Solubility profiles of two-spotted cricket (Grylluss bimaculatus De Geer, 1773) protein powder: Roles of developmental stage, gender, processing, pH level

Vo Lam^1,2, Tran Nghia Khang^1,2, Le Thi Thuy Diem^1,2, Vo Phương Mai^1,2, Nguyen Thi Bich Hanh^1,2 and Bui Phan Thu Hang^1,2

¹ An Giang University, An Giang Province, Vietnam
² Vietnam National University Ho Chi Minh City, Vietnam
bpthang@agu.edu.vn

Abstract

This study evaluated the appropriate age, gender, post-harvest processing, pH level to obtain two-spotted cricket powder with high solubility of protein content for food applications. Crickets were harvested at 4, 5, 6, 7 weeks of age, separated by gender, processed via blanching or left unblanched before freeze-drying. Soluble protein concentration was measured at pH 5, 7, 8 using the Bradford assay.

Protein solubility was influenced by age, gender, processing methods. The highest protein solubility was observed in 5-week-old both male and female cricket powder processed by blanching, particularly at pH 7-based food applications.

Keywords: edible insects, functional protein, processing method, protein solubility

Introduction

The increasing global demand for sustainable protein sources has brought increasing attention to edible insects such as crickets (Acheta domesticus), which offer high nutritional value, efficient feed conversion, low environmental impact (FAO 2013; Huis et al 2013). Cricket powder, derived from whole insect processing, has gained interest for its high-quality protein, vitamins, bioactive compounds, supporting its use as a viable ingredient in functional foods and animal feeds (Mitchaothai et al 2022).

Among the key functional properties of protein ingredients, solubility is particularly important because it influences other techno-functional properties such as emulsification, gelation and digestibility (Zayas 2012; Hettiarachchy et al 1994). The solubility of insect proteins is known to vary with biological factors such as developmental stage and gender, as well as with post-harvest processing techniques including blanching and drying (Finke et al 1985; Bui Phan Thu Hang et al 2020).

Additionally, the pH of the surrounding medium plays a significant role in determining the net charge and aggregation behavior of proteins, thereby affecting solubility (Compton et al 1985). However, limited research has comprehensively evaluated how these factors interact to influence soluble protein yield in cricket powder.

Therefore, this study aimed to investigate the effects of developmental stage (age), gender, post-harvest processing (blanching) on the protein solubility of Gryllus bimaculatuspowder across a range of pH levels. The findings are expected to contribute to the optimized processing and utilization of cricket powder in functional food systems and feed applications.

Materials and methods

Location, source of crickets and design

The study was conducted at HLF/Cricket Farm in Long Xuyen City, An Giang Province, Vietnam. Laboratory analyses were carried out at the Laboratory of An Giang University, under Vietnam National University- Ho Chi Minh City. The crickets (n = 7 200 pinheads) at 7 days old raised in plastic containers of 57 cm x 38 cm x 21 cm, given with cardboard egg cartons for hiding and moulting areas. They were randomly assigned to 24 plastic containers and harvested at 4, 5, 6, 7 weeks of age, respectively.

The experimental design followed a 2 × 4 × 2 factorial structure: processing methods (blanched and unblanched), ages (4, 5, 6, 7 weeks), genders (male and female), with three replicates per treatment. Crickets were fed on a concentrate with 22% crude protein content and fresh sweet cassava leaves. The feed and water were supplied ad libitum. The rearing procedure was similar during the experiment.

Sample preparation

Crickets (Gryllus bimaculatus) were harvested at four developmental stages: (before the last moulting), 5 (adult), 6 (bearing eggs) and 7 (laying eggs) weeks of age. For each age group, male and female crickets were separated. Each gender-age group was divided into two subgroups: one subjected to blanching in hot water at 80°C for 5 minutes, one left unblanched. All samples were immediately frozen at −80°C, freeze-dried to moisture content below 5%, ground into powder using a blender. The powders were vacuum-packed and stored at −20°C until analysis.

Chemical analysis

Soluble protein content was determined using the Bradford assay as described by Compton and Jones (1985). Samples were dissolved in buffer solutions at pH 5, 7, 8, centrifuged, the protein concentration of the supernatant was measured spectrophotometrically.

Statistical analysis

Statistical analysis was performed using the General Linear Model (GLM) in Minitab 17. Mean differences were considered significant at p<0.05 using Tukey’s test. Polynomial equations were fitted to the changes for live weight gain and the conversion using the Excel option in Microsoft software.

Results and discussion

Effects of ages, genders and processing methods on protein solubility of cricket powder at pH 5

Protein solubility varied significantly across age, gender, processing methods, aligning with known physicochemical characteristics of insect proteins as influenced by developmental stage and thermal treatment (Finke et al 1985; Zayas 2012).

