Nutraceutical Potential of Seven “Quelites” Harvested in the Northern Highlands of Puebla-México

Abstract

The northern highlands of Puebla harbor several plant foods consumed since pre-Columbian times. Most of the native vegetables from this geographical region are still uncharacterized at the nutraceutical level. This investigation was focused on the elucidation of chemical and biochemical properties of the edible organs from Rhamnus pompana, Solanum nigrescens, Sechium edule, Yucca aloifolia, Piper auritum, Amaranthus hybridus and Rumex obtusifolius, which are used as traditional foods in this region. The content of basic nutrients (including vitamins of the B complex), nutraceuticals (carotenoids, flavonoids, and fatty acids), phenolics (including antioxidant capacity), and inhibitory properties of these plants on specific enzymes linked to carbohydrate and lipid metabolism were determined. In addition, extracts of these plants were evaluated against pathogenic microorganisms of the gastrointestinal tract. Our results demonstrated statistically significant divergences (p < 0.05) in the content protein (12.5 g/100 g), fiber (9.8 g/100 g), fat (81.2 mg/100 g), beta-carotene (2.48 mg/100 g), beta-cryptoxanthin (1.24 mg/100 g), thiamine (159.6 µg/100 g), riboflavin (106.4 µg/100 g), and vitamin B₆ between Y. aloifolia and other plants studied. All plants showed low amounts of reducing sugars (<150 mg/100 g) and sodium (<70 mg/100 g), but the berries of R. pompana exhibited substantial differences (p < 0.05) in the levels of vitamin C (14.6 mg/100 mg). The edible leaves of R. obtusifolius were particularly rich in quercetin (4.38 mg/100 g), kaempferol (2.38 mg/100 g) and lutein (2.52 mg/100 g), whereas the aerial parts of S. edule contained high concentrations of folate (50.6 µg/100 g). All plants showed variable phenolic content and antioxidant capacity; however, Y. aloifolia had the highest values (23 GAE/mg/g and 440 TEAC/µM/g). The ethanolic extracts from Y. aloifolia efficiently inhibited pancreatic lipase (IC₅₀, 43.76 µg/mL) and alpha-glucosidase (IC₅₀, 60.04 µg/mL), whereas those from S. edule inhibited alpha-glucosidase (IC₅₀, 47.4 µg/mL) and HMG-CoA reductase (IC₅₀, 33.6 µg/mL). Only the ethanolic extracts of R. pompana (IC₅₀, 35.36 µg/mL) and Y. aloifolia (IC₅₀, 53.28 µg/mL) inhibited ornithine decarboxylase. All plant extracts exerted moderated antimicrobial activity in at least one species associated with the gastrointestinal tract.

1. Introduction

The northern highlands of Puebla-México (NHP) comprise 68 municipalities with Indigenous roots. This geographical region is represented by an extraordinary mixture of people with Totonac, Nahua, Otomi, and Tepehua ancestry [1]. Because of the high precipitation rate and permeable soils, this zone stores a great diversity of plants that have an evident impact on the socioeconomic conditions of the Indigenous people [2]. Plantations of coffee, avocados, maize, beans, and manzano peppers are the leading sustainable crops in these places [3]. However, 319 plant species belonging to 238 genera and 99 families were recorded as a source of income [4]. Among these plants, green tissues, or immature fruits from 40 species are frequently consumed in the NHP as an inheritance of pre-Columbian knowledge passed down from one generation to another [2].

The term “quelite” derives from the Nahuatl word “quilitl” meaning young edible vegetable [1,2,3,4]. Nonetheless, in some places, the concept is extended to immature dehiscent fruits, such as those of the Phaseolus genus (ejotes), flowers of Yucca genus, berries of wild Solanaceae, and the seeds of the Ceiba genus [1,2,3,4]. On the other hand, these vegetables are not known enough in other geographical regions, and their consumption has unfortunately decreased in the community itself because of the generational transition [5]. One way to preserve these millenary crops involves the elucidation of their nutritional and nutraceutical benefits, followed by proper promotion in the community and general public through social networks. Such actions may help to create a consciousness on their benefits to human health [6]. Furthermore, the involvement of municipalities and complementary policies should be considered for establishing new social strategies to motivate and facilitate the preservation of these foods [6].

