CRICKET FLOUR-LADEN MILLET FLOUR BLENDS’ PHYSICAL AND CHEMICAL COMPOSITION AND ADAPTATION IN DRIED PASTA PRODUCTS

Increasing the protein and antioxidant content of food products is a constant challenge amongst researchers. Dried pasta products are popular amongst all groups of society. The most important factor in pasta processing is the quality of the ﬂ our. Millet ( Panicum miliaceum ) ﬂ our has high nutritional value, enriching it with cricket ( Gryllus bimaculatus ) ﬂ our is good choice to increase the quality of protein composition and antioxidant properties of products. Flour mixtures of millet and insect ﬂ ours (5% and 10%) were analysed after mixing and pasta processing. Addition of wheat gluten improved both texture and nutrition value of pasta products. Total polyphenol content, antioxidant capacity, total protein content, free and total amino acid composition were studied. Quality analysis of dried pasta products were carried out according to Hungarian standards. Data was analysed with Kruskal-Wallis test, Dunn’s pair-wise post hoc test was used with Bonferroni correction. The correlation was determined by Spearman’s rank. Addition of cricket ﬂ our modiﬁ ed the pH, acid value, moisture content, and colour of the samples, these changes lasted during storage. Enrichment could increase the total phenol content signiﬁ cantly even at the low level of 10%. Heat treatment during pasta processing had negative eﬀ ect on the antioxidant capacity except at higher cricket ﬂ our contents. Cricket ﬂ our’s high protein content proportionately increased millet ﬂ our’s, thus pasta products’. Dried pasta products passed all quality norms. Enrichment of millet ﬂ our with cricket ﬂ our is favourable from both nutritional and quality aspects.

Nutrition is a key factor in food research. The pursuit for new ways to integrate diff erent macro-and micronutrients in the diet of consumers is constant. However, it seems that the majority of human population still consume food one-sidedly. Both social, cultural, and economic factors contribute to the development and change of dietary patterns. Availability and knowledge are key factors in the spread of a food product (R Z , 1985). The rising income level and population growth results an increasing demand for food, especially for protein. The consumption of protein rich food products is necessary to maintain human metabolism. However, the need is not only confi ned to the quantity of protein in human diet, the quality is also crucial. Choosing higher quality protein sources could fi ll the void in essential amino acid consumption, thus preventing malnutrition of an individual (P , 1978). In Hungary, dried pasta products are popular amongst all groups of society. Its position is also safe in the global market as well. The amount of dried pasta products consumed in recent years is elevating. This trend results that the food industry continuously tries to lower production costs, improve profi t and nutritional value of products (K P , 2014).
The most important factor in pasta processing is the quality of the fl our used. Choosing good quality base fl our can radically improve the nutritional value of the product.
Millet (Panicum miliaceum) fl our is a good replacement or addition for wheat fl our. All types of millets were found to have high nutritive value, comparable to the major cereals such as wheat and rice (P S , 1994). T and S (2017) had shown that foxtail millet fl our exhibits higher functional characteristics than refi ned wheat fl our. Furthermore, proso millet varieties have high content of slowly digestible starches, thus are good sources of dietary fi bres, which can help digestion (T et al., 2017). It has also been reported that millet proteins are good sources of essential amino acids except lysine and threonine, but are relatively high in methionine. Millets are also rich sources of phytochemicals and micronutrients (M et al., 2010; S R , 2012). The best way for improving the millet fl our's amino acid composition is enrichment. In case of the millet fl ours -as mentioned above -there is a hiatus of the amino acids: lysine and threonine. To supplement these amino acids, one way is addition of an alternative protein source. To address the hiatus of lysine content of millet fl our and to improve the texture of the pasta products, the best way is the addition of wheat fl our.
The use of insect fl ours is more and more accepted in the western culture, and the shift from animal protein is necessary not just for the health of our planet but for ours as well (G , 2010; T et al., 2011; W , 2014; M , 2018). The production of insect fl our is fast, cost eff ective, and environmentally friendly. The preparation is highly supervised and controlled by the Food and Drug Administration (FDA, 1998), as well as the European Union (EC, 1997). Cricket (Gryllus bimaculatus) fl our is a good choice to add to millet fl our. With its high protein and low carbohydrate content it can positively alter the nutritional value of the product without having a large environmental footprint.
Addition of insect fl our could change the sensory properties of a given food product. Millet fl our has a relatively neutral fl avour and smell. Depending on quantity, rationing of cricket fl our could result aftertaste and unappetizing smell of food products. Studies suggest that consumer acceptance of edible insect-laden products can be enhanced by processing and blending with familiar food products (P et al., 2018). Using a prevalent product, like dried pasta, is a great start in introducing insect fl ours into western diet.
The aim of this study was to analyse the chemical behaviour of millet fl our enriched with cricket fl our in diff erent quantities. Also the chemical and quality properties of the dried pasta products made from these fl our mixtures were investigated. Another goal was to evaluate the possibilities of cricket fl our-laden pasta in industrial processing and on commercial market.

