Analysis of the Composition of Different Instars of Tenebrio molitor Larvae using Near-Infrared Reflectance Spectroscopy for Prediction of Amino and Fatty Acid Content.

Insects are a sustainable protein source for food and feed. The yellow mealworm (Tenebrio molitor L.) is a promising candidate for industrial insect rearing and was the focus of this study. This research revealed the diversity of Tenebrio molitor larvae in the varying larval instars in terms of the nutritional content. We hypothesized that water and protein are highest in the earlier instar, while fat content is very low but increases with larval development. Consequently, an earlier instar would be a good choice for harvest, since proteins and amino acids content decrease with larval development. Near-infrared reflectance spectroscopy (NIRS) was represented in this research as a tool for predicting the amino and fatty acid composition of mealworm larvae. Samples were scanned with a near-infrared spectrometer using wavelengths from 1100 to 2100 nm. The calibration for the prediction was developed with modified partial least squares (PLS) as the regression method. The coefficient for determining calibration (R2C) and prediction (R2P) were >0.82 and >0.86, with RPD values of >2.20 for 10 amino acids, resulting in a high prediction accuracy. The PLS models for glutamic acid, leucine, lysine and valine have to be improved. The prediction of six fatty acids was also possible with the coefficient of the determination of calibration (R2C) and prediction (R2P) > 0.77 and >0.66 with RPD values > 1.73. Only the prediction accuracy of palmitic acid was very weak, which was probably due to the narrow variation range. NIRS could help insect producers to analyze the nutritional composition of Tenebrio molitor larvae fast and easily in order to improve the larval feeding and composition for industrial mass rearing.

Influence of Dietary Protein Content on the Nutritional Composition of Mealworm Larvae (Tenebrio molitor L.).

The use of insects as livestock feed is becoming increasingly accepted because they provide an important source of protein. The purpose of this study was to investigate the chemical composition of mealworm larvae (Tenebrio molitor L.) reared on a range of diets that differed in nutritional composition. Focus was placed on the influence of dietary protein content on larval protein and amino acid composition. For the experimental diets, wheat bran was chosen as the control substrate. The following types of flour-pea protein, rice protein, sweet lupine, and cassava, as well as potato flakes, were mixed with wheat bran and used as the experimental diets. An analysis of the moisture, protein, and fat content was then carried out for all diets and larvae. Furthermore, the amino acid profile was determined. It was shown that supplementing the feed with pea and rice protein was most suitable in terms of high protein yield in larvae (70.9-74.1% dry weight) with low fat content (20.3-22.8% dry weight). The total amino acid content was highest in larvae that were fed with a mixture of cassava flour and wheat bran (51.7 ± 0.5% dry weight), as well as the highest content of essential amino acids (30.4 ± 0.2% dry weight). Moreover, a weak correlation between larval protein content and diet was identified, yet a stronger influence of dietary fats and carbohydrates on larval composition was found. This research could result in improved formulations of artificial diets for Tenebrio molitor larvae in the future.

Near-Infrared Reflectance Spectroscopy for Quantitative Analysis of Fat and Fatty Acid Content in Living Tenebrio molitor Larvae to Detect the Influence of Substrate on Larval Composition.

