Agrarian Academic Journal

agrariacad.com

doi: 10.32406/v7n5/2024/19-33/agrariacad

 

In vitro antioxidant, antimicrobial and antidiabetic properties of the organic fraction of distillate of Salvia hispanica seeds. Propriedades antioxidantes, antimicrobiana e antidiabéticas in vitro da fração orgânica do destilado de sementes de Salvia hispanica.

 

Ibtissem Rahmoune^1,2*, Samira Karoune^2*, Clara Azzam³, Somia Saad², Abdelhamid Foughalia², Madani Sarri⁴, Farid Chebrouk⁵, Hana Abidat^1,2, Mohamed Seif Allah Kechebar²

 

^1- Laboratory of Functional Ecology and Environment, Larbi Ben M’Hidi, University of Oum El Bouaghi, Algeria. E-mail: rahmoune.ibtissam@univ-oeb.dz

^2- Center for Scientific and Technical Research on Arid Regions – CRSTRA, Omar El Bernaoui, Biskra, Algeria. E-mail: karounesamira@yahoo.fr

^3- Cell Research Department, Field Crop Research Institute, Agricultural Research Center, Giza, Egypt.

^4- Department of Nature and Life Sciences, Faculty of Sciences, M’sila University, M’sila, Algeria. E-mail: madani.sarri@univ-msila.dz

^5- Center for Scientific and Technical Research in Physico-Chemical Analyzes, Tipaza, Algeria.

 

Abstract

 

This work aims to test in vitro biological properties and phytochemical composition of the organic fraction of distillate of Salvia hispanica seeds. Twenty-five bioactive compounds were identified, constituting 90.40% of the total organic fraction. The chemical composition of organic fraction revealed that fatty acids were the main component at 78.93%, followed by steroids (6.95%) and tocophérol (1.87%). The organic fraction showed interesting antioxidant activity for DPPH, DMSO, ABTS, PHE and FRAP tests. Furthermore, the in vitro enzyme inhibitory activity against α-amylase exhibited the best activity (IC₅₀ = 34.97 ± 0.68 μg/mL). The results indicated that organic fraction displayed good antimicrobial effect.

Keywords: Chia seeds. Phytochemical analysis. Biological properties. GC/MS. Fatty acids.

 

 

Resumo

 

Este trabalho tem como objetivo testar propriedades biológicas in vitro e composição fitoquímica da fração orgânica do destilado de sementes de Salvia hispanica. Vinte e cinco compostos bioativos foram identificados, constituindo 90.40% da fração orgânica total. A composição química da fração orgânica revelou que os ácidos graxos foram o principal componente em 78.93%, seguidos por esteroides (6.95%) e tocoferol (1.87%). A fração orgânica mostrou atividade antioxidante interessante para os testes DPPH, DMSO, ABTS, PHE e FRAP. Além disso, a atividade inibitória enzimática in vitro contra α-amilase exibiu a melhor atividade (IC₅₀ = 34.97 ± 0.68 μg/mL). Os resultados indicaram que a fração orgânica apresentou bom efeito antimicrobiano.

Palavras-chave: Sementes de chia. Análise fitoquímica. Propriedades biológicas. GC/MS. Ácidos graxos.

 

 

Introduction

 

Salvia hispanica, also known as Chia or Spanish sage, is a food crop newly recognized for its pharmacological and nutritional properties that may have enormous application potential in food systems (GRANCIERI et al., 2019). It is an annual herbal plant from the Lamiaceae family, native to Central America (CAHILL, 2003).

Currently, Chia seeds are becoming increasingly popular in modern diets due to their essential nutrients and health-enhancing activity that have been recognized in some of their components, including their high dietary fiber content (OLIVOS-LUGO et al., 2010), carbohydrates (LIN et al., 1994), fatty acids (ISHAK et al., 2021), proteins (SANDOVAL-OLIVEROS & PAREDES-LÓPEZ, 2013), and high levels of phenolic compounds (MARTÍNEZ-CRUZ & PAREDES-LÓPEZ, 2014) that display various physiological function properties, including antioxidant, antibacterial, anticancer, and anti-inflammatory.

Oxidation, a fundamental metabolic process, is responsible for breaking down organic molecules to produce energy and generating reactive oxygen species (ROS) as byproducts. While these ROS are natural byproducts, they can lead to oxidative stress, a factor implicated in a range of human diseases, including diabetes, cancer, skin pigmentation problems, and obesity (KOEPKE et al., 2008). Type 2diabetes, a prevalent chronic disease, is regulated by oxidative stress through molecular mechanisms (FOLLI et al., 2011). In this condition, the body resists insulin, a hormone crucial for blood sugar regulation. This resistance leads to high blood sugar levels, which is known as hyperglycemia.

The activity of digestive enzymes, particularly α-amylase, significantly influences blood sugar levels. It breaks down complex carbohydrates, such as starch, into simpler sugars, like glucose, which can then be absorbed by the body and released into the bloodstream to provide energy for cells and tissues. Therefore, inhibiting α-amylase activity can reduce the release of glucose into the blood stream (TARLING et al., 2008). The combination of antioxidants and α-amylase suppression holds promise for the treatment of type 2 diabetes, cancer, and obesity.

Several natural compounds, such as polyphenols, fatty acids, tocopherols, and steroids, are present in seeds, fruits, vegetables, and certain herbs. These compounds possess antioxidant properties, α-amylase inhibitory activity, and antibacterial effects. They have enormous potential in preventing and treating various diseases, thus underscoring the importance of exploring natural therapies in medical research.

Based on the reasons indicated above, this research was conducted with the aim of in vitro evaluation of the antioxidant, antidiabetic, and antibacterial properties of the organic fraction distillate from S. hispanica seeds (OFSH) extracted using a Clevenger distillation apparatus. The composition of the organic fraction was examined using GC/MS. The antioxidant properties were evaluated using seven assays, including ABTS, DPPH, PHE, FRAP, alkaline- DMSO, CUPRAC, and PRAC. The antidiabetic activity was evaluated by the α-amylase enzyme. Further, the antimicrobial effect was studied using the disc diffusion method against eight bacterial species and one pathogenic yeast.

 

Materials and methods

 

Organic fraction extraction

 

One hundred grams of powdered Salvia hispanica seeds (purchased from Algeria) were distilled using traditional hydro distillation with 1L of distilled water for four hours using a Clevenger-type apparatus. The resulting fraction yield was 1.9 g. Then this fraction was stored in screw-capped glass vials in a refrigerator at – 4°C. The yield was estimated based on the dry weight of the sample.