As a general overview, the nutritional and nutraceutical contents of the quelites from the NHP have been poorly explored. This fact contrasts with the investigation carried out on medicinal plants from the same geographical region, which has received much more attention. Most studies performed in the quelites from the NHP and other zones of this province, simply present the name of the vegetables and how they are typically cooked [1,2]. Considering that some of these vegetables are eaten raw, their chemical profiling and possible nutraceutical activity must be researched. In this context, the chemical and biological properties of these foods may reveal potential applications for controlling public health problems in Mexico, such as obesity, type 2 diabetes mellitus (DM2), and cancer. These foods can also be visualized as potential sources of chemopreventive molecules.

Rates of obesity and overweight persons in Mexico continue to increase; their prevalence is indicated by comprising 42% proportion of women and people over 50 years old [7]. Similarly, DM2 increased from 7.3 to 9.5% in 2020, and this tendency is well associated with a highly diabetogenic environment in which hypercaloric diets are included [7]. The observed trends strongly suggest that the prevalence of DM2 will continue to rise in the next couple of years if preventive measures are not correctly applied [7]. In Mexico, cancer is considered the third most common cause of death, including prostate, colorectal, testicular, lung, and gastric cancers as the most prevalent in men, whereas breast, thyroid, cervical, uterine corpus, and colorectal cancers are the most common in women [8]. The replacement of fruits and vegetables rich in fiber by ultra-processed foods has increased the exposure to various chemicals acting as carcinogens [8].

In addition, the low consumption of antioxidants like vitamin A, C, flavonoids, minerals, and essential amino acids is also linked to the emergence of these types of cancer [8]. Then, the consumption of green vegetables rich in nutraceuticals (as in the case of quelites) seems to be safe and would be an efficient choice to prevent these disorders. The term nutraceutical is applied to molecules or elements contained in foods that can improve human health and well-being. These chemicals may include isoprenoid derivatives, phenolic compounds, fatty acids, lipids, non-proteinogenic amino acids, fiber, non-caloric carbohydrates, and chemical elements that exert therapeutic properties, such as antioxidant, anti-inflammatory, antimicrobial, antineoplastic action, and as regulators of lipid and carbohydrate metabolisms [9,10].

Previous studies suggest that at least three quelites from the NHP possess nutraceutical properties [11,12,13]. Interestingly, their consumption could ameliorate metabolic illnesses derived from an imbalanced diet [11,12,13]. The edible stalks of Begonia nelumbiifolia (named as “xocoyoles” in the NHP), have substantial amounts of vitamin C, carotenoids, and flavonoids with biological activity. In addition, extracts of this plant inhibited the action of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase) and alpha-glucosidase under in vitro conditions [11]. As is known, these enzymes are blank of anti-hypercholesterolemic and anti-hyperglycemic pharmaceuticals. Interestingly, the volatile fraction of the leaves from Peperomia maculosa (known as “Oreja de Leon” in the NHP) and its main volatile decanal inhibited the enzymatic activity of pancreatic lipase in vitro. Considering that the plant is consumed raw, in vivo studies using the ICR mouse model confirmed its ability to reduce plasma triglycerides in the postprandial stage [12]. Another quelite, Zanthoxylum limoncello (known as “Cilantrillo” in the NHP) produces 2-undecanone, 2-undecenal and 2-dodecenal able to inhibit the activity of ornithine decarboxylase. This enzyme is visualized as a therapeutic target to block the proliferation of several types of cancer [13]. The volatile fraction of this plant also produced an evident growth inhibition on three strains of Helicobacter pylori, which is associated with gastric cancer [13].

On the basis of the latter arguments, this work was focused on the nutritional and nutraceutical screening of seven quelites frequently consumed in the NHP.

2. Materials and Methods

2.1. Chemicals

Solvents for chromatography were from J.T. Baker®. Rezasurin, alpha-glucosidase (AG), alpha-amylase (AA), 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase; catalytic domain kit), a standard mixture of elements (zinc, calcium, copper, sodium, iron, and potassium), quercetin, kaempferol, lutein, beta-cryptoxanthin, beta-carotene, alpha-carotene, linoleic acid, and linolenic acid were obtained from Sigma-Aldrich Co. (St. Louis, MO, USA). Pancreatic lipase (PL) was from Affymetrix/USB and human ornithine decarboxylase (ODC) was obtained from MyBiosource (MBS967514).