At pH 5, protein solubility was the lowest across all treatments, consistent with the proximity to the isoelectric point of many insect proteins, where solubility naturally decreases due to minimal net charge (Hettiarachchy et al 1994). Nonetheless, both age and processing influenced these values. Unblanched female samples harvested at 7 weeks recorded the highest solubility (1.50 mg/mL), while younger samples (4WO) exhibited significantly lower solubility (Table 1), suggesting maturation contributes to higher water-soluble protein content.

Effects of ages, genders and processing methods on protein solubility of cricket powder at pH 7

At pH 7, protein solubility peaked for most groups. Female cricket powder from unblanched samples at 5 weeks yielded the highest solubility (1.67 mg/mL), a trend also visible in the blanched samples. This aligns with observations that the early adult phase (around 5 weeks) marks a physiological stage where protein synthesis and accumulation are highest (Kaewtapee et al 2024).

Unblanched samples consistently retained higher solubility at pH 7 across all age groups compared to blanched ones (Table 2, Figure 1- 4). This suggests that blanching, despite reducing microbial load, may compromise functional protein integrity – a point supported by Mitchaothai et al (2022), who noted a decline in water-holding capacity in thermally processed cricket flour.

Figure 1. Protein solubility at pH 7 for female crickets blanched	Figure 2. Protein solubility at pH 7 for male crickets blanched
Figure 3. Protein solubility at pH 7 for female crickets blanched	Figure 4. Protein solubility at pH 7 for male crickets unblanched

Effects of ages, genders and processing methods on protein solubility of cricket powder at pH 8

Protein solubility remained relatively high at pH 8, but was more affected by age and gender interaction (Table 3). The reduced solubility in blanched samples supports the hypothesis that thermal denaturation disproportionately affects protein solubility at basic pH levels (Compton et al 1985).

Female crickets generally demonstrated higher soluble protein yields compared to males, possibly due to differing body composition, reproductive physiology, or variation in cuticle development rates (Bui Phan Thu Hang et al 2020; Vo Lam et al 2023).

Conclusion

This study demonstrates that soluble protein content in two-spotted cricket (Gryllus bimaculatus) powder is influenced by developmental stage, gender, post-harvest processing methods. Cricket powder both genders from 5-week-old crickets processed by blanching has high protein solubility at pH 7.

The findings underscore the importance of selecting appropriate harvesting time and processing techniques to optimize the functional protein content of cricket powders. These insights provide practical guidance for the development of high-protein, functional insect-based ingredients in the food and feed industry.

Acknowledgement

The authors are grateful for the financial for this research from Vietnam National University Ho Chi Minh City (VNU-HCM) under grand number C 2023-16-05/HĐ-KHCN. We also acknowledge for the infrastructure support for this research from a smallholder cricket farm in Long Xuyen city, An Giang province.

References

Bui Phan Thu Hang, Nguyen Van Cop and Vo Lam 2020 Effect of cassava leaves (Manihot esculenta Crantz), water spinach (Ipomoea aquatica) and Coccinia grandis L. on biomass growth of crickets (Gryllus bimaculatus) fed chicken feed as the basal diet. Livestock Research for Rural Development. Volume 32, Article #28.Retrieved May 14, 2024, from http://www.lrrd.org/lrrd32/2/bpthan32028.html

Compton S J and Jones C G 1985 Mechanism of dye response and interference in Bradford protein assay. Analytical Biochemistry, 151(2):369–374.

FAO 2013 Edible insects: Future prospects for food and feed security. FAO Forestry Paper 171. Rome, Italy.

Finke M D, DeFoliart G R and Benevenga N J 1985 Use of a four-parameter logistic model to evaluate the quality of insect protein. Journal of Nutrition. 115(5):703–710.

Hettiarachchy N S and Ziegler G R 1994 Protein functionality in food systems. Marcel Dekker, New York.

Huis A V, Itterbeeck J V, Klunder H, Mertens E, Halloran A, Muir G and Vantomme P 2013 Edible insects: future prospects for food and feed security. FAO. Rome.

Kaewtapee C, Triwai P, Inson C, Masmeatathip R and Sriwongras P 2024 Effects of protein levels on production performance, nutritional values, phase feeding of two-spotted cricket. Journal of Insect Science, 24(2), p.18. https://doi.org/10.1093/jisesa/ieae039.

Mitchaothai J, Grabowski N T, Lertpatarakomol R, Trairatapiwan T, Chhay T, Keo S and Lukkananukool A 2022 Production performance and nutrient conversion efficiency of field cricket (Gryllus bimaculatus) in mass-rearing conditions. ANimal 12. https://doi.org/10.3390/ani12172263.

Vo Lam, Nguyen Thi Ut Em and Bui Phan Thu Hang 2023 Effect of rearing density on growth performance and survival rate of field cricket (Gryllus bimaculatus). Livestock Research for Rural Development. Volume 35, Article #39. Retrieved May 14, 2024, from http://www.lrrd.org/lrrd35/5/3539bpth.html

Zayas J F 2012 Functionality of proteins in food. Springer science & business media.