 

Free radical scavenging test (DPPH)

 

The antioxidant capacity of the OFSH was tested using DPPH test as published (BOUCHOUKH et al., 2019). Briefly, a 96-well microplate was used to mix 40 μL of diluted fraction dissolved in DMSO at different concentrations (from 12.5 to 800 μg/mL) with 160 μL of a 0.1 mM DPPH solution. At a wavelength of 517 nm, the absorbance was read using a Multiskan Sky Plate reader from Singapore (5111700DP) after incubating the mixture for 30 minutes in the darkness at room temperature. BHT was used as a positive reference. The DPPH inhibition (%) was calculated using the following equation (1). The IC₅₀, or sample concentration that lowered 50% of free radicals was obtained by plotting the acquired inhibitions against the sample concentrations.

(1) DPPH+ scavenging ability (%) = (A₀ – A₁)/A_0,

where A₀ and A₁ are the absorbance of control and fraction at 30 min, respectively.

 

Free radical scavenging test (ABTS)

 

The capacity of ABTS to scavenge free radicals was evaluated using the described procedure (RE et al., 1999). Briefly, 5 mL of 7.7 mM ABTS solution was blended with 5 mL of 2.45 mM potassium peroxide to produce the free radical cation ABTS+. The resulting mixture was diluted with H₂O until it exhibited an absorption of 0.7± 0.02 at a wavelength of 734 nm after incubation for 16 hours in the darkness at room temperature. Then, 40 µL of OFSH at different concentrations (from 12.5 to 800µg/mL) were combined with 160 µL of fresh ABTS+ mixture. At 734 nm, the absorbance was recorded after 10 minutes of incubation in the darkness at room temperature. BHT was used as a positive control. The radical scavenging ratio of ABTS+ was determined based on the formula (2) below:

(2) ABTS+ scavenging ability (%) = (A_b – A_f)/A_b,

where A_b and A_f are the absorbance of control and fraction at 10 min, respectively.

 

Ferric reducing power test

 

The FRAP test involves the reduction of iron oxide to ferrous ions. The potential reduction of OFSH was estimated using the published protocol (OYAIZU, 1986). Briefly, 40 μl of phosphate buffer solution (PBS: 0.2 M with a pH=6.6), 50 μL of potassium ferricyanide (C₆FeK₃N₆, 1%) were mixed with 10 μL of OFSH at various concentrations (from 3.125 to 200 μg/mL). After leaving the mixture in the incubator at 50°C for 20 min, 50 µL of 10% trichloroacetic acid (TCA) solution and 10 µl of 0.1% iron chloride were added to the sample and supplemented with 40 µl of H₂O. At a wavelength of 700 nm, the absorbance was recorded, and the results are presented as A_0.5 values (µg/mL) using ascorbic acid as a positive reference.

 

Superoxide radical test

 

Superoxide radical activity was assessed using the method described (KUNCHANDY & RAO, 1990). Briefly, in a 96-well microplate, 40 μL of OFSH at different concentrations (from 12.5 to 800 μg/ml) were mixed with130 μL of alkaline DMSO (20 mg of sodium hydroxide in 1 mL of H₂O and supplement with DMSO to 100 mL), followed by 30 μl of nitroblue tetrazolium (1 mg/mL). At the wavelength of 560 nm, the absorbance was recorded directly at 560 nm. The findings were expressed as BHT equivalent used as a positive control. The formula below (3) was used to calculate the percentage of superoxide inhibition:

(3) % inhibition = (Ac − At / Ac) × 100; A control absorbance is denoted by (Ac), while OFSH absorbance is denoted by (At).

The resulting inhibitors were graphed against the concentrations of the sample, and the graphs were used to calculate the IC₅₀, which is the concentration that reduces superoxide by 50%.

 

Cupric reducing antioxidant activity

 

The CUPRAC test was conducted to assess the antioxidant capacity of OFSH to reduce copper in the presence of neocuproine, using the published protocol (APAK et al., 2004). The test involves measuring the ability of antioxidants to reduce Cu⁺² to Cu⁺. In a microplate 96 well, 60 µL of ammonium acetate solution (1 M, pH 7), 50 µL of neocuproine and 50 µL of copper chloride (CuCl₂) solution (10 mM) were added to 40 µl of OFSH at various concentrations (from 12.5 to 400 µg/mL). The reaction was incubated for 1h, and the absorption was recorded at a wavelength of 450 nm. A_0.5 (µg/mL) was used to determine antioxidant activity results.

 

Phenanthroline test

 

This test was performed based on the method (SZYDŁOWSKA-CZERNIAK et al., 2008) which includes the capacity of the antioxidant to form a complex with 1,10-phenanthroline to reduce Fe³⁺ to Fe²⁺. Briefly, in a microplate, 50 µL of iron chloride (0.2%), then 30 µL of 1,10-phenanthroline and 110 µL of methanol were added to 10 µL of OFSH at different concentrations (from 3.125 to 200 μg/mL). At a wavelength of 510 nm, the absorbance was recorded after leaving the mixture in the incubator at 30°C for 20 minutes. BHT was used as a standard, and the results are presented in A_0.5 values (µg/mL).

 

The permanganate reducing antioxidant capacity

 

The antioxidant activity of OFSH was measured using the PRAC test as published (EFTIMOVÁ et al., 2018). Briefly, 10 µl of OFSH at various concentrations was placed in a microplate. Then, 130 µL of potassium permanganate (KMnO₄) solution and 60 µL of sulfuric acid (H₂SO₄) were added. After incubating the mixture for 5 minutes, the absorbance at 535 nm was measured. BHT was used as a positive reference. The results are presented as a proportion decrease in potassium permanganate. The results are presented as A_0.5 values (µg/mL).

 

Antimicrobial activity

 

The antimicrobial activity of the organic fraction from S. hispanica seeds was evaluated using the disc diffusion method (PEREZ et al., 1990) against eight pathogenic bacteria Escherichia coli (ATCC 25922), Klebsiella pneumonia (ATCC 13883), Vibrio cholera (ATCC 14035), Staphylococcus aureus (ATCC 25923), Listeria monocytogenes (ATCC 35152), Bacillus cereus (ATCC 14579), Salmonella typhimurium (ATCC 14028), Pseudomonas aeruginosa (ATCC 27853) and one pathogenic yeast: Candida albicans (ATCC 10231).

Two different media were used in this part, Muller-Hinton (MH) agar (Biokar Diagnostics, REF 048HA; for the bacterial activity) and Sabouraud agar (Biokar Diagnostics, REF BK027HA; for the antifungal activity against “C. albicans”. The media were poured into sterile Petri dishes and after solidification their surfaces were inoculated with the corresponding microbial suspensions using sterile swabs (at 0.5 McFarland). Next, sterile 6 mm discs (Whatman filter paper) soaked with the extract (dissolved in 2.5% DMSO (at 100 mg/mL) and sterilized through 0.45 µm microfilters) were placed on the surface of the inoculated media.

