*Proceedings***, 2020, la ValSe-Food 2019**

Editors

**Isabel Castanheira Norma C. Samm´an Loreto Mu ˜noz H. Claudia Monika Haros**

MDPI • Basel • Beijing • Wuhan • Barcelona • Belgrade • Manchester • Tokyo • Cluj • Tianjin

*Editors* Isabel Castanheira National Institute of Health Doutor Ricardo Jorge Portugal

Norma C. Samman´ Universidad Nacional de Jujuy Argentina

Loreto Munoz H. ˜ Universidad Central de Chile Chile

Claudia Monika Haros Instituto de Agroqu´ımica y Tecnolog´ıa de Alimentos (IATA-CSIC) Spain

*Editorial Office* MDPI St. Alban-Anlage 66 4052 Basel, Switzerland

This is a reprint of articles from the Special Issue published online in the open access journal *Proceedings* (ISSN 2504-3900) (available at: https://www.mdpi.com/2504-3900/53/1).

For citation purposes, cite each article independently as indicated on the article page online and as indicated below:

LastName, A.A.; LastName, B.B.; LastName, C.C. Article Title. *Journal Name* **Year**, *Volume Number*, Page Range.

**ISBN 978-3-0365-0368-4 (Hbk) ISBN 978-3-0365-0369-1 (PDF)**

Cover image courtesy of Loreto Munoz. ˜

© 2020 by the authors. Articles in this book are Open Access and distributed under the Creative Commons Attribution (CC BY) license, which allows users to download, copy and build upon published articles, as long as the author and publisher are properly credited, which ensures maximum dissemination and a wider impact of our publications.

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### **Contents**



Reprinted from: *Proceedings* **2020**, *53*, 21, doi:10.3390/proceedings2020053021 ........... **129**

#### **Mar´ıa Gabriela Bord ´on, Gabriela Noel Barrera, Maria C. Penci, Andrea Bori, Victoria Caballero, Pablo Ribotta and Marcela Lilian Mart´ınez**

Microencapsulation of Chia Seed Oil (*Salvia hispanica* L.) in Spray and Freeze-Dried Whey Protein Concentrate/Soy Protein Isolate/Gum Arabic (WPC/SPI/GA) Matrices Reprinted from: *Proceedings* **2020**, *53*, 22, doi:10.3390/proceedings2020053022 ........... **135**

### **About the Editors**

**Isabel Castanheira**, PhD (ORCID: 0000-0001-7273-0676), Principal Researcher, Head of the Food and Nutrition Department, National Institute of Health Doutor Ricardo Jorge.

As Head of Department, she is responsible for consolidating INSA's national and international reputation for developing and applying analytical methods for assessing food quality and food safety. She coordinates activities on the content of nutrients and contaminants of emerging interest in food products in terms of the comparability and reliability of the measurement values. She coordinates the management of activities related to the food–disease relationship. As a member of the WHO Collaborating Centre for Nutrition and Childhood Obesity, she works under WHO recommendations to characterize infant foods and other commodities with relevance to nutritional epidemiology. As a chairperson of the IMEKO Technical Committee for Metrology in Food and Nutrition (2009–2020), she worked on aspects of the traceability routes to SI units involving intermediate reference points which are identified and developed through state-of-the-art analytical methods. She has been a member of international collaborative studies focusing on the certification campaign of values assigned to certified reference materials. She permanently supervises Master's and PhD theses involving: 1) analytical methods; 2) nutrient profile; 3) quality tools applied to food science and technology; 4) inorganic contaminants; 5) speciation analysis; 6) customer perception of label claims; and 7) nutrient losses and gains and yield factors. Research member of several European research projects. She dedicates part of her research activities to the nutritional characterization of traditional foods from Latin America with an impact on health. A reviewer of scientific articles in indexed journals, mainly in the first quartile (SCImago Journal Rank). Associated Editor of Food Chemistry (Elsevier). Member of the editorial boards of the Journal of Food Composition and Analysis (Elsevier), Measurement (Elsevier) and Foods (MPDI). Author of more than 100 scientific publications in various areas, from metrology to food toxicology, with a high share of publications in journals of the first quartile of importance (SCImago Journal Rank).

She has been invited to give lectures in several international food graduate courses under FAO and WHO patronage and at universities around the world. She was chair of 13th International Food Data Conference organized by INFOODS (FAO) held in Lisbon.