Several studies have shown that mealworms (Tenebrio molitor L.) could provide animals and humans with valuable nutrients. Tenebrio molitor larvae were studied to determine whether their rearing diets affected their fat and fatty acid content and to ascertain if it is possible to detect the changes in the larval fat composition using near-infrared reflectance spectroscopy (NIRS). For this reason, a standard control diet (100% wheat bran) and an experimental diet, consisting of wheat bran and the supplementation of a different substrate (coconut flour, flaxseed flour, pea protein flour, rose hip hulls, grape pomace, or hemp protein flour) were used. The results showed lesser weight gain and slower growth rates for larvae raised on diets with a high fat content. A total of eight fatty acids were identified and quantified, where palmitic, oleic, and linoleic acids were the most prevalent and showed a correlation between larval content and their content in the rearing diets. There was a high content of lauric acid (3.2-4.6%), myristic acid (11.4-12.9%), and α-linolenic acid 8.4-13.0%) in mealworm larvae as a result of the high dietary content of these fatty acids. NIR spectra were also influenced by the fat and fatty acid composition, as larval absorbance values differed greatly. The coefficient of the determination of prediction (R2P) was over 0.97, with an RPD value of 8.3 for the fat content, which indicates the high predictive accuracy of the NIR model. Furthermore, it was possible to develop calibration models with great predictive efficiency (R2P = 0.81-0.95, RPD = 2.6-5.6) for all fatty acids, except palmitoleic and stearic acids which had a low predictive power (R2P < 0.5, RPD < 2.0). The detection of fat and fatty acids using NIRS can help insect producers to quickly and easily analyze the nutritional composition of mealworm larvae during the rearing process.

Self-Selection of Feeding Substrates by Tenebrio molitor Larvae of Different Ages to Determine Optimal Macronutrient Intake and the Influence on Larval Growth and Protein Content.

Nutrient self-selection was used to determine the optimal uptake of macronutrients by the yellow mealworm (Tenebrio molitor) larvae. The selection study consisted of four combinations of eight pelleted substrates from a total choice of 25, available to the larvae in a multiple-choice arena. In order to be able to determine the nutrient requirements as a function of the larvae age, six, eight and tenweekold larvae were used for the experiment. The larvae were free to choose between the different feeds for a period of two weeks. Rearing took place at 27 °C, 75% relative humidity and under dark conditions. The optimal ratios of macronutrients were 67.3 to 71.5% for carbohydrates, 19.9 to 22.8% for proteins and 8.6 to 10.0% for lipids to ensure the best results. Biomass growth, food intake and conversion were positively influenced to a significant extent by carbohydrate intake. The protein content, too, varied according to the macronutrient intake and substrate composition; a higher protein consumption increased the larval protein content. Wheat bran and flour, oat bran and flakes, maize hulls, lupine flour and potato flakes, in particular, were considered suitable substrates for the feeding and rearing of Tenebrio molitor larvae and highlighted that these larvae preferred a grain-based diet.

Determination of Moisture and Protein Content in Living Mealworm Larvae (Tenebrio molitor L.) Using Near-Infrared Reflectance Spectroscopy (NIRS).

Yellow mealworm larvae (Tenebrio molitor L.) are a sustainable source of protein for food and feed. This study represents a new approach in analyzing changes in the nutritional composition of mealworm larvae using near-infrared reflectance spectroscopy (NIRS) combined with multivariate analysis. The moisture and protein content of living larvae were scanned with a near-infrared spectrometer using wavelengths from 1100 to 2100 nm. Different feeding groups with varying moisture sources and amount and the difference between low (50%) and high (75%) humidity were tested, and the influence on larval moisture and protein content was measured. A calibration was developed, with modified partial least squares as the regression method. The NIR spectra were influenced by the moisture and protein content of the larvae, because the absorbance values of the larval groups differed greatly. The coefficient of the determination of calibration (R2c) and prediction (R2p) were over 0.98 for moisture and over 0.94 for protein content. The moisture source and content also had a significant influence on the weight gain of the larvae. Consequently, significant differences in protein content could be determined, depending on the water supply available. With respect to wet weight, the larvae moisture content varied from 60 to 74% and protein content from 16 to 24%. This investigation revealed that with non-invasive NIRS online monitoring, the composition of insects can be continuously recorded and evaluated so that specific feeding can be carried out in the course of larval development and composition.

Systematic Studies on the Antioxidant Capacity and Volatile Compound Profile of Yellow Mealworm Larvae (T. molitor L.) under Different Drying Regimes.

