Characteristics of Oils Extracted from Yellow Mealworm (Tenebrio molitor L.) Dried with the Infrared-Convective Method

TL;DR

This study examines how drying yellow mealworms with infrared-convective methods affects the oils extracted from them, finding that pretreatment improves oil quality and yield.

Contribution

The novel contribution is the investigation of pulsed electric field pretreatment and infrared-convective drying on yellow mealworm oil properties.

Findings

PEF pretreatment increased oil extraction yield by up to 29.2%.

PEF and IR-CD drying lowered acid and peroxide values, increased MUFA content, and improved oxidative stability.

Nutritional properties of oils deteriorated despite improved quality indicators.

Abstract

Edible insects are a nutritionally attractive food product, also due to their high fat content and high levels of unsaturated fatty acids. In this work, the effect of pulsed electric field (PEF) pretreatment and infrared-convective (IR-CD) drying on the oil properties extracted from yellow mealworm (Tenebrio molitor L.) larvae was investigated. The oil from raw and dried insects was extracted via the Soxhlet method for yield determination and via the Folch method for quality analysis. The acid value (AV) and peroxide value (PV) via the titration method, fatty acid composition and its distribution in triacylglycerol (TAG) molecules via the gas chromatography method, calculation of nutritional value indices, oxidative stability via pressure differential scanning calorimetry (PDSC) method, and antioxidant activity of methanol extracts were examined. The results show that PEF may enhance the oil extraction yield from dried insects by up to 29.2%. The PEF treatment and drying method made insect oils more valuable by lowering their acid and peroxide values, increasing the MUFA content, and improving oxidative stability. Nonetheless, the nutritional properties of oils deteriorated. Our study demonstrated that oils extracted from dried yellow mealworm could be used as an ingredient in other food products to improve their nutritional value. However, more research in this area is needed to assess the impact on quality properties.

Figures

Full text

1. Introduction

Edible insects have emerged as a promising alternative food source to address common food security problems due to their high nutritional value, efficient feed conversion, and lower environmental footprint [1,2]. However, they are not yet widely accepted in the human diet, primarily due to cultural and psychological barriers [1]. One strategy to overcome this reluctance is to incorporate insect-derived ingredients into familiar food products. This can be achieved by introducing them in processed forms, such as powders or isolated fractions, e.g., protein or fat [3,4]. Insect protein and its application are frequently studied, while the fat fraction remains underexplored. During defatting procedures aimed at protein isolation, the fat fraction is removed [3], which opens up possibilities for its utilization. However, it is necessary to study it from both technological and nutritional perspectives. The fat content of dried yellow mealworm larvae ranges from 22.8% to 38.3% (on a dry basis) [5,6]. Based on the relevant literature [6,7,8,9,10], the fat fraction is mainly composed of unsaturated fatty acids (up to 77%), with major oleic (32.7–53.4%) and linoleic (17.5–38.4%) acids, and saturated palmitic acid (12.8–20.5%). Despite that, the fat fraction showed a higher percentage of n-6 PUFA than n-3 PUFA, resulting in a high n-6/n-3 ratio, ranging from 11.1 to 47.0 [11,12,13]. It is not advised for human health to have a high n-6/n-3 ratio, as it may contribute to the pathogenesis of numerous chronic diseases [14,15]. Despite its high content of unsaturated fatty acids, insect oil is not beneficial for nutritional reasons due to its high n-6/n-3 ratio and the distribution of fatty acids in triacylglycerols. This means it could be used as an ingredient in a food matrix or to produce new fats with desirable technological properties, rather than as a direct edible oil.

Extraction of the fat fraction traditionally relies on mechanical pressing or solvent extraction, such as the Folch method, which can be time-consuming and sometimes yield-limited [3]. To overcome these limitations, innovative extraction technologies have been developed and tested extensively. Among these, techniques such as ultrasound-assisted extraction (UAE), supercritical CO_2_ extraction (SFE-CO_2_), and microwave-assisted extraction are more commonly employed, especially for plant oils [16,17,18,19]. One of the promising techniques appears to be extraction after pulsed electric field (PEF) treatment. The PEF is a non-thermal method that involves applying short, high-voltage electrical pulses to the material placed between two electrodes. The use of a PEF causes cell membranes to become electroporated, altering their permeability, increasing the diffusivity of intracellular substances, enhancing the rate of mass transfer, and ultimately increasing the extraction yield [20,21]. So far, PEF treatment has been successfully applied to oil-rich plant matrices such as sunflower seeds, olives, sesame seeds, and microalgae [22,23,24,25]. Its application to edible insect oil extraction remains insufficiently investigated. Given the unique cellular structure and composition of insect biomass, extrapolations from plant systems may not fully apply, underscoring the need for specific studies.

