Effect of Cereal α-Amylase/Trypsin Inhibitors on Developmental Characteristics and Abundance of Digestive Enzymes of Mealworm Larvae (Tenebrio molitor L.)

Abstract

The objective of this work was to investigate the potential effect of cereal α-amylase/trypsin inhibitors (ATIs) on growth parameters and selective digestive enzymes of Tenebrio molitor L. larvae. The approach consisted of feeding the larvae with wheat, sorghum and rice meals containing different levels and composition of α-amylase/trypsin inhibitors. The developmental and biochemical characteristics of the larvae were assessed over feeding periods of 5 h, 5 days and 10 days, and the relative abundance of α-amylase and selected proteases in larvae were determined using liquid chromatography tandem mass spectrometry. Overall, weight gains ranged from 21% to 42% after five days of feeding. The larval death rate significantly increased in all groups after 10 days of feeding (p < 0.05), whereas the pupation rate was about 25% among larvae fed with rice (Oryza sativa L.) and Siyazan/Esperya wheat meals, and only 8% and 14% among those fed with Damougari and S35 sorghum meals. As determined using the Lowry method, the protein contents of the sodium phosphate extracts ranged from 7.80 ± 0.09 to 9.42 ± 0.19 mg/mL and those of the ammonium bicarbonate/urea reached 19.78 ± 0.16 to 37.47 ± 1.38 mg/mL. The total protein contents of the larvae according to the Kjeldahl method ranged from 44.0 and 49.9 g/100 g. The relative abundance of α-amylase, CLIP domain-containing serine protease, modular serine protease zymogen and C1 family cathepsin significantly decreased in the larvae, whereas dipeptidylpeptidase I and chymotrypsin increased within the first hours after feeding (p < 0.05). Trypsin content was found to be constant independently of time or feed material. Finally, based on the results we obtained, it was difficult to substantively draw conclusions on the likely effects of meal ATI composition on larval developmental characteristics, but their effects on the digestive enzyme expression remain relevant.

Effect of Sample Preparation on the Detection and Quantification of Selected Nuts Allergenic Proteins by LC-MS/MS

“…The extraction protocol using ammonium bicarbonate buffer containing urea is the one optimized and applied in our previous work [ 28 , 49 , 50 ]. Briefly, 100 mg of each defatted nut sample were mixed with 1.5 mL of extraction buffer (100 mM ammonium bicarbonate containing 4 M urea) and the whole was shaken at 95 rpm at room temperature for 60 min, followed by the centrifugation (10,000× g , 4 °C for 5 min).…”

“…Candidates with less than 95% protein sequence similarity to other species were in-silico digested with the Skyline software using trypsin as enzyme. The additional settings are described in our previous works [ 28 , 49 , 50 ]. The species selectivity of the identified individual nut peptides was analyzed once more by the BLAST algorithm (UniProt Universal Protein Resource, ; accessed on 13 June 2021).…”

Insect α-Amylases and Their Application in Pest Management

“…The inhibition of the α-amylase of Coleoptera was stronger than that of human salivary α-amylase [73]. Cereal proteins are the main nutritional proteins of Tenebrio molitor L. larvae; the impact of CTI in food is proven to be correlated to the expression of digestive enzymes, whereas it is currently difficult to determine their potential impact on the developmental characteristics of larvae [74].…”

Structure, Function and Protein Engineering of Cereal-Type Inhibitors Acting on Amylolytic Enzymes

“…A recent review addresses the inhibition of different insect a-amylases by plant proteinaceous inhibitors including CM-proteins from wheat, rye, corn and barley (da Lage, 2018). A more practical approach consists in application of artificial diets containing either recombinantly produced inhibitors or efficient plant fractions to reduce viability of herbivorous insects (Dias et al, 2005;da Silva et al, 2013;Sagu et al, 2021). For crops, the backside of this strategy can be negative consequences of higher levels of CTIs on human health and in feed for livestock.…”