Materials
The fl ours were bought commercially. The bio millet fl our (Panicum miliaceum) and wheat gluten were produced by GreenMark Organik, the Gryllus bimaculatus cricket fl our was purchased from ThailandUnique. The reagents for the chemical measurements were provided by Reanal llc.

Samples
All measurements were carried out on the pure fl ours as well as the fl our mixtures. For the enriched fl ours, the millet fl our and the cricket fl our in two diff erent compositions (5% and 10% of cricket fl our by weight of the whole mix) were mixed.
The pastas were made from both the pure millet fl our and the fl our mixtures. To give structure and further improve protein composition of the pastas, wheat gluten (20% of the weight of dry matter) was added. After mixing the dry matter, the pastas were formed with the addition of water (50% of the weight of dry matter). The quantities of these additional ingredients were kept the same in every recipe.
After the right amounts of both fl ours were weighing, wheat gluten was added. After homogenizing the powder mixture in a mortar, it was stirred in the water. The pastas were kneaded by hand, rolled and cut by Mercato Atlas Deluxe 150 type pasta machine into strips. The drying process was executed in an Armfi eld type fl uid dryer for 20 minutes at 80 °C. From pure cricket fl our no pasta could be made with these test conditions.
The samples were subjected to chemical measurements. In preparation, 100 mg ml -1 extracts were made with distilled water. The pasta products were grated in a mortar, the extracts were made from the powder. Every analytical method was carried out on three parallel samples.

Chemical analysis.
Both the fl our mixtures and the dried pasta products were evaluated with chemical methods.
Moisture content was determined with Sartorius MA 50 type fast moisture analyser. Water activity was measured with Novasina Lab Master. For the colour measurement the Minolta Chromameter CR-310 device with tristimulus objective in Lab coordinate system was used on powdered samples. For pH measurement, a Mettler Toledo SevenGo Duo pH/ conductivity meter SG23 hand device was used. The determination of pH and acid value was carried out based on Hungarian standards (1985a).
Total water-soluble polyphenol content was measured by Folin-Ciocalteu method (S R , 1965) and expressed in gallic acid equivalent (GA). Water soluble antioxidant capacity was measured using FRAP (ferric reducing antioxidant power) assay (B S , 1996) and given in ascorbic acid equivalent (AA). For measuring the water-soluble protein content, Layne method was used (L , 1957). For the spectral analysis, Rayleigh UV1800 type spectrophotometer was applied. Total and free amino acid contents of the samples were measured with an Ignos AAA 400 Automatic Amino acid Analyser, sample preparation went according to the manufacturer's instructions. The amino acid profi les were determined with Chromulan V 0.82 software and given in μg g -1 unit.

Quality analysis of dried pasta products.
To determine the adaptation of the pastas in commercial trade, Hungarian Standards (1985b) were used. Measurement of optimum cooking time, swelling capability, disintegrating and sticking during cooking, and sensory evaluation were carried out.

Statistical evaluation.
All measurements were carried out in fi ve replicates, using XL-stat software. Because of the parallel samples, non-parametric probes were used. The Kruskal-Wallis test was applied to calculate the p-value (α=0.05), and Dunn's pair-wise post hoc test was used with Bonferroni correction on signifi cant results. The correlation between parameters was determined by Spearman's rank correlation (non-parametric equivalent of Pearson's correlation) (α=0.05).