Previous studies on insect oils have investigated mechanical pressing [26], and solvent extraction [27,28], as well as more recent approaches such as supercritical CO_2_ (SFE-CO_2_), ultrasound-assisted (UAE), or high-hydrostatic-pressure (HHPE) extraction [3,5,6]. However, the potential of emerging technologies, such as pulsed electric field (PEF) treatment, remains largely underexplored. Moreover, drying techniques significantly influence oil properties, yet available studies evaluate such drying methods as convective drying, freeze-drying, or microwave drying [9,10]. The effect of the infrared-convective method on insect oil properties is in its infancy.

The mechanism of PEF treatment has been demonstrated to have a beneficial effect on insect tissue properties. From this standpoint, it has the potential to reduce the drying time, thereby contributing to strategies to reduce carbon emissions. Furthermore, structural modifications have been shown to enhance the extraction of fat from the dried material. From a nutritional perspective, mealworm oil is characterized by a high proportion of unsaturated fatty acids; however, its direct impact on human health requires clinical investigations and remains beyond the scope of the present study. Therefore, the present work focuses on technological quality parameters and nutritional indices relevant for food applications rather than clinical health outcomes. In doing so, the present study aims to fill the knowledge gap by investigating how PEF pretreatment prior to infrared-convective drying of yellow mealworm (Tenebrio molitor L.) larvae affects the fatty acid composition, fatty acid distribution between the TAG positions, hydrolysis and oxidative stability, and antioxidant activity of the oils extracted therefrom.

2. Results

2.1. Oil Extraction Yield

Fat is a crucial component in insects, and insect species vary in their content. The extraction yield of oil from dried yellow mealworm without pretreatment was equal to 16.0 g/100 g d.m. (Table 1). This content was comparable to previously reported values, which ranged from 17.8 to 38.3 g/100 g d.m. [6,8,10]. Insect larvae are a very complex matrix, and their composition can be diverse due to species-specific traits (e.g., insect species, developmental stage, and sex of the insects), feed type and rearing conditions, and geographical location, but also due to the processing of the insects before fat extraction [29,30]. In addition, the method of fat extraction also affects its yield [27,28]. The use of PEF before IR-CD drying enhanced the oil extraction yield by up to 29.2% (Table 1).

In yellow mealworm larvae, fat is located in the fat body, consisting of cells containing multiple lipid droplets. Each lipid droplet is surrounded by a phospholipid monolayer, into which specific proteins are embedded or peripherally associated [31]. The use of PEF disrupted a phospholipid monolayer, resulting in a more homogeneous distribution of fat droplets with smaller diameters [32], perhaps more readily available for solvent extraction, and provided an enhanced oil extraction yield (Table 1). Furthermore, PEF may reduce tissue hardness. This allows better material fragmentation, including the fat body, and greater solvent penetration, as observed for convective-dried yellow mealworm larvae [8].

2.2. Acid Value and Peroxide Value

The quality of fat informs us about the impact of processing methods and the potential uses of this fat. The acid value (AV) is a measure of the content of free fatty acids, which are mostly formed as a result of the hydrolysis of triglycerides. Hence, it is a measure of the freshness of fat. In turn, the peroxide value (PV) measures the concentration of peroxides resulting from oxidation. Hence, this parameter serves as an indicator of the degree of rancidity in the fat [33,34]. The results for the AV and PV of fat extracted from raw and dried yellow mealworm are summarized in Table 1. The fat from raw insects was characterized by a significantly lower AV (11.21 mg KOH/g) than fats from dried insects (from 15.12 to 34.81 mg KOH/g). On the contrary, the PV of raw insect fat was significantly higher (1.77 meq O_2_/kg) than for fats extracted from dried samples (1.12 and 1.61 meq O_2_/kg). Among the tested fats isolated from dried samples, the highest AV and PV were noted for the sample without PEF pretreatment. The utilization of PEF significantly reduced the AV and the PV. There were reported some possible explanations for that behavior of the samples. First of all, this may be due to the diverse water content in the dried insects (data not published). The untreated and dried insects exhibited a higher water content (0.06 g H_2_O/g d.m.) than PEF-treated and dried insects (about 0.03 g H_2_O/g d.m.). Water molecules promote the hydrolysis of TAG ester bonds and the formation of free fatty acids [35]. Secondly, the application of PEF and electroporation can affect the activity of endogenous lipolytic enzymes present in insects [35]. As enzymes are proteins, higher PEF energy can cause partial damage to the protein-unfolding polypeptide chains and reduced activity [36], thereby reducing the hydrolysis of TAG ester bonds. It was proven, based on Pearson correlation, that the acid and peroxide values tend to correspond with the PEF energy intake (AV: r ≈ −0.774, PV: r ≈ −0.904). The lower AV and PV were achieved when higher PEF energy levels were used. Additionally, a strong correlation (r ≈ 0.739) was observed between acid and peroxide values.