2.2. Plant Material

Aerial parts from seven plants tagged as quelites in the NHP were collected in the municipality of Yaonáhuac, Puebla, Mexico (19°56’55’’N 97°26’26’’W; 1997 masl) during the year 2021 (

Table 1. Edible quelites consumed in the northern highlands of Puebla-México.

Quelite	Scientific Name	Edible Organ	Harvesting Stage *	Collection Date
Tepeilite	Rhamnus pompana	Berries	Stage 8 of BBCH-Scale	August 2021
Hoja santa	Piper auritum	Fresh leaves	Stage 5 of BBCH-Scale	February 2021
Hierba mora	Solanum nigrescens	Berries	Stage 8 of BBCH-Scale	May 2021
Quintoniles	Amarathus hybridus	Fresh leaves	Stage 5 of BBCH-Scale	February 2021
Guias de erizo	Sechium edule	Terminal stalks with tendrils	Anthesis	June 2021
Lengua de vaca	Rumex obstusifolia	Fresh leaves	Stage 5 of BBCH-Scale	October 2021
Flor de Izote	Yucca aloifolia	Young inflorescences	Anthesis	November 2021

* Development in accordance with BBCH scale for berries and leafy vegetables [14].

). Yaonáhuac-Puebla has an annual temperature range between 12 °C and 18 °C, the average annual rainfall is 2000 to 2500 mm, presenting a humid temperate climate with rain throughout the year. In Yaonáhuac, Puebla, andosol is predominant and occupies more than 75% of the municipality. The identity of Rhamnus pompana M.C. Johnst. & L.A. Johnst., Piper auritum Kunth, Solanum nigrescens M. Martens & Galeotti, Sechium edule (Jacq.) Sw., Amarathus hybridus L., and Rumex obtusifolius L., was corroborated at the herbarium of the FCME-UNAM by Ramiro Cruz Durán where the vouchers 178,871, 152,371, 164,616, 179,056, 179,058, and 179,404, were respectively deposited. The identity of Yucca aloifolia L. was determined by Fermín Tavares at the medicinal plant collection of Northern Highlands Association–Mexico where the reference voucher LY-770014 was stored. More data on the collection of these plants are listed in Table 1.

The edible organs of these plants were used for further analysis. In this context, 5 kg of young berries from Rhamnus pompana and Solanum nigrescens were collected and processed for further tests, whereas 4 kg of young terminal stalks with tendrils were obtained from Sechium edule. Lastly, 5 kg of flowers from Yucca ailofolia, as well as 5 kg of fresh leaves from Piper auritum, Amarathus hybridus and Rumex obtusifolius were processed and collected and processed for further tests (Figure 1).

2.3. Proximate Analysis and Micronutrient Content

The contents of protein (AOAC 920.23), fiber (AOAC 962.09), fat (AOAC 920.39), reducing sugars (AOAC 945.66), vitamin C (AOAC 967.21/90), folate (AOAC 944.12), thiamine (AOAC 942.23), total vitamin B₆ (AOAC 961.15), and riboflavin (AOAC 970.65) were estimated in accordance with the official methods of the AOAC International [15]. Mineral content was calculated on the basis of atomic absorption spectroscopy using an A3F flame atomic absorption spectrometer equipped with a flame atomizer (PERSEE) using aqueous extracts containing 50 g of plant material. The wavelengths, slits, and lamp conditions for the determination of zinc, calcium, copper, sodium, iron, and potassium were the same as those described in a previous investigation [12]. Calibration curves were set using a standard mixture of the six elements. The results were expressed in mg 100 g^–1 fresh weight. Each parameter was assayed ten times (n = 10).