Disks impregnated with Gentamicin (against bacteria) and amphotericin B (against C. albicans) were used as a positive reference. The plates were then incubated at 37°C for 24 h. After the incubation, the antimicrobial activity was assessed by measuring the diameters of the inhibition zones around each disc. Three replicate plates were inoculated with the same microbial suspension and the whole experiment was repeated thrice (independent repetitions).

 

The α-amylase inhibition activity

 

This experiment aimed to demonstrate the ability of organic fraction from S. hispanica seeds to inhibit α-amylase based on the proposed procedure (ZENGIN et al., 2014). Briefly, 50 μL of α-amylase solution prepared in PBS (pH 6.9) was combined with 25 μL of diluted fraction at different concentrations (from 6.25 to 400 μg/mL). After 10 minutes of incubation at 37°C, 50 µl of 0.1% soluble starch was added to the mixture, and the reaction was incubated for an additional 20 minutes at 37°C. Then, 25 µl of 1M HCl solution was added, followed by 100 µl of iodine/potassium iodine (IKI). At 630 nm, the absorbance was read, and the inhibition percentage of the alpha-amylase enzyme was calculated based on the formula (4) below:

(4) I% = 1 − [(Absc − Abse) − (Abss − Absb)/(Absc − Abse)], where:

Absc: Absorbance [starch+ IKI+ HCL+ Volume of studied fraction+ Volume buffer enzyme];

Abse: Absorbance [starch+ IKI+ HCL+ enzyme];

Abss: Absorbance [starch+ IKI+ HCL+ Volume of studied fraction+ enzyme];

Absb: Absorbance [IKI+ Buffer solution+ Volume of studied fraction].

Chemical analyses

 

The Hewlett-Packard computerized system was used to examine the organic fraction of Chia. This system included a 6890-GC connected to a 5973A MSD and a 30 m × 0.25 mm ID HP-5MS capillary column with a film thickness of 0.25 μm. With an injection volume of 1 μL and an injection temperature of 250°C in split less mode, helium was used as the carrier gas, moving forward at a rate of 0.5 mL/min. Starting at 50°C and maintained for 5 minutes, the oven temperature was programmed to rise to 300°C at a rate of 15°C /min, which was held for 10 minutes. The ionization potential of 70 V, scanning time, and mass range were 2.83 s and 55-550 m/z, respectively. The organic fraction components were identified by comparing mass spectral fragmentation patterns with those stored in the NIST 2020 MS database, Wiley 07 libraries and mass spectra reported in the literature. The concentration of each compound was represented as a percentage (content) and calculated from the corresponding chromatographic peak areas using ChemStation software.

 

Results and discussion

 

Chemical composition of the organic fraction

 

The results of the analysis and identification of components of the organic fraction of S. hispanica obtained using GC-MS are presented in Table 1. Twenty-five bioactive compounds were identified, which constitute an average of 90.40% of the total organic fraction. The chemical composition of OFSH revealed that fatty acids were the main component by 78.93%, followed by steroids (6.95%) and tocopherol (1.87%). Among the fatty acids, omega-6 linoleic acid was the dominant compound, accounting for 57.43%, followed by palmitic acid (8.44%), α-glyceryl linoleate (7.26%), ethyl linoleate (3.35%), and methyl linoleate (1.57%). Our findings are consistent with various studies that suggest that the primary components of S. hispanica organic fraction are polyunsaturated fatty acids. For instance, one research revealed that the oil extracted by different solvents contained linoleic acid alpha and linolenic acid (SILVA et al., 2016). In another study conducted by Tolentino et al. (2014), on the fatty acid content of Chia seeds grown in four states in Mexico, chemical analysis revealed nine fatty acids in Chia oil samples, including alpha-acid, omega-6 acid, and palmitic acid. The percentage of linoleic acid reached 20.57% (GHAFOOR et al., 2020).

 

Table 1 – Chemical components of the organic fraction of S. hispanica seeds

N°	RTa	Componentsb	%c
1	7.44	n-Butylbenzène	0.04
2	7.92	1-Methyl-2-n-propylbenzene	0.09
3	8.00	n-Undécane	0.05
4	8.40	β-Ethylstyrene	0.14
5	10.32	p-Ethylguaiacol	0.20
6	11.12	n-Heptylbenzene	0.06
7	11.2	p-Propylguaiacol	0.13
8	11.37	n-Tetradec-1-ene	0.05
9	11.44	3-Methylindole (Skatol)	0.16
10	11.96	(Z)-Isoeugenol	0.16
11	12.06	n-Octylbenzene	0.08
12	12.31	1-Pentadecene	0.15
13	12.88	Pentadecane	0.21
14	15.32	Palmitonitrile	1.13
15	15.48	Methyl palmitate	0.88
16	15.85	Palmitic acid	8.44
17	16.50	Methyl linoleate	1.57
18	17.15	Linoleic acid	57.43
19	18.22	Ethyl linoleate	3.35
20	19.90	α-Glyceryllinoleate	7.26
21	21.61	β-Tocophérol	1.87
22	21.92	Stigmasta-3,5-diene	3.04
23	22.01	16,17-Didehydropregnenolone acetate	0.65
24	22.96	Methyl glycocholate	0.36
25	23.38	β-Sitostérol	2.90
		Total identified (%)	90.40
		Grouped components
		Fatty acid derivatives	78.93
		Steroids	6.95
		Tocopherol	1.87
		Hydrocarbon derivatives	0.46
		Others	2.19

^a Retention times (RT); ^b Components are listed according to their elution from a HP-5MS column; ^c Relative peak area percentage.

 

Antioxidant properties

 

Seven distinct tests were used to understand the antioxidant effect of the organic fraction of S. hispanica that was extracted using hydro distillation. These tests included the DPPH, ABTS, phenantroline, PRAC, FRAP, CUPRAC and alkaline-DMSO. In Table 2, the data are presented in the form of IC₅₀ and A_0.5 values, and the results were compared with those of butylhydroxytoluene (BHT) and ascorbic acid as positive controls. DPPH (2,2-Diphenyl-1-picrylhydrazyl) is a stable free radical frequently used to test the ability of compounds to become a stable diamagnetic molecule by taking an electron or hydrogen radical (BOUCHOUKH et al., 2019). The OFSH showed good antioxidant activity, with an IC₅₀ value = 71.36 ± 2.93 µg/mL, comparable to BHT with an IC₅₀ value of less than 12.5 µg/mL.