Son et al. [13] obtained a lower AV (2.6 mg KOH/g) and a higher PV (3.5 meq O_2_/kg) for fat extracted from blanched and convective-dried yellow mealworm, while Jeon et al. [37] reported the PV from 5.0 to almost 10.0 meq O_2_/kg for fat isolated from freeze-dried and roasted yellow mealworm. There are currently no recommendations for the quality of fat isolated from insects. According to the EFSA’s Scientific Opinion [38], for fat extracted from blanched and thermally dried yellow mealworm, the acid value and peroxide value should be in the range of 0.5–4.8 mg KOH/g and 1.0–16.3 meq O_2_/kg, respectively. In turn, the Codex Alimentarius provides the acid value (4 mg KOH/g) and the peroxide value (15 meq O_2_/kg) as general quality guidelines for fats and oils. The obtained results of the acid value for extracted insect oils have exceeded this limit, which may suggest a higher than expected content of free fatty acids, especially in dried samples. This may result in biased taste of a product and excessive hydrolysis of oil [39]. The PV, however, was below the threshold value in every tested sample, which may indicate that PEF did not induce the oxidation process in the lipid fraction of the tested insects. Given that PEF treatment reduces the AV and PV, especially the AV, PEF application is recommended before IR-CD drying of insects. Although the acid values of oils extracted from dried material exceeded the limits established for edible vegetable oils, this does not preclude their application as food ingredients. An elevated AV indicates the presence of free fatty acids formed during drying, suggesting that further refining steps would be required prior to direct consumption. From a technological standpoint, such oils may still be suitable for incorporation into processed food systems or as raw materials for functional lipid fractions.

2.3. Fatty Acid Composition

Fats are compounds composed primarily of triacylglycerols (TAGs). Triacylglycerols mostly consist of fatty acids, which are divided into three groups: saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), and polyunsaturated fatty acids (PUFAs), depending on the type and number of bonds. The contribution of given fatty acids and fatty acid groups is connected to the characteristics of fat [40,41]. All the investigated insect oils were dominated by unsaturated fatty acids, with a predominance of monounsaturated fatty acids over polyunsaturated fatty acids (Figure 1).

In the SFA fraction, the most abundant fatty acid was palmitic (11.71–14.78%), in a lower percentage of myristic (3.17–5.49%) and stearic (2.36–2.94%) acids (Table 2). The oleic acid (C18:1 n-9c) was the main fatty acid from the MUFA group, comprising between 36.88% and 39.92%, with the lowest percentage (36.88%) in oil extracted from raw larvae (Table 2). Among the PUFA group, the dominant fatty acid was linoleic acid (C18:2 n-6c), with a relative percentage share ranging from 32.57 to 36.44%. The other fatty acid from the PUFA group was α-linolenic acid (C18:3 n-3) at the level of 1.40 to 1.76%.

The low relative percentage of this fatty acid should contribute to a satisfactory oxidative stability of the tested insect oils, as α-linolenic acid is characterized by rapid oxidation [42]. Considering the differences in the results, and based on previous findings, it could be due to free radical formation during PEF treatment and infrared radiation during drying, which may promote the oxidation of fatty acids from the PUFA group and other degradation transformations of TAG molecules [36]. When using insect oil as an ingredient in food products, PEF treatment is beneficial because it increases the MUFA content and decreases the PUFA content. A higher MUFA content may improve oxidative stability, which is beneficial during thermal food processing, such as baking or frying. However, the proportion of PUFA to MUFA content in the tested insect oils remains too high to use them as frying media. From a nutritional standpoint, a decreased PUFA content and increased SFA content in the tested insect oils are not beneficial.