2.4. Nutraceutical Content and Antioxidant Capacity

The contents of flavonoids (quercetin, kaempferol), carotenoids (lutein, beta-cryptoxanthin, alpha- and beta-carotene) were determined by RP-HPLC starting from hydroalcoholic extracts (80% EtOH) using the same protocols of extraction, equipment, chromatographic conditions, and authentic standards reported in previous investigations [11,12]. For carotenoids, 50 g of fresh material were ground in a mortar and extracted with 200 mL of cold hexane:acetone (1:1 v/v) added with 0.5% butylated hydroxytoluene, with vigorous shaking at 4 °C for 12 h in darkness. Extracts were filtered and reduced to 50 mL for further saponification, according to Villa-Ruano et al. [11]. Then, 50 mL of distilled water was added along with 50 mL of 10% NaCl and the two layers of the mixture were separated in a sedimentation funnel after being vigorously shaken. The upper layer containing carotenoids was collected and reduced to dryness and resuspended in n-hexane for further HPLC analysis. For flavonoid extraction, 100 g of fresh material were ground and extracted with 80% MeOH for 24 h with gentle shaking at room temperature. The extracts were filtered and reduced to a green gum, which was subsequently separated by column chromatography (20 X 1.5 cm) using 60.0 g of silica gel (Merck 7744). Elution was made with methanol/ethyl acetate (20, 40, 60, 80%) with fractions of 20 mL. Fractions 4–6 were mixed and reduced to a dried form using a rotary evaporator and resuspended in pure methanol for HPLC analysis. The run conditions were those reported by Villa-Ruano et al. [11,12]. Total phenolic content and antioxidant capacity were estimated in accordance with Awika et al. [16]. Briefly, 1 g of material was ground and extracted with aqueous acetone (70%), and the mixture was shaken for 2 h at low speed and then stored at −20 °C in the dark overnight to allow for maximum diffusion of phenolics from the cellular matrix. Samples were equilibrated to room temperature and centrifuged at 3000× g for 10 min. The phenol content of aqueous acetone extracts were quantified by the Folin−Ciocalteu method [16]. Antioxidant capacity assays were conducted using an MR9600-T-SmartReader (Accuris ^TM) using pH 7.0 phosphate buffer at 37 °C. A peroxyl radical was generated using AAPH, and fluorescein was used as the substrate. Fluorescence conditions were 485 nm excitation and 520 nm emission [16]. The results were expressed as gallic acid equivalents (GAE mg g⁻¹) and Trolox equivalent antioxidant capacity (TEAC µM g⁻¹), respectively. The presence of linoleic and linolenic acid was determined by GC-MS using fatty acid methyl esters prepared from total lipids and authentic standards. For this particular case, one gram of fresh tissue was extracted in n-hexane (20 mL), followed by sonication (VEVOR Ultrasonic Bath) for 30 min. The extract was concentrated until dryness through N₂ stream and resuspended in 500 µL of the same solvent. The analyses were done in an Agilent 5977B GC/MSD chromatograph (Santa Clara, CA, USA) equipped with an HP5-ms capillary column (Palo Alto, CA, USA) using the running conditions reported by Coyotl-Pérez et al. [17]. All assays were replicated fifteen times (n = 15).

2.5. Inhibitory Activity on Key Enzymes with Therapeutic Potential

The enzymatic reactions were performed with AG, AA, HMG-CoA reductase, PL and ODC (see Section 2.1). The assays were performed with ethanolic extracts (80% EtOH) previously treated with lead acetate (10 mM) to remove chlorophylls for further lyophilization until powder. The extracts were resuspended in the same solvent to achieve a final concentration of 100 mg mL⁻¹. All assays were done using a dose-response curve of 10–300 µg mL⁻¹. AG tests were done with 2.5 U/mL enzyme and p-nitrophenyl-α-D-glucopyranoside (1 mM) as a substrate. For AA (2.5 U/mL) assays, the substrate was 4-Nitrophenyl-4,6-Ethylidene-a-D-Maltoheptaoside (ethylidene-pNP-G7; 1 mM). Both tests were followed at 405 nm, as previously described [18]. The inhibitory examination on PL was performed using 3 U/mL enzyme with 2 mM triolein by the turbidimetric method, which measures the hydrolysis of fatty acids from triolein at 340 nm [19]. The assays performed on human HMG-CoA reductase (1.5 U/mL) were based on the decrease of absorbance at 340 nm, which represents the oxidation of NADPH by the catalytic subunit of the enzyme in the presence of the substrate HMG-CoA (1 mM) [11]. Finally, the possible inhibitory effect of the seven extracts was investigated on the human ODC. The conversion of ornithine (2 mM) into putrescine by ODC (1 U/mL) was done in accordance with the protocol of hydrophobic withdrawal of yellow-colored TNP-Putrescine-TNP [13]. The IC₅₀ of each plant extract was calculated by linear regression considering the specific activity of each enzyme under our experimental conditions. Thus, the optimal specific activity (100% activity) for AG, AA, HMG-CoA reductase, PL and ODC was 0.034, 0.063, 0.021, 0.085, and 0.046 mM min⁻¹, respectively. All assays were replicated 25 times (n = 25) for each enzyme.