**Norma C. Samm´an**, Chemical Engineer, Doctor of Food Technology, Full Professor at the National Universities of Jujuy and Tucuman. She has extensive experience in the chemical, nutritional and ´ functional characterization of foods and the development of new food products. She has made important achievements in the academic, research and transfer fields.

Since 1999, she has been a member of the Regional Academic Commission of the Doctorate in Food Science and Technology of the Northwest of Argentina. She has contributed to the training of human resources through the development of national and international courses and in the direction of more than 20 doctoral theses. She was Director of the Jujuy Research and Transfer Center (CIT-Jujuy) Universidad Nacional de Jujuy (UNJu)-Consejo Nacional de Investigaciones Cient´ıficas y Tecnicas (CONICET) (2012–2018). ´

She was the President (2006–2009; 2015–2018) and is now the current Geographic Representative (2018–2021) for South America of LATINFOODS (Latin American Food Composition Network). She has contributed to the development of capacities and standards in food composition. She is a member of the INFOODS Task Force.

Currently, her research activities are oriented to the study of Andean crops in the Argentine Northwest with the main objective of promoting sustainable productive activities. The research group that she directs carries out activities related to the following: i) Survey of the productive and economic-social characteristics of agricultural producers to detect potentialities, making organizational proposals, many of them accepted by the communities involved and compatible with their worldview, values, culture and history. ii) Preservation of genetic material and characterization of the different varieties of Andean potatoes and corn. This material was distributed to agricultural cooperatives to encourage its cultivation and conservation. iii) Development of low-cost and environmentally compatible technologies for the elaboration of new food products using native raw materials that allow high retention of nutrients, biofunctional compounds and their sensory characteristics.

She has published numerous scientific articles, book chapters and technical or policy guidance documents and has received awards for her work in the field of food and nutrition.

**Loreto A. Munoz**, Food Process Engineer from the Universidad de Santiago de Chile (Chile), Master's in Food Science from Universidad de Chile (Chile), PhD in Science and Food Engineering from Universidad de Santiago de Compostela (Spain) and PhD in Engineering Science from Pontificia Universidad Catolica de Chile (Chile). At present, she is full professor at the Universidad Central de Chile and Head of the Food Science Lab (LabCial).

She has contributed to the training of advanced human capital through the direction of undergraduate and graduate theses.

She has participated and directed numerous projects of national and international scientific and academic significance, as well as significant impact, through ISI scientific publications. In 2013, she received the BIMBO Panamericano Award for her research on the chia seed.

Currently, her research has been focused on the extraction and characterization of food materials of plant origin of interest for human consumption such as seeds, grains and legumes in terms of their microstructural and morphological characterization and nutritional, physical (optical, rheological, textural and thermal properties), chemical and functional properties, as well as studying the digestibility, bioaccessibility and bioavailability of nutrients. In addition, she has experience in the extraction, separation and concentration of their components, as well as their application in food matrices with functional properties.

Finally, her current projects are related to the evaluation of new sources of dietary fiber in terms of the potential contribution on the reduction in risks associated with metabolic syndrome.

**Claudia Monika Haros**, Chemist, graduated from the School of Exact and Natural Sciences, University of Buenos Aires (UBA), Argentina, in 1990. She also gained her MSc in Bromatology and Food Technology (1992) and MSc in Biology Analysis (1997) from the UBA. She has a PhD in Chemistry (UBA—1999), officially approved by the Ministry of Education, Culture and Sport of Spain. From 1991 to 2003, she worked as university professor in the Organic Chemistry Department, Food Science and Technology Area, UBA. From 1991 to 1999, she was a Research Assistant in the Cereals and Oilseeds Group, Department of Industrial Chemistry, UBA. Later, from 2000 to 2002, she worked in Spain as a visiting professor in the Cereal Group of the Institute of Agrochemistry and Food Technology (IATA) in Valencia. During 2003, she was a postdoc fellow at the Department of Food Microbiology, Institute of Animal Reproduction and Food Research (CENEXFOOD-EU), Polish Academy of Science, Olsztyn, Poland. From 2003 to 2004, she received an award for working with Prof. Sandberg of the Department of Chemical and Biological Engineering, Life Science Division, University of Chalmers, Gothenburg, Sweden. In 2005, she became a Research Associate (Ramon y Cajal Contract) of the Spanish Council for Scientific Research of the Ministry of Economy and Competitiveness (CSIC-MINECO). Since 2008, she has been a permanent staff member of CSIC and continues her investigation in the Cereal Group, Department of Food Science of IATA. Since 2015, she has been a coordinator of the International Chia-Link Network, and since 2018, she has been the coordinator of la ValSe-Food Group-CYTED.