Generally, our results are consistent with previous studies [7,11,12]. The differences may have been due to variations in feeding and processing conditions. Tzompa-Sosa et al. [27] found a higher percentage of palmitic (18.52%) and oleic (49.50%) acids in oil from freeze-dried yellow mealworm, while the contribution of linoleic (21.82%) and α-linolenic (0.84%) acids was lower. Lenaerts et al. [10] reported that fat isolated from microwave-dried and freeze-dried yellow mealworm contained a higher proportion of palmitic acid (17.92–20.52%) and a lower percentage of linoleic acid (23.15–30.37%). Also, Messina et al. [11] demonstrated a lower proportion of linoleic acid (25.73%).

2.4. Nutritional Value Indices

Three nutritional value indices calculated for insect oils are presented in Table 2. The nutritional and dietary values of insect oils were significantly affected by the PEF treatment and the drying method. The obtained oils outperformed raw insect oil in terms of nutritional characteristics, with lower atherogenicity index (AI) and thrombogenicity index (TI) values and a greater hypocholesterolemic-to-hypercholesterolemic (HH) ratio. Drying with the IR-CD method significantly decreased the n-6/n-3 ratio and improved the HH ratio, both of which are beneficial compared to raw insect oil. In turn, the use of PEF before drying significantly reduced the values of the HH ratio and significantly increased the n-6/n-3 ratio compared to oil from untreated and dried insects. Based on previous findings and based on the literature, this can be the effect of the free radicals formed after PEF treatment, which aids in the oxidation of unsaturated fatty acids [36]. The results showed that PEF had an adverse effect on the nutritional value of the oils tested.

The results achieved in our study showed a lower nutritional value than vegetable oils, e.g., for coriander seed oil (AI: 0.04, TI: 0.09, HH: 28.34) and parsley seed oil (AI: 0.06, TI: 0.18, HH: 16.19) [43], or for strawberry seed (AI: 0.06, TI: 0.06, HH: 18.81) and cranberry seed (AI: 0.07, TI: 0.06, HH: 15.50) oils [44], or a better nutritional value, especially due to a higher HH ratio than provided, e.g., for chicken fat (AI: 0.37–0.39, TI: 0.76–0.78, HH: 2.66–2.79) [45], for lamb fat (AI: 0.49–0.52, TI: 1.10–1.15, HH: 1.92) [46], and for herring (AI: 0.70, TI: 0.26), carp (AI: 0.46, TI: 0.31) or bream (AI: 0.37–0.42, TI: 0.23–0.24) oils [47].

The AI and TI are associated with the potential ability of fat to induce platelet aggregation. The greater the value of AI and TI, the less antiatherogenic the fatty acids, and hence the lower the ability to prevent coronary heart disease. The HH ratio considers the specific impact of fatty acids on cholesterol metabolism. Considering the above-mentioned, fats with a low value of AI (<1.0) and TI (<0.5) and with a high value of HH ratio are nutritionally recommended [48]. The drying process improved the nutritional properties of insect oil compared to those of raw insect oil. In turn, PEF treatment disadvantages those properties by increasing the n-6/n-3 ratio, along with the values of AI and TI, and decreasing the HH ratio. For this reason, the PEF application does not seem to be recommended.

2.5. Fatty Acid Distribution

The positional distribution of fatty acids in triacylglycerols (TAGs) affected the physical, thermal, and nutritional properties of fat. The fatty acids are selectively esterified in the glycerol backbone at three stereospecific number positions (sn), where sn-1,3 are external positions, and sn-2 is the inner (central) position of the TAG molecules [44,49].

The fatty acid composition of the external and inner TAG positions of the tested insect oils is presented in Table 3, while Figure 2 shows the percentage share of the main fatty acids in the sn-2 position of TAG. The results showed that unsaturated fatty acids were in both external and inner TAG positions; nonetheless, their content in the inner position was higher (Table 3). This is generally a characteristic feature of vegetable oils [19,34]. In turn, the main saturated fatty acids detected, especially palmitic acid, were located primarily in the sn-1,3 TAG positions (Table 3). The percentage share of oleic acid and linoleic acid in the sn-2 TAG position was higher than 33.3%, indicating that these fatty acids were predominantly esterified in that position. What is more, the percentage share of oleic acid in the sn-2 TAG position was higher than linoleic acid, regardless of the processing of materials (Figure 2).