2.6. In Vitro Antimicrobial Activity on Selected Pathogens

The antimicrobial activity of aqueous and ethanolic extracts (previously lyophilized) was determined by the broth microdilution method using resazurin as an indicator of cell viability [17]. Dose–response curves of 10–550 µg mL⁻¹ were done to obtain the minimum inhibitory concentration (MIC) [17]. The assayed species and strains were Escherichia coli ATCC 25922, E. coli ATCC 11303, Staphylococcus aureus ATCC 25923, Enterococcus faecalis ATCC 29212, Salmonella typhi ATCC 6539, and Candida albicans ATCC 90028. Each point of the curve was assayed 25 times.

2.7. Statistical Analysis

Analysis of variance coupled with Tukey’s test was carried out to establish statistically significant differences among the levels of nutrients and nutraceuticals from the seven edible plants using GraphPad Prism 8.0 software. The IC₅₀ of each plant extract on assayed enzymes was calculated by linear regression using the same software. The optimal specific activity of each enzyme was used to calculate IC₅₀.

3. Results and Discussion

3.1. Nutritional Content

Proximate analysis revealed that the flowers of Y. aloifolia had a higher content of protein (12.5 g 100 g⁻¹), fiber (9.8 g 100 g⁻¹), and fat (81.2 mg 100 g⁻¹) than other plants studied (Figure 2a–c). Remarkably, the levels of protein from the leaves of Y. aloifolia were seven folds higher than those reported for B. nelumbiifolia and P. maculosa, which are edible plants frequently consumed in the NHP [11,12]. Besides, the levels of proteins were higher than those found in six wild vegetables regularly consumed in the Mediterranean region [20]. The berries of R. pompana had acceptable amounts of protein; however, its reducing sugar content was higher (368.2 mg 100 g⁻¹) than that of other vegetables (Figure 2d).

Interestingly, the amounts of reducing sugars and protein from the berries of S. nigrescens were comparable to those of the other edible greens studied (Figure 2d). In any case, the low levels of reducing sugars for the studied berries seem to be safe for people with DM2 and their obesogenic properties are probably negligible under moderate consumption [11,12]. In addition, the presence of specific antioxidants, including anthocyanins, needs to be considered for these fruits [11,12]. Preliminary data on the pigment HPLC-profiling of R. pompana and S. nigrescens suggest the presence of specific anthocyanins with known nutraceutical activity (data no shown). Aside from Y. aloifolia, our results also revealed that S. edule, R. obtusifolius, and R. pompana had better levels of fiber (>4 g 100 g⁻¹) than that reported for B. nelumbiifolia and P. maculosa [11,12]. This parameter could be associated with favorable gastrointestinal benefits, such as the induction of peristalsis and the protection of gut probiotics [21].

It is known that the leaves of Rumex spp. may also accumulate sennosides (anthraquinones) that promote peristalsis in the gut of mammals [22]. Then, further chemical screening should be performed in R. obtusifolius to determine anthraquinones with biological activity. The fat content of R. obtusifolius and that from the berries of R. pomapana showed no statistically significant differences (Figure 2c). A similar trend was observed for the aerial parts of S. edule and A. hybridus (Figure 2c). The amount of fat in these plants was five folds lower than that reported for the edible stalks of B. nelumbiifolia and the leaves of P. maculosa, and it was comparable to that of Bryonia dioica (red bryony), Humulus lupulus (common hop), and Tamus communis (black bryony) [11,12,20]. Although the levels of protein, fiber, fat, and reducing sugars of the studied plants were adequate for human consumption, the effect of cooking on these parameters should be further studied.

