LWT ood Scene an Tehaoogy 912018) 158-174 Contents lists available a ScienceDirect, hae LWT - Food Science and Technology ELSEVIER journal homepage: www. slsevier.comilocate/lwt Comparison of functional properties of edible insects and protein preparations thereof Ewelina Zielitiska", Monika Karas, Barbara Baraniak Digan of chair and fod Chenin, Overy of Life eens in ain, Slow $8, 20.704 Lain. Pld ARTICLE INFO ABSTRACT Komerte “Tis study investigted the functional properties of thee spectes of elbe insects: Gryodes alas, Schistocerca ae sees eget, snd Tenebrio moliar. The water and oll holding apse, solubility, and fosming and emulsion prop- onopbeny ‘les were evaluated. The protein salbilty showed minimm values at pit 5. The highest water and oil holding Fancoal rope ‘capacity was noticeable for the T: molor protein preparation (2954/8) and forthe 6, sii protein pre Datetion (3.35 g/g), respectively. The 6. sigan protein preparation also showed the highest foaming eapeet, ‘oam stably, and emulsion att (98.0%, 92.0%, and 72.52%, respectively, while the poten preparation ‘rom 5 gregoria exhibited the highest emulsion stability (51.31%) This study has shown that whole insets and protein preparations thereat can be rable for development of net fod formations. 1. Introduction In the last decade, che interest of entomophagy has been con- Linuously growing. Currently, insects are cossumed by two billion ‘people worldwide end even insect foads have recently became available in the US and Europe. More than 2100 insect species have been doce ‘mented in literature as edible (ongema, 2017). Moreover, insects are still promoted asa good source of protein and the production of edible insects in developing countries is supported by various institations such ‘as the Food and Agriculture Organization of the United Nations However, the use of insects in food production requires further ine vestigations at different levels, for example to search for opporunities ‘to use chem in various forms. This may be necessary since Western consumers may be reluctant to accept insects as a protein source (Ghelomi, 2025). In many countries, whole insects are often consumed but they can also be processed to pastes and powders; furthermore, insect proteins, fats and chitin can be isolated before use in food pro ducts as well This could be a useful way for increasing acceptability among wary consumers. Edible insects can also be processed into & ‘more palatable form by grinding or milling. Iis an easy way to obtain high-protein insect flour with other valuable components such as i tamins of minerals (Viet sl, 2013) ‘Several studies have shown that edible insects area good source of protein (Ramos-Forduy, Moreno, & Camacho, 2012; Rumpold & Schluter, 2013; Zielifska, Baraniak, Karas, Rybezyiska, & Jakubezyk, 2015) but there are only few data about the funetional properties of insect protein. These properties could be helpful to clarify the use of Insect powder or protein extracts in diferent food products, for ex ample bread, pasta, and dairy products Currently, many commercial food products are fortified in order to Increase their nutritional valve, For example, ham is enriched with protein derived fcom legumes and fruit juices aze enriched with vite ‘mins. Edible insects area great material for food fortification for several reasons, First ofall, they are rich in protein of high biological value with 2 good amino acid profile and a high level of digestibility Moreover, insects are a good source ofa variety of micronutrients such as minerals: copper, iron, magnesium, manganese, phosphorous, sle nium, and zine and vitamins: riboflavin, pantothenic acid, biotin, and folic acid (Ramos-Florduy et a, 2012; Rumpold & Schluter, 2013). “Their lipid profile is desirable for humans. They are a source of um saturated fatty acids, for example omega-3 (Zielska et l., 2015). Given their nucitional value, insects can be a good product {or food supplementation and entomophagy does not have to be associated with ‘he consumption of whole insects any more. In this study, three species of insects (Tenebrio moliter, Schistocerca, gregaria, Grylldes sigilans) were selected, which are well knovn and esay to breed in Furope; each of them belongs to different order oF family and is bred widely in Europe. These species have also been re ported to have the biggest potential to be used as food and feed in the BU (EPSA, 2015). Moreover, the nutritional value of these insects was Hibrvtane: 6 sions, Gods ilu S pega, Seiterea gear: Tle, Ten malar TNBS, pylon ae: ONE, al Kalding capac, WHC water Baleg ‘ape foaming capacity 13, fam cerrepending tor [mod ed cna len 9 (Zeit it EA emaien ati, emelnon abystudied, The most important information for determination of the functional properties is the protein content, and the studied species ‘contain 52.35, 76.0, and 70.0% of protein, respectively (Zielisska eal, 2015). The physleochemieal properties of proteins, protein size, and flexibility play an important role in determining their functional properties, for example small molecular weight proteins give very good ‘emilsionsforming abilities because of rapid diffusion to the interface Proteins are commonly used to improve the functional properties of food compositions. in faet, the functional properties of proteins are dependent on pH ‘The aim of this study was to determine the functional properties of fours and protein preparations obtained (rom edible insects. this study, the solubility, water and oil holding capacity, and foaming and ‘emulsifying properties were determined. 