1016/j.foodcont.2016.07.011

    Article  Google Scholar 

  38. Fombong, F.T., Van Der Borght, M., Vanden Broeck, J.: Influence of freeze-drying and oven-drying post blanching on the nutrient composition of the edible insect Ruspolia differens. Insects. 8, 102 (2017). https://doi.org/10.3390/insects8030102

    Article  Google Scholar 

  39. Lenaerts, S., Van Der Borght, M., Callens, A., Van Campenhout, L.: Suitability of microwave drying for mealworms (Tenebrio molitor) as alternative to freeze drying: Impact on nutritional quality and colour. Food Chem. 254, 129–136 (2018). https://doi.org/10.1016/j.foodchem.2018.02.006

    Article  Google Scholar 

  40. Jonas-Levi, A., Martinez, J.J.I.: The high level of protein content reported in insects for food and feed is overestimated. J. Food Compos. Anal. 62, 184–188 (2017). https://doi.org/10.1016/j.jfca.2017.06.004

    Article  Google Scholar 

  41. Janssen, R.H., Vincken, J.-P., van den Broek, L.A.M., Fogliano, V., Lakemond, C.M.M.: Nitrogen-to-protein conversion factors for three edible insects: Tenebrio molitor, Alphitobius diaperinus, and Hermetia illucens. J. Agric. Food Chem. 65, 2275–2278 (2017). https://doi.org/10.1021/acs.jafc.7b00471

    Article  Google Scholar 

  42. Meneguz, M., Schiavone, A., Gai, F., Dama, A., Lussiana, C., Renna, M., Gasco, L.: Effect of rearing substrate on growth performance, waste reduction efficiency and chemical composition of black soldier fly (Hermetia illucens) larvae. J. Sci. Food Agric. 98, 5776–5784 (2018). https://doi.org/10.1002/jsfa.9127

    Article  Google Scholar 

  43. Bosch, G., van Zanten, H.H.E., Zamprogna, A., Veenenbos, M., Meijer, N.P., van der Fels-Klerx, H.J., van Loon, J.J.A.: Conversion of organic resources by black soldier fly larvae: legislation, efficiency and environmental impact. J. Clean. Prod. 222, 355–363 (2019). https://doi.org/10.1016/j.jclepro.2019.02.270

    Article  Google Scholar 

  44. Barragan-Fonseca, K.B., Dicke, M., van Loon, J.J.A.: Influence of larval density and dietary nutrient concentration on performance, body protein, and fat contents of black soldier fly larvae (Hermetia illucens). Entomol. Exp. Appl. 166, 761–770 (2018). https://doi.org/10.1111/eea.12716

    Article  Google Scholar 

  45. Del Valle, F.R., Mena, M.H.: An investigation into insect protein. J. Food Process. Preserv. 6, 99–110 (1982)

    Article  Google Scholar 

  46. Joseph, J.D., Ackman, R.G.: Capillary column gas chromatographic method for analysis of encapsulated fish oils and fish oil ethyl-esters: collaborative study. J. AOAC Int. 75, 488–506 (1992)

    Article  Google Scholar 

  47. Brugnerotto, J., Lizardi, J., Goycoolea, F.M., Argüelles-Monal, W., Desbrières, J., Rinaudo, M.: An infrared investigation in relation with chitin and chitosan characterization. Polymer 42, 3569–3580 (2001). https://doi.org/10.1016/S0032-3861(00)00713-8

    Article  Google Scholar 

  48. Bußler, S., Rumpold, B.A., Jander, E., Rawel, H.M., Schlüter, O.K.: Recovery and techno-functionality of flours and proteins from two edible insect species: Meal worm (Tenebrio molitor) and black soldier fly (Hermetia illucens) larvae. Heliyon. 2, e00218 (2016). https://doi.org/10.1016/j.heliyon.2016.e00218

    Article  Google Scholar 

  49. D’Mello, J.P.F.: Amino Acids in Animal Nutrition, 2nd edn. CABI publishing, Cambridge. (2003) https://doi.org/10.1079/9780851996547.0000.

  50. Leni, G., Caligiani, A., Sforza, S.: Killing method affects the browning and the quality of the protein fraction of black soldier fly (Hermetia illucens) prepupae: a metabolomics and proteomic insight. Food Res. Int. (2018). https://doi.org/10.1016/J.FOODRES.2018.08.021

    Article  Google Scholar 

  51. Cárdenas, G., Cabrera, G., Taboada, E., Miranda, S.P.: Chitin characterization by SEM, FTIR, XRD, and13C cross polarization/mass angle spinning NMR. J. Appl. Polym. Sci. 93, 1876–1885 (2004). https://doi.org/10.1002/app.20647

    Article  Google Scholar 

  52. Waśko, A., Bulak, P., Polak-Berecka, M., Nowak, K., Polakowski, C., Bieganowski, A.: The first report of the physicochemical structure of chitin isolated from Hermetia illucens. Int. J. Biol. Macromol. 92, 316–320 (2016). https://doi.org/10.1016/j.ijbiomac.2016.07.038

    Article  Google Scholar 

  53. Kaya, M., Baran, T., Erdoʇan, S., Menteş, A., Aşan Özüsaʇlam, M., Çakmak, Y.S.: Physicochemical comparison of chitin and chitosan obtained from larvae and adult Colorado potato beetle (Leptinotarsa decemlineata). Mater. Sci. Eng. C. 45, 72–81 (2014). https://doi.org/10.1016/j.msec.2014.09.004

    Article  Google Scholar 

  54. Oonincx, D.G.A.B., Van Broekhoven, S., Van Huis, A., Van Loon, J.J.A.: Feed conversion, survival and development, and composition of four insect species on diets composed of food by-products. PLoS ONE 10, 1–20 (2015). https://doi.org/10.1371/journal.pone.0144601

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Microbial and Molecular Systems, Faculty of Engineering Technology, Lab4food, KU Leuven, Kleinhoefstraat 4, 2440, Geel, Belgium

    Ruben Smets, Johan Claes & Mik Van Der Borght

  2. Thomas More University of Applied Sciences, RADIUS, Kleinhoefstraat 4, 2440, Geel, Belgium

    Bert Verbinnen

  3. Laboratory of Enzyme, Fermentation & Brewing Technology, Department of Microbial and Molecular Systems, Faculty of Engineering Technology, KU Leuven, Gebroeders de Smetstraat 1, 9000, Ghent, Belgium

    Ilse Van De Voorde & Guido Aerts

Authors
  1. Ruben Smets
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Bert Verbinnen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ilse Van De Voorde
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Guido Aerts
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Johan Claes
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Mik Van Der Borght
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mik Van Der Borght.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Smets, R., Verbinnen, B., Van De Voorde, I. et al. Sequential Extraction and Characterisation of Lipids, Proteins, and Chitin from Black Soldier Fly (Hermetia illucens) Larvae, Prepupae, and Pupae. Waste Biomass Valor 11, 6455–6466 (2020). https://doi.org/10.1007/s12649-019-00924-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12649-019-00924-2

Keywords

  • Hermetia illucens
  • Life stages
  • Chemical extraction
  • Lipids
  • Proteins
  • Chitin