As a result of drying, the proportion of myristic acid and palmitic acid in the sn-2 position increased compared to the proportion of these acids in oil from the raw material (Figure 2). The opposite trend was observed for stearic, oleic, and linoleic acids. Moreover, noteworthy changes in the positional distribution of fatty acids were observed following the application of PEF. It was noted that saturated fatty acids were directed towards the sn-1,3 position, while unsaturated fatty acids were moved towards the sn-2 position. Previous studies on oil extracted from convective-dried yellow mealworm larvae have shown that unsaturated fatty acids prefer the inner (sn-2) TAG position. In contrast, saturated fatty acids are mainly in the outer TAG positions [8]. This is less beneficial in terms of nutritional aspects. Free saturated fatty acids released from the external TAG positions are less efficiently absorbed and combine with free Ca^2+^ ions to generate insoluble calcium salts, which are subsequently eliminated from the body via stools [19]. To summarize, compared to the fatty acid distribution in TAG molecules of oil from raw insects, the IR-CD drying process was beneficial. That process decreased the share of unsaturated fatty acids and increased the share of saturated fatty acids in the sn-2 TAG position. In turn, PEF treatment impairs the nutritional properties by increasing the share of unsaturated fatty acids in the sn-2 TAG position, thereby negatively influencing oil metabolism. For this reason, the PEF application does not seem to be recommended.

2.6. Oxidative Stability

One of the most important markers of the quality of oils or fats and their durability is oxidative stability, which determines their resistance to oxidation processes [50]. The oxidative stability depends directly on the composition of the fatty acids, especially the PUFA group, but is also affected by the content of antioxidant compounds, as well as primary and secondary oxidation products [34,43].

The primary principle for oxidative stability is that the longer the oxidation induction time (OIT), the greater the resistance of fat to the oxidation processes. The OIT was about 7.9 min for the oil from raw insects, and between 11.2 and 11.5 min for oils from dried insects (Table 4). The results correspond with the peroxide values, which were the highest for oil from raw larvae. Previous studies on oil extracted from convective-dried yellow mealworm larvae have shown that only after PEF at 5 kJ/kg, OIT was longer (10.13 min) than for the oil from the untreated sample (8.50 min) [8]. There are also studies in which oxidative stability was measured by the Rancimat method. The presented OIT was equal to 36.0 h for yellow mealworm oil from larvae dried at 60 °C [13] and in the range of 10.6 to 64.6 h for oils from freeze-dried and roasted yellow mealworm [37].

The utilization of PEF and the drying process significantly increased the oxidative stability. It was noted, based on Pearson correlation, that there was a very strong correlation (r ≈ 0.945) between PEF energy input and oxidation stability. This is likely due to changes in the proportions of individual fatty acid groups (Table 2) and alterations in their distribution within TAG molecules (Figure 2). Furthermore, the presence of free fatty acids or primary and secondary oxidation products increases the fat’s susceptibility to oxidation [51,52]. The results obtained in our study do not confirm this theory. It could be that the oils may have a greater content of bioactive compounds with antioxidant properties, which was also observed for oil from rapeseed treated by PEF [53]. What is more, a greater oxidative stability may be observed due to unsaturated fatty acids positioned mainly at the internal (sn-2) position in TAG molecules, since several studies show that TAG containing unsaturated fatty acids located in the sn-2 position indicate greater oxidative stability [54,55]. PEF treatment increased the share of unsaturated fatty acids in the sn-2 TAG position, thereby improving the oxidative stability of the tested oils. For this reason, PEF application seems to be recommended before IR-CD drying.

2.7. Antioxidant Properties

The antioxidant properties, especially the presence of antioxidant compounds, affect the oxidation process occurring and the oxidative stability of fats [56]. Yellow mealworm oil, obtained from freeze-dried material, was characterized by a high tocopherol content (196 mg/kg of oil) in a study by Purschke et al. [57], while oil extracted from convective-dried insects presented a high tocopherol content (144.3 mg/kg of oil) and polyphenol content (18 mg GAE/kg of oil) in a study conducted by Son et al. [13]. The antioxidant activity of the insect oil extracts, as determined using the DPPH assay method, is presented in Table 4. Although polyphenols were extracted into methanol prior to analysis, the DPPH assay remains solvent-dependent [58] and does not fully represent antioxidant activity in the native lipid matrix; therefore, the results should be interpreted with caution.