The yellow mealworm (Tenebrio molitor L., Coleoptera: Tenebrionidae) is an edible insect and due to its ubiquitous occurrence and the frequency of consumption, a promising candidate for the cultivation and production on an industrial scale. Moreover, it is the first insect to be approved by EFSA 2021 following the Novel Food Regulation. Industrial production of mealworms necessitates optimized processing techniques, where drying as the first postharvest procedure is of utmost importance for the quality of the final product. The focus of the present study was to analyse the chemical composition, antioxidant capacity, volatile compound profile and colouring of mealworm larvae dried in various regimes (freeze-drying, microwave drying, infrared drying, rack-oven drying and high-frequency drying). Proximate composition and fatty acid profile were similar for all dried larvae. Freeze dried larvae were predominantly marked by lipid oxidation with significantly higher peroxide values, secondary/tertiary oxidation products in the headspace GC-MS profiles and lower antioxidant capacity. High-temperature treatment in the rack oven-and to some extent also infrared or microwave drying-led to mealworm larvae darkening and the appearance of volatile Maillard secondary products such as 2-methylpropanoic acid, 2-/3-methylbutanoic acid and alkylpyrazines. High-frequency drying as a new emerging technology in insect processing was the most cost-effective method with energy costs of solely 0.09 Є/kg T. molitor L. leading to final larval material characterized by both lipid oxidation and nonenzymatic Maillard-browning.

Metabolic fate and toxicity reduction of aflatoxin B1 after uptake by edible Tenebrio molitor larvae.

The use of insects as food and feed is gaining more attention for ecological and ethical reasons. Despite the high tolerance of edible yellow mealworm (Tenebrio molitor) larvae to aflatoxin B1 (AFB1), the metabolic fate of the toxin along with its toxic potential in the insect is uncertain. The present study aimed at investigating the AFB1 mass balance and the metabolite formation in a feeding trial with AFB1-contaminated grain flour. T. molitor larvae tolerated the AFB1 level of 10,700 µg/kg in the feed, however, weight gain was decreased by 15% over a 4-weeks feeding period. The investigation of the phase I metabolite pattern revealed the formation of AFM1 and a novel presumably monohydroxylated compound in larvae extracts that was not formed by reference incubation with rat, bovine or porcine liver microsomes. Mass balance quantification of ingested AFB1 revealed that 87% of the initial toxin remain undetected in larval body or residue. Analysis of histone H2Ax phosphorylation in human liver cells as a surrogate for genotoxicity showed that extracts from exposed larvae did not exhibit an elevated toxic potential. Although toxicological uncertainties remain due to the undetected transformation products, the resulting mutagenicity of the edible larvae appears to be low.

Dietary zinc enrichment reduces the cadmium burden of mealworm beetle (Tenebrio molitor) larvae.

The industrial production of Tenebrio molitor L. requires optimized rearing and processing conditions to generate insect biomass with high nutritional value in large quantities. One of the problems arising from processing is a tremendous loss in mineral accessibility, affecting, amongst others, the essential trace element Zn. As a feasible strategy this study investigates Zn-enrichment of mealworms during rearing to meet the nutritional requirements for humans and animals. Following feeding ZnSO4-spiked wheat bran substrates late instar mealworm larvae were evaluated for essential micronutrients and human/animal toxic elements. In addition, growth rate and viability were assessed to select optimal conditions for future mass-rearing. Zn-feeding dose-dependently raised the total Zn content, yet the Znlarvae/Znwheat bran ratio decreased inversely related to its concentration, indicating an active Zn homeostasis within the mealworms. The Cu status remained stable, suggesting that, in contrast to mammals, the intestinal Cu absorption in mealworm larvae is not affected by Zn. Zn biofortification led to a moderate Fe and Mn reduction in mealworms, a problem that certainly can be overcome by Fe/Mn co-supplementation during rearing. Most importantly, Zn feeding massively reduced the levels of the human/animal toxicant Cd within the mealworm larvae, a technological novelty of outstanding importance to be implemented in the future production process to ensure the consumer safety of this edible insect species.