3.2. Selected Vitamin Content

According to our determinations, R. pompana had higher levels of vitamin C (14.6 mg 100 g⁻¹) than other vegetables studied, and these levels were almost twice those observed for blueberries (Figure 3a) [23]. Nevertheless, the berries from S. nigrescens were also rich in vitamin C (Figure 3a). To the best of our knowledge, the presence of ascorbic acid in both berries is reported for the first time. From the edible leaves included in this study, those from R. obtusifolius had larger amounts of vitamin C than the others (Figure 3a). Interestingly, the levels of vitamin C in P. auritum were not observed (Figure 3a). Although these results suggest that this source is poor in ascorbic acid, complementary investigation on the seasonal and geographical analysis of this plant species should be envisioned.

Y. aloifolia had the highest amount of thiamine (159.6 µg 100 g⁻¹), which was similar to that observed in A. hybridus (149.8 µg 100 g⁻¹) (Figure 3b). These levels of vitamin B₁ were related to those found in edible insects such as Zophobas morio and Tenebrio molitor as well as in common meat sources such as duck carcass and turkey breast [24]. No levels of thiamine were found for Solanum nigrescens, whereas those of S. edule, R. obtusifolius, P. auritum, and R. pompana, showed statistically significant differences (Figure 3b). Y. aloifolia also had the highest levels riboflavin (106.4 µg 100 g⁻¹) followed by R. pompana (84.4 µg 100 g⁻¹) (Figure 3c). The levels of this vitamin were comparable to those found in chicken breast, turkey breast, and duck carcass [24]. The lowest levels of vitamin B₂ were detected in S. edule (18.0 µg 100g⁻¹) (Figure 3c).

Vitamin B₆ was better accumulated in Y. aloifolia (91.2 µg 100 g⁻¹) and R. obtusifolius (29.0 µg 100 g⁻¹) than in other plants studied (Figure 3d). These levels were comparable to those found in purslane, cucumber, crisp lettuce, tomato, cabbage, peas, and green beans [25]. Considering that vegetarians obtain most of their vitamin B₆ from legumes, cereals, and fruits, these foods may complement their requirements for this vitamin [25]. Remarkably, the edible organs from S. nigrescens, S. edule, and A. hybridus had no detectable levels of this vitamin (Figure 3d).

The levels of folate in the seven quelites studied revealed that the edible stalks of S. edule (50.6 µg 100 g⁻¹) were the best source of this vitamin, followed by the edible leaves of A. hybridus (34.4 µg 100 g⁻¹) and R. obtusifolius (26.4 µg 100 g⁻¹) (Figure 3e). The amounts of folate in R. pompana and Y. aloifolia had no statistically significant differences, whereas no detectable levels of this micronutrient were determined in the samples of S. nigrescens and P. auritum (Figure 3e). Although the levels of folate were low, these were comparable to those found is specific USDA and AVRDC spinach accessions such as PI604792 and TOT7339-B [26].

3.3. Mineral Content

The results of mineral analysis showed that S. nigrescens and P. auritum had the higher concentration of potassium (351.4 and 349.4 mg 100 g⁻¹, respectively), whereas S. nigrescens (116.2 mg 100g⁻¹) had the lowest levels of this mineral (Figure 4a). These levels were comparable to those found in common cultivars of Beta vulgaris and Lactuca sativa [27]. However, the concentrations of this mineral were lower than in the edible leaves of P. maculosa, a quelite frequently consumed in the NHP [12]. Since potassium helps in the regulation of fluid balance, muscle contraction, and neuronal signaling, the discovery of alternative sources in rural areas should be relevant to the nutrient intake of the local population. According to our results, all plants contained acceptable amounts of calcium, but Y. aloifolia (552.2 mg 100 g⁻¹) and the berries from S. nigrescens (>400 mg 100 g⁻¹) had the highest concentration of this mineral (Figure 4b). These amounts were pretty higher than those reported for P. maculosa, Plantago major, Spinacia oleracea, Beta vulgaris and Lactuca sativa [27]. From the analyzed edible organs, the berries of R. pompana showed the lowest values of calcium (55.0 mg 100 g⁻¹) (Figure 4b).