Oil extracted from raw larvae exhibited significantly lower antioxidant activity compared to oils extracted from dried samples. Significant differences in DPPH antioxidant activity were observed depending on the PEF energy. A decreasing trend with increasing PEF energy intake was observed, with a strong Pearson correlation coefficient (r ≈ −0.908). The reduction in antioxidant activity may be related to the formation of free radicals because of PEF treatment [59]. This results in greater use of antioxidant compounds to scavenge/neutralize them [20,60] and to reduce unfavorable reactions, such as oxidation, resulting in lower antioxidant activity in the tested insect oils. Furthermore, a strong Pearson correlation (r ≈ 0.793) was observed between the oxidative stability of insect oils and their antioxidant activity, suggesting that some antioxidant compounds may have been used to mitigate oxidative changes in the fat during the material’s drying process. To conclude, despite the lower antioxidant activity of the tested oils from PEF-treated insects, PEF treatment may result in structural changes that improve the availability of antioxidant compounds used to protect the fat fraction against oxidation during drying. For this reason, the PEF application seems to be recommended before IR-CD drying.

3. Materials and Methods

3.1. Material

Raw (alive) yellow mealworm larvae were purchased from a local Polish producer (Cirwins, Kamień Duży, Poland), sourced from the same batch and under the same environmental conditions (temperature and humidity). After purchasing, larvae were kept at 4 ± 1 °C for 24 h to discard their bowel content. Before the experiments, insects were sieved through a metal kitchen sieve to remove impurities and feces, and then washed with tap water. The tested material exhibited a water content of 2.64 ± 0.02 g H_2_O/g d.m.

3.2. Technological Part

3.2.1. Pulsed Electric Field (PEF)

The use of PEF was considered the first step in the killing procedure of insect larvae.

The PEF pretreatment with PEF was carried out using a PEF Pilot™ Dual batch system (Elea Technology GmbH, Quakenbrück, Germany) in duplicate. Approximately 350 g of yellow mealworm larvae were loaded into the treatment vessel and submerged in tap water (22 ± 1 °C, electrical conductivity 220 μS/cm) to obtain a total batch mass of approximately 1 kg. The prepared batch was then placed inside the pulse chamber of the reactor between two stainless-steel electrodes separated by a distance of 280 mm. The parameters during PEF treatment were as follows: electrode voltage of 24 kV, pulse frequency of 20 Hz, pulse duration of 40 µs, and electric field strength of 1.07 kV/cm. The specific energy intake W_spec_ (kJ/kg) was supplied by applying a specific number of pulses depending on the specific energy intake (5, 20, and 40 kJ/kg), and it was calculated as presented below [8]:

$$[eqn]$$

where C is the capacitance (μF), U is the voltage (kV), n is the number of pulses (–), and m is the mass of the treated samples (kg).

3.2.2. Infrared-Convective Drying (IR-CD)

The drying was performed in a prototype laboratory dryer (Warsaw, Poland) equipped with nine lamps, positioned 0.25 m from the surface of the dried material and providing a radiation intensity of 7.8 kW/m^2^. Heated air at 40 °C was supplied parallel to the insect layer at a velocity of 0.8 m/s. The process was conducted until a constant mass was reached, determined based on at least 4 repeatable values obtained by means of indications of a balance (±0.1 g) connected to the dryer tray. The final water content of the dried untreated material was about 0.06 g H_2_O/g d.m., and that of dried PEF treated material was about 0.03 g H_2_O/g d.m. The process was done in duplicate.

3.3. Analytical Part

The dried insects, before starting the analytical part, were packaged in PET/AL/PE bags (Pakmar, Warsaw, Poland) to prevent access to external vapor and light and were kept for a week to stabilize them.

3.3.1. Chemicals

The reagents and solvents used in this study were purchased from POCH S.A. (Gliwice, Poland). All chemicals used for extraction, analysis, and sample preparation for GC were of chromatographic or analytical purity. The fatty acid methyl ester (FAME) standard (Supelco 37 Component FAME Mix) was provided by Sigma-Aldrich GmbH (Schnelldorf, Germany).