Since the early stages of her career, she has mainly been engaged in research in respect to the cereal science and technology field. The major theme in Dr. Haros's research is the utilization of different strategies to improve the nutritional and/or functional value of cereal by-products or cereal ingredients. These strategies include the use of different physical, biochemical or biological treatments during the cereal milling process; the development of new cereal by-products by including novel ingredients; and the use of new starter phytase producers for regulating the content and composition of lower myo-inositol phosphates in cereal by-products with clear nutritional and health benefits. In recent years, her research has focused on nutritional studies of raw vegetable materials and/or their by-products to determine their biological activity for their subsequent integration into new food matrices. For this purpose, different in vitro and in vivo strategies are utilized for determining nutritional and/or biological activities. These assays include the determination of the bioaccessibility/bioavailability of minerals, glycaemic index and nutrient inputs according to Dietary Reference Intakes/Adequate Intakes (DRIs/AIs). The ultimate objective is to identify dietetic solutions and innovations to prevent diseases and to improve consumers' well-being/health.

https://www.iata.csic.es/en/staff/claudia-monika-haros https://publons.com/researcher/1751589/monika-haros/ ID: H-6839-2012 http://orcid.org/0000-0001-7904-0109

The editors belong to the Chia-Link Network and la ValSe-Food CYTED Network http://www.chialink.es/ http://www.cyted.org/es/valse food

### **Preface to "***Proceedings***, 2020, la ValSe-Food 2019 "**

Iberoamerican crops are underutilized and cultivation levels are low, but, recently, their worldwide demand has significantly increased, resulting in a production increase as well as a price increase. For many years, these valuable seeds have been widely recognized by food scientists and food producers because of their nutritional value. They contain high-quality proteins, and some contain abundant amounts of starch and/or fiber (with unique characteristics) and large quantities of micronutrients such as minerals, vitamins and bioactive compounds; moreover, they are gluten-free, which makes them suitable for people suffering from gluten intolerances/allergies. For these reasons, the Iberoamerican valuable seeds interest has immensely increased, since in recent decades, the research efforts have been intensified.

This book summarizes the Proceedings of the II International la ValSe-Food, Development of Food Ingredients from Iberoamerican Ancestral Crops and V Symposium of Chia-Kink Network held at the National Institute of Health Doutor Ricardo Jorge (INSA), 20–21 October, Lisbon, Portugal. This book gathers the recent investigations of la ValSe-Food Group on these valuable seeds and other crops and provides comprehensive and up-to-date knowledge within all the relevant fields of food science. It provides information on production and utilization, structure and chemical composition, paying special attention to carbohydrates, fibers, bioactive compounds, proteins and lipids of kernels and other parts of the plants. It includes their processing, various food products and applications and the nutritional and health implications.

We hope that this book will contribute to the increased utilization of Iberoamerican valuable seeds in human nutrition.

This publication was financially supported by Ia ValSe-Food-CYTED (119RT0567).

#### **Isabel Castanheira, Norma C. Samm´an, Loreto Mu ˜noz H., Claudia Monika Haros** *Editors*

### *Proceedings*  **Nutritional Value and Bioactive Compounds in Andean Ancient Grains †**

#### **Ritva Repo-Carrasco-Valencia**

Centro de Investigación e Innovación en Productos Derivados de Cultivos Andinos CIINCA, Universidad Nacional Agraria La Molina, Avenida La Molina s/n Lima 18, Lima 15024, Peru; ritva@lamolina.edu.pe † Presented at the 2nd International Conference of Ia ValSe-Food Network, Lisbon, Portugal,

21–22 October 2019.