Results and discussion
2.1. Physical and chemical properties 2.1.1. Moisture content and water activity. The moisture content and water activity of all samples were below the limit given by Hungarian Standards. Addition of the cricket fl our lowered both values in fl our mixtures and pastas as well. In the case of mixtures, addition of cricket fl our at 5% and 10% lowered moisture content by 2.58% and 5.79%, respectively.
Water activity of the mixtures increased at 5% supplementation by 1.88% and at 10% supplementation by 4.93%. These values are relatively high given the added content. In the case of both types of enriched pastas, the increase was 10%.

pH and acid value.
The cricket fl our's pH was higher than the millet fl our's (6.21±0.10 and 6.01±0.02, respectively). The pH of the mixtures followed the same trend as the moisture content and the water activity. The dried pasta's pH was lower than the relating fl our's (pure millet: 5.97±0.07; 5%: 6.00±0.01; 10%: 6.01±0.01).
The cricket fl our had relatively high acid value at the start (23.78±0.042), which enhanced the value of the mixtures (from 5.02±0.021; 5% mix: 7.24±0.04; 10% mix: 15.78±0.01). The acid value of pasta products was lower than the base fl our's. The millet fl our pasta had 4.59±0.04, the 5% pasta: 2.02±0.02 and the 10%: 4.95±0.04 acid value. This means that the method of pasta processing protects from lipid oxidation that could occur in the fl our samples.

Colour measurement.
The addition of cricket fl our changed the colour of fl our mixtures and dried pastas visibly. For evaluating the obtained colour data, statistical analysis on fl ours and pastas was used together.
The lightness (L*) value decreased with increasing the added amount of cricket fl our, because the cricket fl our has darker colour (L*=43.11±0.21) than millet fl our (L*= 90.16±0.13). The a* value gives the greenred axis, the b* the blueyellow axis of the colour chart. The increasing ratio of cricket fl our resulted redness (rising value of a* value) and blueness (lowering in b* value) in the mixtures.

Antioxidant properties
2.2.1. Water-soluble total polyphenol content. The water-soluble (WS) total polyphenol content of the fl ours and pastas are given in mg gallic acid/g dry matter (Fig. 1). The lower amount in the 10% fl our mixture can be caused by inhomogeneity of the sample. The pairwise comparison gave two homogeneous groups and at least two samples were signifi cantly diff erent amongst all samples. The least amounts of polyphenolic compounds were measured in 100% millet fl our and 100% millet fl our-based pasta. The highest amount group contained the 100% cricket fl our and the 10% cricket fl our-based pasta. According to the data, the addition of cricket fl our could increase the phenol content signifi cantly even at the low level of 10%. This means a two-fold elevation from 0.125±0.008 mg GA/g DS (100% millet fl ourbased pasta) to 0.252±0.043 mg GA/g DS (10% cricket fl our-based pasta).
Acta Alimentaria 49, 2020 Fig. 1. Water-soluble total polyphenol content of the fl our and pasta samples given in mg gallic acid/ g dry matter.
White, squared, striped, and grey columns represent 100% millet fl our and pasta, 5% cricket fl our mixture and pasta, 10% cricket fl our mixture and pasta, and 100% cricket fl our, respectively. Stars above the columns mark signifi cantly diff erent values (P<0.05). Figure 2. The Dunn-type post hoc test determined three groups amongst the samples. Two of those were homogeneous groups. The lowest antioxidant capacity was for 100% millet fl our and 100% millet fl our-based pasta, the highest value belonged to the 10% cricket fl our-based pasta. The pairwise comparison found that there was signifi cant diff erence between the dried pasta samples made with 5% and 10% cricket fl our.

Water-soluble total antioxidant capacity. WS antioxidant capacity can be seen in
According to these results, we are able to say that addition of cricket fl our to the base mixture signifi cantly increases the antioxidant capacity both of fl our mixtures and pasta products. Also, it can be stated that heat treatment has negative eff ect on the antioxidant capacity except in case of higher cricket fl our content, where the value signifi cantly rose after pasta processing.

Protein content
2.3.1. Water-soluble total protein content. From the measured values of WS protein, the statistical analysis defi ned three groups, two of those were homogeneous. There were at least two signifi cantly diff erent samples as well. The lowest content was measured in the 100% millet fl our (32.98±2.52 mg g -1 ) and the highest in the 100% cricket fl our (138.11±5.41 mg g -1 ) and the 10% cricket fl our-based pasta (84.32±4.46 mg g -1 ). The addition of cricket fl our elevated the protein content of the mixtures. However, this trend was not clear in the pasta samples. Fig. 2. Water-soluble total antioxidant capacity of the fl our and pasta samples given in mg ascorbic acid/g dry matter. White, squared, striped, and grey columns represent 100% millet fl our and pasta, 5% cricket fl our mixture and pasta, 10% cricket fl our mixture and pasta, and 100% cricket fl our, respectively. Stars above the columns mark signifi cantly diff erent values (P<0.05).
There was signifi cant diff erence between the protein content of the millet fl our and the cricket fl our and between the 10% cricket fl our-based pasta and the 5% cricket fl our-based pasta.
In the case of 100% millet fl our and 10% cricket fl our mixture, the processing of pasta products resulted in signifi cant elevation of protein content. In the pasta samples made from the 5% cricket fl our mixture the protein content has not changed signifi cantly, this low amount of added cricket fl our was not enough to relevantly change the nutritional value of the fi nal product.