3.3.2. Oil Yield Determination

The oil was extracted in triplicate via the Soxhlet method using a Behrotest ET2 Control Unit (Behr Labor-Technik GmbH, Düsseldorf, Germany) with petroleum ether as the solvent. The extraction process lasted 6 h [61]. The oil extraction yield was calculated as the oil’s weight to the powdered insects’ weight and expressed per 100 g of dry matter.

3.3.3. Oil Extraction for Analysis

The oil from raw and dried insects was extracted in duplicate using the Folch procedure, as described by Kozłowska et al. [62]. In brief, 50 g of previously ground, dried material was placed into a glass bottle sealed with a screw cap. Subsequently, 100 mL of a chloroform:methanol solution (1:1, v/v) was added, and the mixture was stirred for 3 min. The obtained mixture was maintained at 60 °C for 20 min, and after cooling, an additional 100 mL of chloroform was added, then stirred intensively for 3 min and filtered through the filter paper. In the next step of the procedure, 70 mL of a 0.1 mol/L potassium chloride solution (KCl) was added to the filtrate. The obtained solution was thoroughly mixed and left overnight at 4 °C to achieve phase separation. The lower phase (chloroform fraction) was collected, dried with anhydrous magnesium sulfate for 40 min in the dark, and filtered through a filter paper. The chloroform was evaporated from the filtrate using a Büchi R-300 Rotavapor rotary evaporator (Büchi Labortechnik AG, Flawil, Switzerland) at 40 °C.

3.3.4. Acid Value and Peroxide Value

The acid value (AV) and the peroxide value (PV) of the oil samples were determined according to the method described by Brzezińska et al. [34]. The AV was assessed by titration with an ethanolic potassium hydroxide solution (0.1 mol/L) and expressed as the amount of KOH (mg) required to neutralize free fatty acids present in 1 g of insect oil. The PV was evaluated using a titration procedure with sodium thiosulfate solution (0.001 mol/L) as the titrant, and the results were reported as milliequivalents of active oxygen per kilogram of oil (meq O_2_/kg). Both indices were measured by using a titrator TitraLab AT100 (HACH LANGE, Wrocław, Poland) in triplicate.

3.3.5. Fatty Acid Composition

Fatty acid profiles were determined using the analytical approach reported by Bryś et al. [63]. Derivatization of fatty acids into methyl esters (FAMEs) was prepared by the oil sample esterification with methanol according to the ISO standard [64]. Gas chromatographic separation was carried out using a YL6100 gas chromatograph (Young Lin Bldg., Anyang, Hogye-dong, Republic of Korea) equipped with a flame ionization detector (FID) and a BPX-70 fused-silica capillary column (SGE Analytical Science, Milton Keynes, UK; 60 m × 0.25 mm i.d., 0.25 μm film thickness). The oven temperature for the separation of FAMEs was programmed as follows: 70 °C for 5 min, 15 °C/min to 160 °C, 1.1 °C/min to 200 °C, 200 °C for 12 min, 30 °C/min to 225 °C, and 225 °C for 1 min. The injector temperature was set at 225 °C, and that of the detector was set at 250 °C. The carrier gas used was nitrogen, with a flow rate of 1 mL/min and a split ratio of 1:50. The results were expressed as relative percentages of each fatty acid (% of the fatty acid peak area was calculated). The relative retention times of the FAME peaks were compared with those of the FAME chemical standard for the identified fatty acids.

3.3.6. Nutritional Value Indices

The atherogenicity index (AI), the thrombogenicity index (TI), and the hypocholesterolemic/hypercholesterolemic (HH) ratio were calculated [8,48]:

$$[eqn]$$ $$[eqn]$$ $$[eqn]$$

3.3.7. Fatty Acid Distribution

The positional distribution of fatty acids at the sn-2 and sn-1,3 positions of triacylglycerols (TAGs) was determined according to the method presented by Bryś et al. [63]. Briefly, 0.1 g of oil was mixed with regiospecific pancreatic lipase (Lipozyme RM IM, Type II, Merck KGaA, Darmstadt, Germany), Tris buffer, bile salts, and calcium chloride. After 5 min of incubation at 40 °C, the reaction was stopped by HCl, and the products were extracted with diethyl ether. The samples were then separated and loaded onto a thin-layer chromatography (TLC) plate. The 2-monoacylglycerols (2-MAGs) were then extracted using diethyl ether, and their fatty acid composition was identified using the previously described GC method. The percentage of given fatty acid in the starting TAG molecules and in the sn-2 position of TAG was used to calculate the percentage share of fatty acid in the sn-2 TAG position of insect oils.