Several studies have used the DPPH assay to estimate the antioxidant activity of S. hispanica. According to the report (SCAPIN et al., 2016), who examined the antioxidant effect of the ethanolic extract of S. hispanica purchased from commercial establishments in Brazil, in January 2013 and the IC₅₀ value obtained was 3.841 mg/mL. In a study (TUNÇİL & ÇELİK, 2019) to compare the compositional properties of white and black Chia seeds commercially available in Turkey, reported an excellent IC₅₀ values for white and black Chia extract (IC₅₀: 1.735 mg/mL and 2.001 mg/mL), respectively. A recent investigation (ABOU ZEID et al., 2022) reported high DPPH scavenging percentage of ethyl acetate extract from aerial parts of S. hispanica with IC₅₀ = 13.11 μg/mL.

Regarding the ABTS test, the OFSH displayed the best activity (IC₅₀ = 14.7 ± 1.7μg/mL). This activity was similar to that of BHT, which exhibited an IC₅₀ value less than 12.5 μg/mL. This result is similar to the IC₅₀ value obtained for black Chia seed ethanol extract (IC₅₀ = 18.11μg/mL) (BANU et al., 2021). However, our sample showed more significant ABTS radical scavenging activity compared to the findings published in previous studies on salvia species (KATANIĆ STANKOVIĆ et al., 2020; MOKHTAR et al., 2023).

The analyzed data in Table 2 for the superoxide radical scavenging test using the alkaline DMSO assay demonstrated that the OFSH seeds exhibited the best activity, with an IC₅₀ value of less than 12.5 μg/mL which is higher than the activity of BHT (IC₅₀ = 40.21 ± 0.3μg/mL). The present results of our study are consistent with the findings (BANU et al., 2021), who reported an IC₅₀ value equal to 14.10 μg/mL. The ability of different Salvia species to scavenge the superoxide (O₂ •-) anion radical has also been the subject of investigations conducted similarly and published (MAMACHE et al., 2020).

 

Table 2 – Antioxidants activity of the organic fraction of S. hispanica seeds

Samples	OFSH	BHT	Ascrobic acid
DPPH IC50(μg/mL)	71.36 ± 2.93	< 12.5	–
ABTS IC50 (μg/mL)	14.9 ± 1.7	<12.5	–
PHE A0.5 (μg/mL)	17.2±2.8	9.71±0.9	–
FRAP A0.5 (μg/mL)	39.3±3.5	–	6.52±0.07
Alkaline-DMSO IC50 (μg/mL	<12.5	40.21±0.3	–
CUPRAC A0.5 (μg/mL)	360.74± 8.6	–	123.02±1.6
PRAC A0.5 (μg/mL)	189.0 ± 0.9	<3.125	–

 

Based on the evaluation of the reduction of metallic iron by the phenanthroline test, it was shown that the OFSH showed a significant reducing capacity (A_0.5 = 17.2 ± 2.8 μg/mL) which is very close to the BHT (A_0.5 = 9.71 ± 0.9 μg/mL). These findings are consistent with the study (MAMACHE et al., 2020) for methanolic extracts of S. verbenaca and S. aegyptiaca obtained from aerial parts (A_0.5 = 18.71 ± 1.03 and 27.03 ± 1.54 μg/mL, respectively).

The reductive ability of organic fraction of S. hispanica seeds depends on reducing ferric ions to the ferrous form in the presence of a reducing agent (an antioxidant). OFSH exhibited a significant reduction potential (A_0.5 = 39.3 ± 3.5 μg/mL) compared with standard ascorbic acid (A_0.5 = 6.52 ± 0.07 μg/mL). The current findings for the studied fraction agreed with the activity value previously reported for ethyl acetate extract from S. hispanica seeds (DIB et al., 2021). The results of our FRAP investigations are better than those reported in a similar study on other Sage species, such as S. nutans, S. nemorasa and S. austriaca (A_0.5= 52.08 ± 0.01; 55.61 ± 0.3 and 80.02 ± 0.05 μg/mL, respectively) (LUCA et al., 2023).

For the permanganate-reducing antioxidant capacity (PRAC) assay, which is based on the reduction of Mn(VII) to Mn(II), the OFSH showed low antioxidant activity (IC₅₀ = 189.0 ± 0.9 μg/mL) compared to BHT < 3.125. The chemiluminescence detection of permanganate is a suitable method to evaluate the antioxidant activity of various beverages, including fruit juices, teas, and other drinks (POPOVIĆ et al., 2012). However, this test has not been examined with salvia species.

On the other hand, the CUPRAC (Cupric ion reducing antioxidant capacity) test involves a reduction in the presence of antioxidants, as indicated by the change from a stable copper (II) – neocuproine complex (blue color) to a stable copper (I) – neocuproine complex (orange color). In this test, the OFSH exhibited low activity (A_0.5 = 360.74 ± 8.68 μg/mL) compared to BHT (A_0.5 = 123.02 ± 1.6 μg/mL). Salvia species showed comparable findings (CHOUIT et al., 2021; MOKHTAR et al., 2023).

 

Antimicrobial activity

 

Using the disc diffusion method, the antimicrobial properties of the organic fraction of S. hispanica seeds dissolved in DMSO were examined against Gram-positive, Gram-negative bacteria and a pathogenic yeast: C. albicans. The results of this evaluation are shown in Table 3. OFSH demonstrated good antagonistic effect against Salmonella, L. monocytogenes and B. cereus, where diameters of the zone of inhibition ranged from 10.33 mm to 12.67 mm. However, OFSH showed no antagonistic effect against S. aureus, E. coli, K. pneumoniae, V. cholerae, P. aeruginosa, and C. albicans. Salmonella was the most susceptible bacterium to OFSH (inhibition zone of 12.67 mm). It should be noted that these results were obtained using a concentration of 100 mg/mL of OFSH. Our findings are in keeping with previous studies. For instance, Elshafie et al. (2018), studied the antimicrobial effect of the essential oil of the aerial parts of S. hispanica against some of phytopathogenic fungi and bacteria. The study showed that the essential oil of Chia significantly reduced the growth of C. michiganensis, B. megaterium, P. savastanoi, B. mojavensis, X. vesicatoria, X. campestris and P. syringae pv. phaseolicola.

In addition, Banu et al. (2021) reported the antibacterial effect of the ethanol fraction of Chia seeds against S. aureus, E. coli, M. luteus, B. subtilis and S. flexneri. Among these, S. aureus was the most susceptible bacterium, with a zone of inhibition of 31 mm at a concentration of 500 μg/mL. Mokhtar et al. (2023) showed the antibacterial effect of the Chia leaves extracts against Gram-positive bacteria, including E. faecalis, M. luteus, S. aureus, and S. epidermidis.