As for the levels of calcium, the levels of iron were also abundant in Y. aloifolia (101.2 mg 100 g⁻¹) (Figure 4c). Interestingly, the levels of this element were over 70 mg 100 g⁻¹ in the berries of S. nigrescens and the fresh leaves of A. hybridus (Figure 4c). These levels were higher than those reported for P. maculosa, Plantago major, Spinacia oleracea, Beta vulgaris, and Lactuca sativa [27]. Even, the low levels of iron found in S. edule were higher than those reported for the same vegetables [27]. As a general finding, all plants studied contained low levels of sodium (<50 mg 100 g⁻¹), which could be safe for people with hypertension (Figure 4d). Nevertheless, statically significant differences were observed among sodium levels of A. hybridus, Y. aloifolia, and S. nigrescens (Figure 4d). The observed concentrations were lower than those reported for Spinacia oleracea, Beta vulgaris and Lactuca sativa [27]. Zinc levels in the edible organs of R. obtusifolius, S. ngrescens, A. hybridus, P. auritum, R. pompana, and Y. aloifolia were below 20 mg 100 g⁻¹, whereas those of S. edule were 32.4 mg 100 g⁻¹ (Figure 4e). The levels of zinc in these plants were substantially higher than those found in S. oleracea, B. vulgaris, L. sativa, and P. maculosa [12,27]. Coincidently, the concentration of magnesium in S. edule was the highest (40.6 mg 100 g⁻¹) among studied quelites (Figure 4f). These results clearly stated that the seven quelites studied may provide significant amounts of minerals to complete the daily requirements for maintaining optimal metabolic performance. Nevertheless, since the accumulation of minerals depends on abiotic factors such as soil composition and climate, further studies are required to determine possible influence of seasons and geographical location on the mineral concentration in plant organs [11,12,27].

3.4. Molecules with Nutraceutical Activity

Rumex obtusifolius, A. hybridus, and Y. aloifolia accumulated the highest amount of quercetin (3–4 mg 100 g⁻¹), whereas S. nigrescens stored the lowest levels of this flavonoid (0.82 mg 100 g⁻¹) (Figure 5a). The concentration of quercetin in these plants was comparable to that contained in Daucus carota (carrot), Raphanus sativus (white radish), and Amaranthus gangeticus (red spinach) [28]. On the other hand, R. obtusifolius (1.28 mg 100 g⁻¹) had the highest level of kaempferol (~2 mg 100 g⁻¹), followed by S. edule, A. hybridus, and Y. aloifolia, which contained similar levels of this flavonol (~1 mg 100 g⁻¹) (Figure 5b). The berries of S. nigrescens (0.42 mg 100 g⁻¹) showed the lowest concentration of kaempferol.

As is known, quercetin and kaempferol exert antioxidant and free radical scavenging activity in foods and possess significant vitamin C sparing activity [28]. Their nutraceutical properties are also extended to anti-inflammatory, cardioprotective and antihypertensive benefits under in vivo conditions [29]. Then, the presence of these flavonoids in foods is highly desirable. Coincidently, lutein was also abundant in the edible leaves of R. obtusifolius (2.52 mg 100 g⁻¹) and the inflorescences of Y. aloifolia (2.48 mg 100 g⁻¹), whereas no levels of this carotenoid were detected in the berries of R. pompana (Figure 5c). Such levels were higher than those reported in B. nelumbiifolia but lower than those reported in P. maculosa [11,12]. Interestingly, the edible leaves of R. obtusifolius contained the highest levels of β-cryptoxanthin (2.12 mg 100 g⁻¹), followed by the berries of S. nigrescens and the flowers of Y. aloifolia (Figure 5d). A. hybridus and P. auritum presented low amounts of this carotenoid and no detectable levels were recorded for S. edule and R. pompana (Figure 5d). The levels of β-cryptoxanthin in the studied plants was higher than those reported in B. nelumbiifolia but lower than those reported in P. maculosa [11,12]. Alpha-carotene was abundant in Y. aloifolia (0.5 mg 100 g⁻¹), while no detectable levels were found in other plants studied (Figure 5e). The levels of α-carotene were higher than those reported in B. nelumbiifolia but lower than those reported in P. maculosa [11,12]. It has been demonstrated that α-carotene supplementation has a positive effect on the prevention of cancer and cardiovascular diseases under in vivo conditions [30]. The content of β-carotene was evidently higher in Y. aloifolia (2.42 mg 100g⁻¹) than in other plants analyzed (Figure 5f). Although the levels of β-carotene were low in all plants studied, those from R. obtusifolius (0.44 mg 100 g⁻¹) and S. nigrescens (0.26 mg 100 g⁻¹) showed statically significant differences in comparison with other plants (Figure 5f). As is known, the western diet is based on the consumption of β-carotene, α-carotene, and β-cryptoxanthin, which are major provitamin A carotenoids that contribute to vitamin A supply [31]. Vitamin A is essential for the promotion of growth and visual function. It is also known that fruits and vegetables provide around 70% of the vitamin A intake in Third World countries [31]. On the basis of these facts, the elucidation of new sources of edible plants rich in carotenoids is required to improve the local “eat-well plate”.