Effect of Different Drying Methods on Nutrient Quality of the Yellow Mealworm (Tenebrio molitor L.).

Yellow mealworm (Tenebrio molitor L.) represents a sustainable source of proteins and fatty acids for feed and food. Industrial production of mealworms necessitates optimized processing techniques, where drying as the first postharvest procedure is of utmost importance for the quality of the final product. This study examines the nutritional quality of mealworm larvae processed by rack oven drying, vacuum drying or freeze drying, respectively. Proximate composition and fatty acid profile were comparable between the dried larvae. In contrast, larvae color impressions and volatile compound profiles were very much dependent on processing procedure. High-temperature rack oven drying caused pronounced darkening with rather low content of volatiles, pointing toward the progress of Maillard reaction. On the other hand, vacuum drying or freeze drying led to enrichment of volatile Maillard reaction and lipid oxidation intermediates, whose actual sensory relevance needs to be clarified in the future. Beyond sensory and visual importance drying intermediates have to be considered with regard to their metal ion chelating ability; in particular for essential trace elements such as Zn2+. This study found comparable total zinc contents for the differently dried mealworm samples. However, dried larvae, in particular after rack oven drying, had only low zinc accessibility, which was between 20% and 40%. Therefore, bioaccessibility rather than total zinc has to be considered when their contribution to meeting the nutritional requirements for zinc in humans and animals is evaluated.

Feeding study for the mycotoxin zearalenone in yellow mealworm (Tenebrio molitor) larvae-investigation of biological impact and metabolic conversion.

Edible insects as additional food and/or feed source may represent one important component to solve the problem of food security for a growing human population. Especially for covering the rising demand for protein of animal origin, seven insect species currently allowed as feed constituents in the European Union are gaining more interest. However, before considering insects such as yellow mealworm larvae (Tenebrio molitor) as suitable for, e.g. human consumption, the possible presence and accumulation of contaminants must be elucidated. The present work investigates the effects of the mycotoxin zearalenone (ZEN) and its metabolites on insect larvae. Seven different diets were prepared: toxin-free control, spiked and artificially contaminated (both containing approx.500 µg/kg and approx. 2000 µg/kg of ZEN) as well as two naturally contaminated diets (600 µg/kg and 900 µg/kg ZEN). The diets were used in a multiple-week feeding trial using T. molitor larvae as model insects. The amount of ZEN and its metabolites in the feed, larvae and the residue were measured by HPLC-MS/MS. A significantly enhanced individual larval weight was found for the insects fed on the naturally contaminated diets compared to the other feeding groups after 8 weeks of exposure. No ZEN or ZEN metabolites were detected in the T. molitor larvae after harvest. However, ZEN, α- and ß-stereoisomers of zearalenol were found in the residue samples indicating an intense metabolism of ZEN in the larvae. No further ZEN metabolites could be detected in any sample. Thus, ZEN is not retained to any significant amount in T. molitor larvae.

Basic aspects of phase changes under high pressure.

Some substances of technological importance reveal phase change phenomena in the pressure and temperature range typically applied in biotechnology and food processing. For example, media with high molar volumes like edible oils and fats undergo liquid-solid phase transition at pressure increases up to several hundred megapascals. This article is concerned with theoretical considerations of the line of coexistence of solid and liquid phases in the pressure and temperature domain that corresponds to the phase boundary as a function of temperature and pressure. A universal model equation based on the equilibrium thermodynamics allowing prediction of the phase transition line of homogeneous media with high molar volume is presented. Approximate solutions of the model equation are discussed, which allow the phase boundary of substances with high molar volume to be described by a linear relation at least sequentially. The methods of experimental determination of the phase boundary under high pressure are presented and an attempt is made to validate the theoretical model with respect to the experimental data.