 

Table 3 – Antimicrobial activity of organic fraction of S. hispanica seeds

Strains	Inhibition zone (mm)	Positive control
E. coli	6.67 ± 0.5	22.33 ± 0.5
K. pneumoniae	6.17 ± 0.2	28.67 ± 1.5
V. cholerae	6.17 ± 0.2	36.67 ± 0.5
S. aureus	7.17±0.2	36.33 ± 0.5
L. monocytogenes	12 ± 1	26.67 ± 05
B. cereus	10.33 ± 0.5	34.67 ± 0.5
S. typhimurium	12.67 ± 0.5	33.67 ± 1.1
P. aeruginosa	6.17 ± 0.2	26.67 ± 0.5
C. albicans	6.17 ± 0.2	19.33 ± 0.5

 

On the other hand, our findings showed that OFSH was able to inhibit the growth of two Gram-positive (L. monocytogenes and B. cereus) and one Gram-negative (S. typhimurium).

These bacteria are known to be responsible for some food poisoning and can develop new antibio-resistance to some antibiotics, which allows us to say that our OFSH can constitute a good alternative to antibiotics.

 

In vitro enzyme inhibitory activity

 

Enzyme inhibition is one of the most intriguing and extensively researched therapeutic strategies for the pharmaceutical and cosmetic sectors. These drugs are utilized in clinical settings to treat a variety of illnesses, including diabetes, obesity, and Alzheimer’s disease (ÖKTEN et al., 2019).

It has been found that synthetic inhibitors can cause adverse effects such as gastrointestinal problems and liver toxicity. However, there is great interest in discovering natural inhibitors without side effects compared to synthetic inhibitors (GONÇALVES & ROMANO, 2017; RODRIGUES et al., 2018). OFSH showed the highest α-amylase inhibitory activity (IC₅₀ = 34.97 ± 0.68 μg/mL) compared to the reference Acarbose enzyme (IC₅₀ = 6.96 ± 0.4 μg/mL) (Figure 1). A recent study (ABDEL GHANI et al., 2023) to evaluate the anti-diabetic effect of dichloromethane extract from the aerial parts revealed biological results (IC₅₀ = 673.25 μg/mL) (BANU et al., 2021), found that the inhibitory effect of Chia seeds was IC₅₀ = 121.46 μg/mL. However, the enzyme inhibitory activity assay results in our studies are better compared to many chemical studies that have been reported on Salvia species; for example, a study reported an inhibitory effect on α-amylase of methanolic fractions of S. officinalis, S. macilenta, S. mirzayanii and S. santolinifola (IC₅₀ = 54.9 ± 5.7; 103.7 ± 1.1; 114.8 ± 11.1 and 54.7 ± 9.6ug/mL) (JAVID et al., 2022), S. aegyptiaca and S. verbenaca (IC₅₀= 86.12 ± 0.08 and 101.30 ± 0.08 μg/mL), respectively (MAMACHE et al., 2020).

In this study, the organic fraction from Chia seeds showed higher antioxidant, antidiabetic, and antibacterial activities, which can be attributed to the richness of bioactive compounds. These compounds include fatty acids, steroids, and tocopherols. The organic fraction contained 78.92% fatty acid derivatives (such as methyl palmitate, palmitic acid, methyl linoleate, linoleic acid, ethyl linoleate, and α-glyceryl linoleate), 6.95% steroids (including stigmasta-3,5-diene, 16,17 didehydropregnenolone acetate, methyl glycocholate, and β-sitosterol), and 1.87% tocopherols. Fatty acids (FAs) play essential roles in preventing many diseases and abnormal differentiation problems. A recent study reported the importance of fatty acids in fighting cardiovascular diseases, inflammatory reactions, and their antioxidant activities (GAWRON-SKARBEK et al., 2023). Linoleic acid, or omega 6 (LA), is one of the polyunsaturated fatty acids (PUFA) that plays a crucial role in the human body and works to reduce triglyceride levels, thus protecting against the risks of heart disease, strokes, and cancer. It is found in many foods, such as nuts, seeds, and vegetable oils. The percentage of linoleic acid in the extracted sample was 57.43%. These data indicate that linoleic acid (LA) would protect against free radicals according to the study (HENRY et al., 2002), which reported that several essential fatty acids FAs including (linolenic acid, linoleic acid, palmitic acid, oleic acid, and lauric acid ) can act as antioxidants or pro-oxidants (LALITHADEVI et al., 2018) reported that conjugated linoleic acid (CLA) has potential health benefits such as antioxidant properties, weight loss, and anti-aging. LA has been shown to inhibit the growth of colorectal cancer cells by enhancing the cellular redox state and promoting the prevention of type 2 diabetes (LU et al., 2010; ZONG et al., 2019). Another study, (YOON et al., 2021), reported that omega 6 contains a multi-targeted inhibitor of non-receptor type protein tyrosine phosphatase (types 1, 9, and 11) and has antidiabetic activity (T2DM). FAs displayed antibacterial activity against some strains (S. aureus and B. subtilis). Stigmata 3,5-diene is a well-known plant sterol that has gained popularity in traditional medicine due to its various biological properties, including antioxidant, anti-inflammatory, antibacterial, antiviral, antidiabetic, antifungal, anti-immune, antiparasitic, and neuroprotective properties (KUSUMAH et al., 2020).

 

Figure 1 – IC₅₀ values of α-amylase inhibitory activity of the organic fraction of Chia seeds.

 

One report indicated that Sigmasta 3,5-diene has significant anti-inflammatory effects and can exert antidiabetic properties by lowering fasting glucose and blood insulin levels (BAKRIM et al., 2022). The β-sitosterol derivative showed antimicrobial activity against Salmonella typhimurium, Staphylococcus aureus, Klebsiella pneumoniae, Escherichia coli, Proteus mirabilis and Bacillus subtilis (NJINGA et al., 2016). The results of one study revealed that β-sitosterol in 1,2-dimethylhydrazine caused an augmentation of enzymatic and non-enzymatic antioxidants in mice, and this compound has been recommended as an effective drug for colorectal cancer (BASKAR et al., 2012). Vitamin β-Tocopherol is essential as an antioxidant and anti-inflammatory (RIZVI et al., 2014).

 

Conclusion

 

Recently, there has been an increasing demand for natural additives instead of synthetic preservatives. As a result, academic research on the antioxidant and antibacterial properties of extracts, essential oils, and fixed oils from various plant species has also increased. In this context, studying the organic fraction of distilled Salvia hispanica seeds (OFSH) may be of great interest. Chia is rich in bioactive compounds, such as fatty acid derivatives, steroids, and tocopherol, which showed high antioxidant, antidiabetic, and antibacterial activities. Our data show that S. hispanica could be an excellent alternative source for therapeutic medicine, especially for the treatment of chronic diseases such as diabetes, coronary heart disease, cancer and fungal infections.

 

Conflicts of interest

 

The author declares no conflicts of interest regarding the work presented here.