Published: 3 August 2020

**Abstract:** Quinoa (*Cheopodium quinoa*), kañiwa (*Cheopodium pallidicaule*), kiwicha (*Amaranthus caudatus*) and tarwi (*Lupinus mutabilis*) are ancient crops from the Andean region of South America. Recently, interest in these crops has grown, and worldwide demand for them has increased considerably. The aim of this study was to study the bioactive compounds and nutritional compositions of different varieties/ecotypes of quinoa, kañiwa, kiwicha and tarwi. Proximate, mineral, dietary fibre, fatty acid and amino acid compositions were evaluated. The content of phenolic compounds, tocopherols and phytosterols, and the folic acid and antioxidant capacity, were determined as well. The protein content of the grains was between 13.00% and 20.00%. More important than protein quantity is protein quality, which is demonstrated by the composition of the amino acids. All analysed grains, and especially the kañiwa, had very high lysine content. This amino acid is especially important in vegetarian diets because it is the limiting amino acid in cereal protein. The content of the total phenolic compounds in the studied grains was 27–58 mg gallic acid/100 g of sample. In quinoa, the principal flavonoids found were quercetin and kaempferol, in kañiwa quercetin and isorhamnetin. In kiwicha, no detectable amounts of flavonoids were found. Plant sterols (phytosterols) were another group of biologically active compounds detected. Andean lupin, tarwi, is very rich in oil, which has a beneficial nutritional composition. In conclusion, all studied grains have a very high nutritional value, are interesting sources of bioactive compounds and could be used as ingredients in health-promoting functional foods.

**Keywords:** quinoa; kañiwa; kiwicha; tarwi

#### **1. Introduction**

The Andean region of South America is an important center of the domestication of food crops. This area has a diversity of landscapes and agroecological zones, due to several climates and altitude differences (1500–4200 m). Compared with other regions in the world where crops have been domesticated, the Andean region has its own characteristics. There are no vast, unending plains of uniformly fertile, well-watered land, as in Asia, Europe and the Middle East. Instead, there is an almost total lack of flat, fertile, well-watered soil. The Andean people have always cultivated their crops on tiny plots, one above another up mountain sides, rising thousands of meters [1].

At the time of the European conquest, the Incas cultivated almost as many species of plants as the farmers of all Asia or Europe. It has been estimated that Andean native people domesticated as many as 70 separate crop species [1]. On mountain sides up to four km high along the whole continent, and in climates varying from tropical to polar, they grew roots, grains, legumes, vegetables, fruits and nuts.

1

Andean indigenous grains, such as quinoa (*Chenopodium quinoa*), kañiwa (*Chenopodium pallidicaule),* kiwicha (*Amaranthus caudatus*) and tarwi (*Lupinus mutabilis)*, are good sources of highquality proteins. They contain also dietary fibre and oil with polyunsaturated fatty acids. Dietary fibre is especially important in diets designated for disease risk reduction and the prevention of diabetes and heart disease. Quinoa, kañiwa and kiwicha are sometimes called pseudocereals because of their similarity in chemical composition with common cereals, such as wheat and rice. Tarwi is a leguminous seed grown mainly in Andean highlands.

The Andean indigenous food crops have enormous potential to be used as functional foods in the prevention of chronic diseases, such as cardiovascular diseases, cancer and diabetes. High variability, not only in colours and shapes, but also in primary nutrient constituents and bioactive compounds, has recently been reported. The health-related properties of Andean crops claimed by local people could be partially attributed to the presence of these bioactive compounds.

The objective of this research was to study the chemical composition and some bioactive compounds of different varieties/ecotypes of quinoa, kañiwa, kiwicha and tarwi.

#### **2. Materials and Methods**

Two varieties of quinoa and kiwicha and one variety of tarwi and kañiwa were acquired from the southern Andes of Peru.

Water content, proteins (N × 6.25), fat, crude fibre and ashes were determined according to American Association of Cereal Chemists, AACC (2005) [2].

The total dietary fibre was analysed by an enzymatic-gravimetric method according to the Approved Method of AACC (2005) [2] using the TDF-100 kit from Sigma Chemical Company (St. Louis, MO, USA)

Radical scavenging activity was determined according to the method of Brand-Williams et al. [3] (217) based on the decrease of absorbance at 515 nm produced by reduction of DPPH (2, 2Diphenyl-1-picrylhydrazyl) by an antioxidant. Trolox was used as the reference compound.

The content of total phenolics was analysed according to the method of Swain and Hillis (220).

Tocopherols and tocotrienols were determined based on the direct hydrolysis method reported by Fratianni et al. [4].