2, Materials and methods 2.1, Raw materials The mealworme Tenebrio molar (Linnaeus, Coleoptera ‘Tenebrionidae) (larvae), locusts Schistocerca gregeria (Forskal, Orthoptera: Acrididae) (adult), and crickets Grylodes sigilans Cabriius, Orthoptera: Grylidse) (adult) were obtained from a com- mercial supplier from Poland. All individuals of these species were fasted for approximately 48) to clear ther gastrointestinal tract of any residual food, For each species tested, approximately 0.5kg of material ‘was frozen and lyophilized. The insects were ground in a laboratory srinder 2.2. Method for obtaining the protein preparation Proteins were isolated according to the Girés-Calle, Alaiz, and) Viogue (2010) method with slight modification. Briefly, insect Nour was stired for 1 with 0.2% NaQH ata ratio of 1:10 (w/¥), pH 11, at room temperature. After centrifugation at 8,000g, precipitation of proteins vias carried out at the isoelectric point pl 4.5 and room temperature. Precipitated proteins were centrifuged at 4°C for 2ominat 8000 g and washed with distilled water. Afterwards, the protein preparations were lyophilized and kept at ~18 °C until further analysis. 23. Solubily ‘The protein solubility was determined according to the method of Castellani, Martine, David-Briand, Guérin-Dubiard, and Anton (2003) with a slight modification. The sample was dispersed in distilled water and the pH ofthe mixture was adjusted (02, 3, 4, 5, 6,7, 8, 9, 10, and 11 using 1 or 6mol/L HCl and 1 or 6 mol/L NaOH. The volume of the mixture was adjusted to obtain the final concentration of protein (Lomg/mi. Total protein content inthe sample was determined after solubilization of the sample in O.Smol/L NaOH. The mixture was stirred for 90min and centrifuged at 8,000g for 15 min, The protein ‘content in the supernatant was determined sing the Bradford method (1976). Protein solubility was calculated from the formula: Solubility (96) = @®/P) x 100 ‘where: P,~ protein content inthe supernatant, P,— total protein content in the sample 24. Water holding capacity ‘Water holding capacity (WHC) was determined according te the method of Diniz and Martin (1997) with a slight modification. The sample (0.5 g) was dispersed in 20 ml of distiled water and stirred with 2 shaker at S40zpm for 30min. Afterwards, the dispersion was Lond Sine and Teco 91 (2018) 156-174 centrifuged at 8,000g for 18 min. The precipitate was weighed and the Aiference in the weight was calculated. The results were presented as gram of water absorbed per gram of the sample. 25. Oil holding capacity Oil holding capacity (OC) was determined according to the sethod of Haque and Mozaffar (1992) witha sight modification. The sample (0.5 g) was added to 10 of vegetable oi and mixed for 30 in a vortex mixer. Afterwards, the dispersion was centrifuged at 8,000 for 15min. The precipitate was weighed and the difference inthe weight ‘was calculated. The results were presented as gram of oll absorbed per ram of the sample, 26, Roaming properties Foaming capacity (FC) and foam stability (FS) were determined according (othe method of Guo etal. (2015). Tweaty milliiter of 1% sample was homogenized in a high shear homogenizer mixer (pol-eko |H500, Poland) ata speed of 16,000 rpm for 2 min. The whipped sample was immediately ransferred into a cylinder. The total volume was read at time zero and 30min after homogenization. The foaming capacity and foam stability were calelated fcom the formals [ev-vy/¥1 x 100 Wag Va) x 100 oaming capacity (FC) (4) foam stability (FS) 94) Where: V = volume before whipping (ml), Vo = volume after whipping (ia), Vag~ volume after standing (ml), 27, Bmulsifying properties Emulsifying properties were determined according to the method of ‘Wu, Wang, Ma, and Ren (2008). The sample was dispersed in distilled water (196 w/) and 15 ml ofthe dispersion were homogenized (po-eko 1500, Poland) with 15 ml of vegetable ol ata speed of 20,000 7pm for ‘min. Afterwards, the samples were centrifuged at 3000 g for Smin and the volume of the individual layers were read. Emulsion stability was evaluated by heating the emulsion for $0 min at 80°C, Then, the samples were centrifuged at 3000 g for Sin. Emulsion activity and emulsion stability were calculated from the formula imulsion activity (A) (86) = (V«/V) x 100 rmlsion stability (ES) (8) = (Vso/VO % 100 Where: ¥- total volume of tube contents, Ve -volume of the emulsified layer, Vao~ volume ofthe emulsified layer after heating 28, The sensory evaluation ‘The panel for sensory analysis was composed of 75 members aged from 21 to 30 years (58 women, 17 men). The characteristics of the ‘ours and protein preparations, such as color, consistency, smell, and