 

Authors’ contribution

 

Ibtissem Rahmoune – methodology, data curation, writhing original draft; Samira Karoune – validation, reviewing and ressources; Madani Sarri – wrote the paper; Clara Azzam – review; Somia Saad – methodology, conceptualization, review, validation; Abdelhamid Foughalia – antimicrobial activity; Farid Chebrouk – GC/MS analysis; Hana Abidat – experimental assays; Mohamed Seif Allah Kechebar – review.

 

Disclosure statement

 

This work was financed by ERANET FOSC under the title of Integrated Chia and Oyster Mushroom System for a Sustainable Food Value Chain in Africa (CHIAM), supported by the General Directorate of Scientific Research and Technological Development, Algeria.

 

References

 

ABDEL GHANI, A. E.; AL-SALEEM, M. S. M.; ABDEL-MAGEED, W. M.; ABOUZEID, E. M.; MAHMOUD, M. Y.; ABDALLAH, R. H. UPLC-ESI-MS/MS Profiling and cytotoxic, antioxidant, anti-Inflammatory, antidiabetic, and antiobesity activities of the non-polar fractions of Salvia hispanica L. aerial parts. Plants, v. 12, n. 5, p. 1-2, 2023. https://doi.org/10.3390/plants12051062

ABOU ZEID, E. M.; ABDEL GHANI, A. E.; MAHMOUD, M. Y.; ABDALLAH, R. H. Phytochemical investigation and biological screening of ethyl acetate fraction of Salvia hispanica L. aerial parts. Pharmacognosy Journal, v. 14, n. 1, p. 226-234, 2022. https://doi.org/10.5530/pj.2022.14.28

APAK, R.; GÜÇLÜ, K.; ÖZYÜREK, M.; KARADEMIR, S. E. Novel total antioxidant capacity index for dietary polyphenols and vitamins C and E, using their cupric ion reducing capability in the presence of neocuproine: CUPRAC method. Journal of Agricultural and Food Chemistry, v. 52, n. 26, p. 7970-7981, 2004. https://doi.org/10.1021/jf048741x

BAKRIM, S.; BENKHAIRA, N.; BOURAIS, I.; BENALI, T.; LEE, L. H.; EL OMARI, N.; SHEIKH, R. A.; GOH, K. W.; MING, L. C.; BOUYAHYA, A. Health benefits and pharmacological properties of stigmasterol. Antioxidants, v. 11, n. 10, p. 1-32, 2022. https://doi.org/10.3390/antiox11101912

BANU, S. A.; JOHN, S.; MONICA, S. J.; SARASWATHI, K.; ARUMUGAM, P. Screening of secondary metabolites, bioactive compounds, in vitro antioxidant, antibacterial, antidiabetic and anti-inflammatory activities of Chia seeds (Salvia hispanica). Research Journal of Pharmacy and Technology, v. 12, n. 14, p. 6289-6294, 2021. https://doi.org/10.52711/0974-360X.2021.01088

BASKAR, A. A.; AL NUMAIR, K. S.; GABRIEL PAULRAJ, M.; ALSAIF, M. A.; MUAMAR, M. AL.; IGNACIMUTHU, S. β-sitosterol prevents lipid peroxidation and improves antioxidant status and histoarchitecture in rats with 1,2-dimethylhydrazine-induced colon cancer. Journal of Medicinal Food, v. 15, n. 4, p. 335-343, 2012. https://doi.org/10.1089/jmf.2011.1780

BOUCHOUKH, I.; HAZMOUNE, T.; BOUDELAA, M.; BENSOUICI, C.; ZELLAGUI, A. Anticholinesterase and antioxidant activities of foliar extract from a tropical species: Psidium guajava L. (Myrtaceae) grown in Algeria. Pharmacy and Medical Sciences, v. 32, n. 3, p. 160-167, 2019. https://doi.org/10.2478/cipms-2019-0029

CAHILL, J. P. Ethnobotany of Chia, Salvia hispanica. Economic Botany, v. 57, n. 4, p. 604-618, 2003. https://www.jstor.org/stable/4256743

CHOUIT, H.; TOUAFEK, O.; BRADA, M.; BENSSOUICI, C.; FAUCONNIER, M. L.; EL HATTAB, M. GC-MS analysis and biological activities of Algerian Salvia microphylla essential oils. Journal of the Mexican Chemical Society, v. 65, n. 4, p. 582-601, 2021. https://doi.org/10.29356/JMCS.V65I4.1581

DIB, H.; SELADJI, M.; BENCHEIKH, F. Z.; FARADJI, M.; BENAMMAR, C.; BELARBI, M. Phytochemical screening and antioxidant activity of Salvia hispanica. Journal of Pharmaceutical Research International, v. 33, n. 41, p.167-174, 2021. https://doi.org/10.9734/jpri/2021/v33i41a32314

EFTIMOVÁ, Z.; EFTIMOVÁ, J.; BALÁŽOVÁ, L. Antioxidant activity of Tokaj essence. Potravinarstvo Slovak Journal of Food Sciences, v. 12, n. 1, p. 323-329, 2018. https://doi.org/10.5219/829

ELSHAFIE, H. S.; ALIBERTI, L.; AMATO, M.; DE FEO, V.; CAMELE, I. Chemical composition and antimicrobial activity of Chia (Salvia hispanica L.) essential oil. European Food Research and Technology, v. 244, n. 9, p. 1675-1682, 2018. https://doi.org/10.1007/s00217-018-3080-x

FOLLI, F.; CORRADI, D.; FANTI, P.; DAVALLI, A.; PAEZ, A.; GIACCARI, A.; PEREGO, C.; MUSCOGIURI, G. The role of oxidative stress in the pathogenesis of type 2 Diabetes Mellitus micro-and macrovascular complications: avenues for a mechanistic-based therapeutic approach. Diabetes Reviews, v. 7, n. 5, p. 313-324, 2011. https://doi.org/10.2174/157339911797415585

GAWRON-SKARBEK, A.; GULIGOWSKA, A.; PRYMONT-PRZYMIŃSKA, A.; NOWAK, D.; KOSTKA, T. The anti-inflammatory and antioxidant impact of dietary fatty acids in cardiovascular protection in older adults may be related to vitamin C intake. Antioxidants, v. 12, n. 2, p. 1-12, 2023. https://doi.org/10.3390/antiox12020267

GHAFOOR, K.; AHMED, I. A. M.; ÖZCAN, M. M.; AL-JUHAIMI, F. Y.; BABIKER, E. E.; AZMI, I. U. An evaluation of bioactive compounds, fatty acid composition and oil quality of Chia (Salvia hispanica L.) seed roasted at different temperatures. Food Chemistry, v. 333, n. 5, p. 1-7, 2020. https://doi.org/10.1016/j.foodchem.2020.127531