The fatty acid composition of the fat fraction was determined after methylation using a modification of the procedure described by Slover and Lanza [5]. Profiling analysis of fatty acid methyl esters was conducted on a 6890N GC-FID gas chromatograph (Agilent) equipped with an Omegawax 250 fused silica capillary column (30 m × 0.25 mm × 0.25 ΐm, Supelco Inc., Bellefonte, PA, USA). The initial oven temperature was held at 160 °C for 1 min, raised to 240 °C at a rate of 4 °C/min, and kept there for 5 min. The injector and detector temperatures were 240 °C and 260 °C, respectively. Helium was used as the carrier gas at a flow rate of 1.1 mL/min. Identification of fatty acid methyl esters was carried out by comparing their retention times with those of standards (Sigma, USA). Results were expressed as percentage of total fatty acid methyl esters analysed.

Mineral: Ca, Mg, P, Fe. Samples were digested in concentrated nitric acid in a Tecator block digestor. ICP-OES = Inductively Coupled Plasma-Optical Emission Spectrometry was used for the determination of mineral and trace elements [6].

#### **3. Results and Discussions**

The proximate composition of the grains is presented in Table 1.


**Table 1.** Chemical composition of Andean grains (g/100 g).

Analysis was made as triplicates.

The protein content of the three Andean grains was between 15.0% and 17.9%. The tarwi had a very high protein content (36.9%). The values of the proteins detected in this study are similar to the values found by Repo-Carrasco-Valencia [7]. The differences between the samples in protein content seem to be more related to the place of cultivation than to variety. The plants use the nitrogen from the soil to produce the proteins. If the soil is rich in nitrogen, the plant produces more proteins than a plant which has been cultivated in nitrogen-poor soil.

In the comparison of the proximate compositions of the Andean grains with the proximate compositions of common cereals, we can find some similarities and differences. The content of total carbohydrates in cereals and Andean grains is similar, about 60–75%. The main carbohydrate is starch in all grains. Thus, they could be used as materials for starch industries. The fat content in Andean grains is considerably higher than the fat content in common cereals (6–7% vs. 2–4%) [8]. The oil is of good nutritional quality, containing the essential fatty acids in proper proportions (see Table 2). Quinoa, kañiwa, kiwicha and tarwi could serve as raw materials to produce edible oils.


**Table 2.** Fatty acid composition of Andean grains (% of total fatty acids).

In Table 3, the contents of phenolic compounds and the antioxidant capacities of Andean grains can be observed. The kañiwa had the highest value compared with the other grains, followed by tarwi, and the kiwicha samples had the lowest values. This sample tendency can be observed in the values of antioxidant capacity. This demonstrates that the phenolic compounds are the main compounds responsible of the antioxidant activity of these Andean grains.

Tocopherols are compounds with a high antioxidant capacity and other important physiological functions. Some of them have the function of vitamin E. The contents of different tocopherols in the Andean grains are presented in Table 4. The highest content of tocopherols was found in black quinoa from Cusco. Kiwicha samples were low in ΅-tocopherol, but interestingly, high in ·-tocopherol. This tocopherol was found in high amounts in kañiwa as well. Wheat has about 1.3 mg/100 g of total ΅ tocopherol, and it does not contain ·-tocopherol [9]. Barley, oat, rye, rice and corn contain the following amounts of total tocopherols: 0.75–0.9, 0.6–1.3, 1.8, 0.2–0.6 and 4.4–5.1 mg/100 g, respectively [8]. Thus, in comparison with common cereals, the Andean grains could be considered good sources of these tocopherols.


**Table 3.** Phenolic compounds and antioxidant capacity in Andean grains.


The contents of some important minerals in Andean grains can be observed in Table 5. In comparison with quinoa and kañiwa samples, the kiwicha samples had the highest calcium content. The black kiwicha was exceptionally rich in this element. This sample had the highest iron and magnesium content, as well. The content of calcium and magnesium in Andean grains is higher than the content of these minerals in wheat, barley, oats and rice, whereas the content of iron is similar [9].


**Table 5.** Content of minerals in Andean grains.

#### **4. Conclusions**

All Andean grains had a relatively high protein and fat content. The pink kiwicha had the highest oil content. This is interesting because kiwicha oil is a very good source of phytosterols, essential fatty acids and, according to the literature, of squalene. This compound is found in olive oil, as well. Squalene is a terpenoid compound and it has some health-promoting properties; in particular it lowers the cholesterol level in blood by inhibiting its synthesis in the liver. In addition, it is hypothesized that the decreased risk of various cancers associated with high olive oil consumption could be due to the presences of squalene. This variety could be an interesting material for producing edible high-quality oil, with bioactive compounds, such as squalene, essential fatty acids and phytosterols. Kañiwa's oil is very interesting, as well. It has a very high content of tocopherols and unsaturated fatty acids.