3.3.8. Oxidative Stability

A pressure differential scanning calorimeter (DSC Q20, TA Instruments, Newcastle, WA, USA) coupled with a high-pressure cell (Q20P) was applied to determine the oxidative stability of insect oils in accordance with the procedure described by Bryś et al. [63]. Oil samples weighing 3–4 mg were placed in an aluminum pan and transferred into the calorimeter pressure sample chamber. Measurement was carried out at a constant temperature of 140 °C under 1400 kPa pressure of oxygen in triplicate. The oxidation induction time (OIT) was determined based on the maximum rate of oxidation.

3.3.9. Antioxidant Properties

Methanolic extracts of the isolated oils were prepared as follows. Insect oil samples (0.25 g) were combined with 5 mL of n-hexane and transferred to the separatory funnel. Then, 10 mL of methanol:water mixture (80:10 (v/v)) was added, and the system was agitated for 3 min. After phase separation, the lower methanol–aqueous layer was collected into a round-bottomed flask. The extraction procedure was repeated in triplicate for each sample. The collected mixed extracts were evaporated using a Büchi R-300 Rotavapor rotary evaporator (Büchi Labortechnik AG, Flawil, Switzerland) at 40 °C. The extracts were reconstituted with 1 mL of methanol.

The antioxidant activity of methanolic oil extracts was measured using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical, following the procedure described by Ugur et al. [5] with minor modifications. Briefly, a 0.6 mM methanolic DPPH solution was prepared and diluted to a concentration at which the absorbance measured at a wavelength of 515 nm was equal to 0.700. Test tubes were filled with 100 mL of diluted polyphenolic fractions extracted from oils and 4 mL of DPPH solution. The mixture was incubated for 30 min in the dark, after which absorbance was recorded at 515 nm. Results were determined based on the Trolox calibration curve and given as Trolox equivalent per 100 g of oil (µmol TE/100 g). All spectrophotometric measurements were carried out using a UV-VIS Jenway 6305 (Cole-Parmer, Vernon Hills, IL, USA) spectrophotometer.

3.4. Statistical Analysis

The statistical analysis was carried out using Statistica 13.3 (TIBCO Software, Palo Alto, CA, USA). A one-way analysis of variance (ANOVA) and Tukey’s HSD multiple comparison test were applied. Pearson correlation was used to determine the relationship between the input energy of PEF and the chosen fat quality indicators. The tests were applied at a significance level of α = 0.05.

4. Conclusions

This research has demonstrated how the application of PEF-assisted infrared-convective (IR-CD) drying of yellow mealworm larvae affects the properties of the isolated fat fraction.

The analyzed insect oils were characterized by a high content of unsaturated fatty acids (about 73.8% for raw insect oil and 78.0–79.4% for dried insect oils), with a higher MUFA content (39.9–42.5%) than PUFA content (34.0–38.2%). The dominant fatty acids were oleic acid, linoleic acid, and palmitic acid. The tested oils were also characterized by a higher n-6 PUFA-to-n-3 PUFA ratio than recommended. Unsaturated fatty acids (oleic and linoleic acids) were located mainly in the sn-2 TAG position, making them similar to those of vegetable oils.

The IR-CD drying process increased the acid value, decreased the peroxide value, increased oxidation stability and antioxidant activity, and improved nutritional indices of insect oil. Despite the higher acid value, IR-CD drying appears to be beneficial for processing insects before fat extraction compared to raw insects.

The PEF-assisted IR-CD drying improved the oil extraction yield, decreased acid and peroxide values, and increased the oxidative stability of insect oils. Given that, this is beneficial from an industrial standpoint. However, PEF treatment decreased antioxidant activity and the PUFA content, deteriorated nutritional indices, and increased the share of unsaturated fatty acids in the sn-2 TAG position. For this reason, the PEF application does not seem to be recommended.

Future studies should focus on optimizing PEF parameters to balance its influence on the drying process, as well as the effects on nutritional and qualitative properties of insect oils. The integration of PEF technology and infrared-convective drying presents a promising pathway for developing novel lipid sources with tailored functional properties, potentially contributing to sustainable and nutritionally valuable food systems.