GONÇALVES, S.; ROMANO, A. Inhibitory properties of phenolic compounds against enzymes linked with human diseases. In: SOTO-HERNANDEZ, M.; PALMA-TENANGO, M.; GARCIA-MATEOS, M. R. (Eds.). Phenolic compounds – biological activity. InTech Open, p. 1-20, 2017. https://doi.org/10.5772/66844

GRANCIERI, M.; MARTINO, H. S. D.; GONZALEZ DE MEJIA, E. Chia Seed (Salvia hispanica L.) as a source of proteins and bioactive peptides with health benefits: a review. Comprehensive Reviews in Food Science and Food Safety, v. 18, n. 2, p. 480-499, 2019. https://doi.org/10.1111/1541-4337.12423

HENRY, G. E.; MOMIN, R. A.; NAIR, M. G.; DEWITT, D. L. Antioxidant and cyclooxygenase activities of fatty acids found in food. Journal of Agricultural and Food Chemistry, v. 50, n. 8, p. 2231-2234, 2002. https://doi.org/10.1021/jf0114381

ISHAK, I.; HUSSAIN, N.; COOREY, R.; GHANI, M. A. Optimization and characterization of Chia seed (Salvia hispanica) oil extraction using supercritical carbon dioxide. Journal of CO₂ Utilization, v. 4, n. 5, p. 1-14, 2021. https://doi.org/10.1016/j.jcou.2020.101430

JAVID, H.; MOEIN, S.; MOEIN, M. An investigationof the inhibitory effects of dichloromethane and methanol extracts of Salvia macilenta, Salvia officinalis, Salvia santolinifola and Salvia mirzayanii on diabetes marker enzymes, an approach for the treatment diabetes. Clinical Phytoscience, v. 8, n. 1, p. 1-8, 2022. https://doi.org/10.1186/s40816-022-00339-y

KATANIĆ STANKOVIĆ, J. S.; SREĆKOVIĆ, N.; MIŠIĆ, D.; GAŠIĆ, U.; IMBIMBO, P.; MONTI, MIHAILOVIĆ, V. Bioactivity, biocompatibility and phytochemical assessment of lilac sage, Salvia verticillata L. (Lamiaceae) – a plant rich in rosmarinic acid. Industrial Crops and Products, v. 143, p. 1-11, 2020. https://doi.org/10.1016/j.indcrop.2019.111932

KOEPKE, J. I.; WOOD, C. S.; TERLECKY, L. J.; WALTON, P. A.; TERLECKY, S. R. Progeric effects of catalase inactivation in human cells. Toxicology and Applied Pharmacology, v. 232, n. 1, p. 99-108, 2008. https://doi.org/10.1016/j.taap.2008.06.004

KUNCHANDY, E.; RAO, M. N. A. Oxygen radical scavenging activity of Curcumin. International Journal of Pharmaceutics, v. 58, n. 3, p. 237-240, 1990. https://doi.org/10.1016/0378-5173(90)90201-E

KUSUMAH, D.; WAKUI, M.; MURAKAMI, M.; XIE, X.; YUKIHITO, K.; MAEDA, I. Linoleic acid, α-linolenic acid, and monolinolenins as antibacterial substances in the heat-processed soybean fermented with Rhizopus oligosporus. Bioscience, Biotechnology and Biochemistry, v. 84, n. 6, p. 1285-1290, 2020. https://doi.org/10.1080/09168451.2020.1731299

LALITHADEVI, B.; MUTHIAH, N. S.; MURTY, K. S. N. Antioxidant activity of conjugated linoleic acid. Asian Journal of Pharmaceutical and Clinical Research, v. 11, n. 11, p. 169-173, 2018. https://doi.org/10.22159/ajpcr.2018.v11i11.27700

LIN, K.Y.; DANIEL, J. R.; WHISTLER, R. L. Structure of chia seed polysaccharide exudate. Carbohydrate Polymers, v. 23, n. 1, p. 1-6, 1994. https://doi.org/10.1016/0144-8617(94)90085-X

LU, X.; YU, H.; MA, Q.; SHEN, S.; DAS, U. N. Linoleic acid suppresses colorectal cancer cell growth by inducing oxidant stress and mitochondrial dysfunction. Lipids in Health and Disease, v. 9, n. 106, p. 1-11, 2010. http://www.lipidworld.com/content/9/1/106

LUCA, S. V.; SKALICKA-WOŹNIAK, K.; MIHAI, C. T.; GRADINARU, A. C.; MANDICI, A.; CIOCARLAN, N.; MIRON, A.; APROTOSOAIE, A. C. Chemical profile and bioactivity evaluation of Salvia species from Eastern Europe. Antioxidants, v. 12, n. 8, p. 1-21, 2023. https://doi.org/10.3390/antiox12081514

MAMACHE, W.; AMIRA, S.; BEN SOUICI, C.; LAOUER, H.; BENCHIKH, F. In vitro antioxidant, anticholinesterases, anti-α-amylase, and anti-α-glucosidase effects of Algerian Salvia aegyptiaca and Salvia verbenaca. Journal of Food Biochemistry, v. 44, n. 11, p. 1-15, 2020. https://doi.org/10.1111/jfbc.13472

MARTÍNEZ-CRUZ, O.; PAREDES-LÓPEZ, O. Phytochemical profile and nutraceutical potential of Chia seeds (Salvia hispanica L.) by ultra-high performance liquid chromatography. Journal of Chromatography A, v. 1346, p. 43-48, 2014. https://doi.org/10.1016/j.chroma.2014.04.007

MOKHTAR, A.; SOUHILA, T.; NACÉRA, B.; AMINA, B.; ALGHONAIM, M. I.; ÖZTÜRK, M.; ALSALAMAH, S. A.; MIARA, M. D.; BOUFAHJA, F.; BENDIF, H. In vitro antibacterial, antioxidant, anticholinesterase, and antidiabetic activities and chemical composition of Salvia balansae. Molecules, v. 28, n. 23, p. 1-26, 2023. https://doi.org/10.3390/molecules28237801

NJINGA, N. S.; SULE, M. I.; PATEH, U. U.; HASSAN, H. S.; ABDULLAHI, S. T.; ACHE, R. N. Isolation and antimicrobial activity of β-Sitosterol-3-O-Glucoside from Lannea Kerstingii Engl. & K. Krause (Anacardiacea). Journal of Health Science and Allied Sciences, v. 6, n. 1, p. 4-8, 2016. https://doi.org/10.1055/s-0040-1708607

ÖKTEN, S.; EKIZ, M.; KOÇYIĞIT, Ü. M.; TUTAR, A.; ÇELIK, İ.; AKKURT, M.; GÖKALP, F.; TASLIMI, P.; GÜLÇIN, İ. Synthesis, characterization, crystal structures, theoretical calculations and biological evaluations of novel substituted tacrine derivatives as cholinesterase and carbonic anhydrase enzymes inhibitors. Journal of Molecular Structure, v. 11, n. 75, p. 906-915, 2019. https://doi.org/10.1016/j.molstruc.2018.08.063