The best source of antioxidants (phenolic compounds) out of the studied grains was kañiwa. Andean grains contain flavonoids, a type of phenolic compound with important antioxidant activity. Berries, such as blueberries, are generally considered to be excellent sources of flavonoids. Regarding the content of tocopherols, in comparison with common cereals, the Andean grains could be considered good sources of these compounds. In general, these Andean native grains are very rich in

#### *Proceedings* **2020**, *53*, 1

health-promoting compounds, and should be explored in future studies on the bioavailability of these compounds.

**Acknowledgments:** This work was supported by grant IaValSe-Food-CYTED (Ref. 119RT0567), International Trade Centre (ITC), Geneva, Switzerland and PROTEIN2FOOD Project (European Union's Horizon 2020, No. 635727).

#### **References**


© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

*Proceedings* 

### **Potential Beneficial Effects of** *Chenopodium quinoa* **and** *Salvia hispanica* **L. in Glucose Homeostasis in Hyperglycemic Mice Model †**

#### **Raquel Selma-Gracia 1,2, Claudia Monika Haros 2 and José Moisés Laparra 1,\***


Published: 3 August 2020

**Abstract:** Impaired glucose homeostasis is associated with an increased risk of developing metabolic alterations. In this study, a model in which mice treated with streptozotocin were fed a high-fat diet was used to mimic early stages of the onset of metabolic disorders, and different bread formulations were administrated to evaluate the effect of replacing wheat flour with *Chenopodium quinoa* (Q) (25%) and *Salvia hispanica* L. (Sh) (20%). Plasmatic glucose and insulin concentrations were quantified and the homeostasis model assessment (HOMAir) was calculated. Q and Sh showed a lower tendency to hyperglycemia compared to wheat bread (WB). Besides, these low glucose levels were accompanied by three-fold lower values of HOMAir respect to WB, suggesting an improved insulin sensitivity. Thus, inclusion of *C. quinoa* and *S. hispanica* into bread formulations could improve the control of glucose homeostasis, which could help to prevent/ameliorate metabolic glucose alterations.

**Keywords:** *Chenopodium quinoa*; *Salvia hispanica* L.; hyperglycemia; HOMAir

#### **1. Introduction**

A good control in glucose homeostasis has shown to be determinant in the prevention and improvement of metabolic disorders, such as insulin resistance, obesity, or type 2 diabetes (T2D) [1]. In recent years, Latin-American crops have become a focus of study because of their beneficial nutritional properties [2]. *Chenopodium quinoa* and *Salvia hispanica* L. in bread formulations [3] have shown an enhanced insulin signaling modulating peroxisome proliferator-activated receptor (PPAR)-· activation, which has been related to a marked improvement of whole-body insulin sensitivity in T2D patients [2,4].

Thus, the objective of the present investigation was to evaluate the inclusion of flour from *C. quinoa* and *S. hispanica* as a substitute to wheat flour in bread formulations in the control of glucose homeostasis in a hyperglycemic-mouse model.

#### **2. Materials and Methods**

#### *2.1. Sample Breads*

White quinoa seeds (Organic quinoa Real©) from ANAPQUI, La Paz, Bolivia, were purchased from Ekologikoak (Bizkaia, Spain). Chia flour was purchased from the Primaria Premium Raw Materials Company (Valencia, Spain). Wheat flour was provided from a local market. Three bread formulations were prepared: white quinoa (Q) at 25% and chia (Sh) at 20% and were compared to wheat bread (WB), as a control [3].

#### *2.2. Experiment Design*

Female C57BL/6 mice were obtained from Centro de Investigaciones Biológicas (CIB-CSIC) in Madrid, Spain. All animals were injected intraperitoneally with 2 low doses of streptozotocin (25 mg/kg) and they received a high-fat diet for 3 weeks. Animals were distributed into groups depending on the administered bread, and they received 3 doses (14 mg/day/animal) per week during all the treatment.