Total and free amino acid content.
The reason of measuring both the total and free amino acid compositions was the fact that insect fl ours contain essential amino acids for humans, thus they not only increase the quantity of protein content but the quality of protein composition as well (C -V et al., 2017). The evaluation of free amino acid composition was carried out by Chromulan V 0.82 software provided by the manufacturer, and the values were given in mg g -1 unit. The total amino acid composition was given in g/100 g unit.
The lowest amounts of total amino acid content were in the 100% millet fl our and 100% millet fl our-based pasta. The cricket fl our had twice as much total amino acid value than the millet fl our (59.2±0.7 g/100 g and 25.6±1.6 g/100 g, respectively) and this deviation was signifi cantly diff erent. The addition of cricket fl our increased the protein content proportionately.
Acta Alimentaria 49, 2020 As mentioned above, millet fl our is defi cient in the amino acids threonine and lysine. The contents of these amino acids were relatively low in the 100% millet fl our, 3.116 mg g -1 and 1.802 mg g -1 , respectively. In contrast, the amounts of these amino acids were high in the 100% cricket fl our (21.1 mg g -1 and 13.6 mg g -1 , respectively). The diff erence caused a favourable increase of the amounts of these amino acids in the mixtures and the fl our mixturebased pastas. The total amino acid content was always lower in the fl our mixtures, which means that pasta processing could improve protein composition.

Quality of dried pasta products
The quality properties were analysed according to Hungarian Standards. Cooking times of all three types of pastas were between 3.5-4 minutes. The highest swelling value belonged to the 10% cricket fl our based dried pasta, which means that the addition of cricket fl our improved the swelling capability of the millet fl our. The disintegrating and sticking values of pasta were below the allowed limit.
Sensory analysis of alternative pasta products measures taste, smell, outer appearance, consistency, and cooking properties. According to these values, there are three quality classes for pasta products. The 100% millet fl our-based pasta fell into the I. class of quality groups. The cricket fl our content lowered the sensory properties of pastas resulting in quality descent to II. class.

Statistical evaluation
On the signifi cant diff erences between our data, Pearson's type correlation studies were carried out. The correlation between four factors: WS total polyphenol content, WS total antioxidant capacity, total amino acid content, and WS total protein content were examined.
Based on the obtained "r" and "p" values (Tables 1 and 2) we found that all parameters correlate positively in some degree. Strongly positive correlation was found between WS total polyphenol content and WS total antioxidant capacity. This relationship was determined by K and co-workers (2010) as well, while studying antioxidant properties of diff erent types of millets.
Two other moderately strong correlations were determined between the groups. One was found between WS total polyphenol content and total amino acid content. The cause of this correlation could lay in polyphenolicprotein interactions in the matrix of samples. For evaluating molecular changes happening in the food matrix, further measurements are necessary. The second was found between WS total antioxidant capacity and amino acid content. The concept that proteins can act as antioxidants are discussed in recent years based on their molecular structures, hydrolysates, and amino acid sequences (A , 2015)

Conclusions
Addition of cricket fl our could improve physical, chemical, and storage properties of millet fl our-based fl our mixtures and dried pasta products. All measured parameters passed Hungarian Standards. Antioxidant capacity, total polyphenol content, and protein content improved with enrichment. Cricket fl our compensated for the hiatus in millet fl our's amino acid composition, increasing the amounts of threonine and lysine. The processing method of pasta products improved antioxidant capacity and protein content. Enrichment of millet fl our with cricket fl our is favourable from both nutritional and quality aspects. Although millet fl our is a popular gluten free option, the focus was rather on the nutritional improvement than the gluten free status of our products. In the future, development of insect fl our enriched pasta products as well as designing gluten free options could be one of the prospects of the food industry. *