OLIVOS-LUGO, B. L.; VALDIVIA-LÓPEZ, M. Á.; TECANTE, A. Thermal and physicochemical properties and nutritional value of the protein fraction of Mexican Chia seed (Salvia hispanica L.). Food Science and Technology International, v. 16, n. 1, p. 89-96, 2010. https://doi.org/10.1 177/1082013209353087

OYAIZU, M. Studies on products of browning reaction: antioxidative activities of browning reaction prepared from glucosamine, Japan Journal of Nutrition, v. 44, n. 6, p. 307-315, 1986. https://doi.org/10.5264/eiyogakuzashi.44.307

PEREZ, C.; PAULI, M.; BAZERQUE, P. An antibiotic assay by agar well diffusion method. Acta Biologiae et Medicinae Experimentalis, v. 15, p. 113-115, 1990.

POPOVIĆ, B. M.; ŠTAJNER, D.; SLAVKO, K.; SANDRA, B. Antioxidant capacity of cornelian cherry (Cornus mas L.) – Comparison between permanganate reducing antioxidant capacity and other antioxidant methods. Food Chemistry, v. 134, n. 2, p. 734-741, 2012. https://doi.org/10.1016/j.foodchem.2012.02.170

RE, R.; PELLEGRINI, N.; PROTEGGENTE, A.; PANNALA, A.; YANG, M.; RICE-EVANS, C. Original contribution antioxidant activity applying an improved abts radical cation decolonization assay. Free Radical Biology & Medicine. v. 26, n. 10, p. 1231-1237, 1999. https://doi.org/10.1016/s0891-5849(98)00315-3

RIZVI, S.; RAZA, S. T.; AHMED, F.; AHMAD, A.; ABBAS, S.; MAHDI, F. The role of vitamin E in human health and some diseases. Sultan Qaboos University Medical Journal, v. 14, n. 2, p. 157-165, 2014. https://pubmed.ncbi.nlm.nih.gov/24790736/

RODRIGUES, M. J.; PEREIRA, C. A.; OLIVEIRA, M.; NENG, N. R.; NOGUEIRA, J. M. F.; ZENGIN, G.; MAHOMOODALLY, M. F.; CUSTÓDIO, L. Sea rose (Armeria pungens (Link) Hoffmanns. & Link as a potential source of innovative industrial products for anti-ageing applications. Industrial Crops and Products, v. 12, n. 1, p. 250-257, 2018. https://doi.org/10.1016/j.indcrop.2018.05.018

SANDOVAL-OLIVEROS, M. R.; PAREDES-LÓPEZ, O. Isolation and characterization of proteins from Chia seeds (Salvia hispanica L.). Journal of Agricultural and Food Chemistry, v. 61, n. 1, p. 193-201, 2013. https://doi.org/10.1021/jf3034978

SCAPIN, G.; SCHMIDT, M. M.; PRESTES, R. C.; ROSA, C. S. Phenolics compounds, flavonoids and antioxidant activity of Chia seed extracts (Salvia hispanica) obtained by different extraction conditions. International Food Research Journal, v. 23, n. 6, p. 2341-2346, 2016. http://www.ifrj.upm.edu.my/23%20(06)%202016/(5).pdf

SILVA, C.; GARCIA, V. A. S.; ZANETTE, C. M. Chia (Salvia hispanica L.) oil extraction using different organic solvents oil. International Food Research Journal, v. 23, n. 3, p. 998-1004, 2016. http://www.ifrj.upm.edu.my/23%20(03)%202016/(13).pdf

SZYDŁOWSKA-CZERNIAK, A.; DIANOCZKI, C.; RECSEG, K.; KARLOVITS, G.; SZŁYK, E. Determination of antioxidant capacities of vegetable oils by ferric-ion spectrophotometric methods. Talanta, v. 76, n. 4, p. 899-905, 2008. https://doi.org/10.1016/j.talanta.2008.04.055

TARLING, C. A.; WOODS, K.; ZHANG, R.; BRASTIANOS, H. C.; BRAYER, G. D.; ANDERSEN, R. J.; WITHERS, S. G. The search for novel human pancreatic α-amylase inhibitors: high-throughput screening of terrestrial and marine natural product extracts. ChemBioChem, v. 9, n, 3, p. 433-438, 2008. https://doi.org/10.1002/cbic.200700470

TOLENTINO, R. G.; VEGA, L. R.; VEGA, Y.; LEON, S. V.; FONTECHA, J.; RODRÍGUEZ, L. M.; MEDINA, A. E. Fatty acid content in Chia (Salvia hispanica L.) seeds grown in four Mexican states. Revista Cubana de Plantas Medicinales, v. 19, n. 1, p.199-207, 2014. https://www.medigraphic.com/pdfs/revcubplamed/cpm-2014/cpm143h.pdf

TUNÇİL, Y. E.; ÇELİK, Ö. F. Total phenolic contents, antioxidant and antibacterial activities of Chia seeds (Salvia hispanica L.) having different coat color. Akademik Ziraat Dergisi, v. 8, n. 1, p. 113-120, 2019. https://doi.org/10.29278/azd.593853

YOON, S. Y.; AHN, D.; HWANG, J. Y.; KANG, M. J.; CHUNG, S. J. Linoleic acid exerts antidiabetic effects by inhibiting protein tyrosine phosphatases associated with insulin resistance. Journal of Functional Foods, v. 83, n. 9, p. 1-6, 2021. https://doi.org/10.1016/j.jff.2021.104532

ZENGIN, G.; SARIKURKCU, C.; AKTUMSEK, A.; CEYLAN, R.; CEYLAN, O. A comprehensive study on phytochemical characterization of Haplophyllum myrtifolium Boiss. endemic to Turkey and its inhibitory potential against key enzymes involved in Alzheimer, skin diseases and type II diabetes. Industrial Crops and Products, v. 5, n. 3, p. 244-251, 2014. https://doi.org/10.1016/j.indcrop.2013.12.043

ZONG, G.; LIU, G.; WILLETT, W. C.; WANDERS, A. J.; ALSSEMA, M.; ZOCK, P. L.; HU, F. B.; SUN, Q. Associations between linoleic acid intake and incident type 2 diabetes among U.S. men and women. Diabetes Care, v. 42, n. 8, p. 1406-1413, 2019. https://doi.org/10.2337/dc19-0412

 

 

 

Received on August 17, 2024

Returned for adjustments on October 7, 2024

Received with adjustments on October 8, 2024

Accepted on October 15, 2024