overall acceptability were evaluated on a scale of 1-5 (I-bad, S-very 00d), 29. Statistical analysis All experiments were run in triplicate and the results were presented as means © standard deviation. Statistical analysis was performed tusing the STATISTICA v. 10.0 for one-way analysis of variance (ANOVA) and the differences of the means berween the samples were determined using the Tukey test, P-values below 0.05 were considered significantaioe tat 3. Results and discussion Functional properties of proteins are determined by the specific properties oftheir molecules such a size, structure, or configuration GGlara & Harwalker, 1996). In an earlier study, Zilissks, Baraniak, sd Karas (2017) obtained the protein profile of insect flour. The pro: teins from three insect species exhibited a varied range of molecular ‘mass. The Schistocerca gregaria showed too major proteins with mole- ‘ular mass of ~36kDa and ~97 kDa, Grylodes silts had ~ 35.3 kDa ‘and ~116kDa proteins, while the Tenebrio molitor protein profile did not show dominant proteins. Nevertheless, the bands observed inthe T. ‘molto protein profile ranged from 6.5 to 95 kDa. The differences inthe ‘molecular mass clearly differentiated the functional properties of the studied insect species. ‘Functional properties can be also associated with a varying amount ‘of protein and other constituents in flours. The protein content in the studied species was af follows: T. molitor 52.358, G. sigllaus 70.0%, and 5. gregaria 76.0% The fat content was 24.7%, 18.23%, and 12.97%, respectively, and the content of carbohydrates was 2.2%, 0.1%, and 1.7%, respectively (Ziel et 4), 2015), The amino acid and fatty ‘acid spectra for the studied species were presented in a previous study elisha et a, 2015). 2.1. Solty ‘The results forthe protein solubility are presented in Fig. 1. The protein solubility ofall samples shoved minimum values at around pH 5 with values of 3% for T. malitor, 4% for G.sigilars, and 8% for S aregoria. A significant increase in protein solubility was observed ‘around pll 8 for T. moltor and 6, sgilats, and around pl 9 for S, asregoria. The bighest solubility ofall samples was noticeable at pH 11 (7 molitor - 97%, G.sigilaus ~ 9686, and S. gregaria 90%), but a high solubility value was shown also at pI 2 and 3 for T: motor (86 and 5286, respectively) and S gregaria (87 and 69%, respectively), and at pH 2,3, and 4 for 6. sgilats (72, 95, and 57%, respectively). These results correspond well with those obtained by Zhao, Vézquex-Gutiérrez, ‘Johansson, Landberg, and Langton (2016) in similar assay conditions Dut for defatted 1: moltor protein extraction, Generally, the insect protein solubility was similar to data reported for legumes, for example kidney bean flour (Wani, Sogi, Wanl, & Gill, 2013), especially the Kidney bean globulin faction (und & Aluko, 2012) and other plants like the Ginko biloba seed albumin fraction (Deng et 81, 2011) or fe hugteek (Hl Nast! & 1 Tinay, 2007) It should be noted that the sol bility of the materials mentioned above was similar although the ai thors used different dilutions of the samples and the protein was determined with different methods, Solubility is one of the most im portant physicochemical and functional properties of protein and de pends on hydration and the degree of hydrophobicity of protein mo: ecules (Sathe & Salunkhe, 1881). Good solubility of proteins is ‘important in many uses, mainly for formation of emulsions, Foams, and sey 8) ceeesssased Fig. 1. Protein slubiy wth anges at ph 2-1, Teners motor uae), Os elas (ang), Scere pegs ele). Lond Sine and Teco 91 (2018) 156-174 ‘water holding capacity (e/e) Tenebrio molter Gros sigilus _Schistoerea gregaria ‘oll holding capacity (e/s) Tenebrio moltor Grades sgitats Fe, 2 Wate blding capac (A) andl lding capac (Bf Inet poe pre ratons gle) and noe ac (ark Difeent ters ete sigue ierece = 003) Schistocerca gregaria sels in designed food products. As a result, molecules in colloidal sys tems are homogeneously dispersed, which improves the interfacial properties (23925, 1997), 3.2, Water holding capacity ig, 2 shows the result of the water holding capacity. Higher water holding capacity was noticeable for the protein preparations than the whole inseets and the highest water holding eapacity was noted in the Tenebrio molitor protein preparation (3.95 g/g), while the lowest value ‘was noted for the whole ground T. molitor (129 g/). This is important Information for the use of these forms in food industry, The big dif ference in water holding capacity between protein preparations and Insect lour might bea good indicator ofthe applications ofthese forms for diferent food products. An opposite situation was observed in the case of Schistocerca gregaria ~ similar WH values were noted for the protein preparation and the insect flour (231 g/g and 2.18 4/, ce spectively). This is also important information that could be helpful in the analysis ofthe cost-ffectiveness and benefits brought by the use of fone of the presented forms of insects. Protein isolates are obviowsly richer in protein than whole insects but do not contain vitamins,