#### *2.3. Glucose and Insulin Concentration*

Insulin and glucose concentrations were determined in plasma samples. Insulin was measured by ELISA kit (RAB0817-1KT, Sigma-Aldrich, Darmstadt, Germany) and glucose (MAK263-1KT, Sigma-Aldrich, Darmstadt, Germany).

#### *2.4. Homeostatic Model Assessment of Insulin Resistance (HOMAir)*

Insulin resistance (IR) was defined as the HOMAir value and was calculated according to the following formula: [insulin (ΐU/mL) × glycaemia (mg/dL)]/405 [5].

#### *2.5. Statistical Analysis*

Multiple ANOVA and Fisher's least significant differences (LSD) were applied to establish statistically significant differences. Statistical analyses were performed with the software Statgraphics Centurion XVI and the significance level was established at *p* < 0.05.

#### **3. Results and Discussion**

This study showed that metabolic alterations occurring at early stages, onset, and disease progression derived from impairment of glucose homeostasis, improved with the inclusion of flours obtained from *C. quinoa* and *S. hispanica* into bread formulations. Hyperglycemia is a hallmark of a profoundly altered metabolism, which is associated with stress oxidative and inflammatory response in the liver [1]. In this study, glucose blood concentrations decreased, following this order: WB > Q > Sh (Figure 1), displaying a positive effect in Q and Sh and suggesting a more controlled glucose homeostasis, which could have important consequences limiting the hepatic endogenous glucose output and liver metabolic stress [6]. The glycemic index is used to determinate the total rise in blood glucose level; however, it could not reflect the real absorption and modulation in glucose homeostasis. The latter is supported by the lack of differences in the glycemic index reported between *C. quinoa* and WB and the higher transcript levels of PPAR-· in *C. quinoa* [2]. These observations are concordant with the low HOMAir value reported in animals fed *C. quinoa* (Figure 1).

HOMAir is considered an index of insulin resistance, which is a risk factor of the progression of type 2 diabetes. Animals administered with WB showed values above 3.8, which is indicative of insulin resistance, meanwhile Q and Sh were below. In line with the glucose levels, Sh obtained 1.5 fold lower HOMAir than Q. The effect of *S. hispanica* seeds in a state of insulin resistance has shown an improved sensitivity associated with increased expression levels of the peroxisome proliferatoractivated receptor-· coactivator-1΅ (PGC-1΅) and promoting the absorption of glucose by muscle in obese rats [7]. In line with these positive effects of flour from S. *hispanica*, it was demonstrated that

#### *Proceedings* **2020**, *53*, 2

long-term supplementation with *S. hispanica* attenuates cardiovascular risk factors with the reduction of systolic blood pressure and serum C-reactive protein concentration in diabetic patients [8].

**Figure 1.** Glucose and HOMAir in mice fed with a high-fat diet and administered with the different bread formulations: WB, white bread; Q, quinoa flour (25%)-containing bread; Sh, chia flour (20%) containing bread. \* Indicates statistically significant (*p* < 0.05) differences in relation to WB.

#### **4. Conclusions**

Replacing flour wheat with *S. hispanica* and *C. quinoa* flour could exert beneficial effects, improving insulin resistance and control of glucose homeostasis and, thus, promoting a better metabolic modulation that could prevent the onset or progress of early metabolic alterations.

**Funding:** This work was financially supported by Ia ValSe-Food-CYTED (119RT0567) and QuiSalhis-Food (AGL2016-75687-C2-1-R) from the Ministry of Economy, Industry and Competitiveness (MEIC).

**Acknowledgments:** The contract given to R. Selma-Gracia by LINCE (PROMETEO/2017/189) from the Generalitat Valenciana (Spain) is gratefully acknowledged. JML thanks MICINN for his 'Ramon y Cajal' contract (RYC-2015-18083).

#### **References**


*Proceedings* **2020**, *53*, 2

8. Vuksan, V.; Whitham, D.; Sievenpiper, J.L.; Jenkins, A.L.; Rogovik, A.L.; Bazinet, R.P.; Vidgen, E.; Hanna, A. Supplementation of conventional therapy a with the novel grain salba (*Salvia hispanica* L.) improves major and emerging cardiovascular risk factors in type 2 diabetes: Results of a randomized controlled trial. *Diabetes Care* **2007**, *30*, 2804–2810, doi:10.2337/dc07–1144.

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

#### *Proceedings*