Linoleic and linolenic acids are the two first (parent) members of ω-6 (n-6) and ω-3 (n-3) fatty acid families, respectively. These fatty acids are considered essential nutrients and they must be supplied by the diet since humans do not have any biochemical pathway to biosynthesize them [32].

Omega-3 and -6 fatty acids have anti-inflammatory, antithrombotic, anti-arrhythmic, hypolipidemic and vasodilatory properties proved in animal models [33]. Then, their regular consumption may avoid cardiovascular diseases. The analysis of fatty acids showed that the inflorescences of Y. aloifolia are a rich source of linoleic acid (54.4 mg 100 g⁻¹) and linolenic acid (51.2 mg 100 g⁻¹) (Figure 6). Interestingly small amounts of these fatty acids were observed in other plants except for S. edule, which did not produce detectable amounts of linoleic acid. The levels of these fatty acids were close to 20 mg 100 g⁻¹, which were higher than those reported for lettuce, broccoli, red cabbage, and soja sprouts [33]. Although the levels of these substances could contribute to the dairy intake of healthy fat, the effect of cooking should be further studied in these edible plant organs.

3.5. Antioxidant Potential

It is well known that all plant extracts possess antioxidant activity; however, this feature should be complemented with the identification of specific compounds associated with this activity [34]. As stated in the previous section, all studied plants contained common nutraceuticals such as minerals, quercetin, kamepferol, carotenoids, fatty acids, as well as other unidentified substances which synergistically work to enhance antioxidant activity. The total phenolic content of the studied plants, revealed that the edible leaves of Y. aloifolia accumulate a substantial amount of phenolics (GAE 23.82 mg/g), which could be correlated with the high antioxidant capacity of this plant (TEAC 440.24 µM g⁻¹) (Figure 7a,b).

Surprisingly, total phenol content was not directly proportional to the antioxidant capacity for all plants studied. In this context, the phenol content of R. obtusifolius was around 12 GAE (mg g⁻¹), which was lower than that of A. hybridus, but the antioxidant capacity of R. obtusifolius was higher than that of A. hybridus (Figure 7a,b). A similar trend was observed for S. nigrescens, S. edule, and R. pompana where no clear correlation was perceived between total phenolic content and antioxidant capacity. Remarkably, the antioxidant capacity of the seven quelites described in this investigation was higher than that previously observed for common cultivars of spinach, red radish, carrot, cauliflower, and broccoli [34]. The role of polyphenols as free radical stabilizers and inhibitors of specific enzymes linked to the metabolisms of carbohydrates and lipids is already known [35]. Therefore, basic knowledge of the total phenol content and the determination of specific phenols may help to correlate the possible nutraceutical effects of the studied plants in forthcoming in vitro or in vivo studies. Since the bioavailability of these substances is considerable modified in the gastrointestinal tract, the evaluation of these substances in murine model, as well as the use of materials to avoid their degradation, should be addressed [36]. Nevertheless, it should be considered that the digestion of extracts and intact foods varies since the cell walls of non-processed plant foods protect inner metabolites from physicochemical factors of the gastrointestinal tract and promote their slow release to the medium [37].

3.6. Inhibitory Activity on Key Enzymes

PL was inhibited by the ethanolic extracts from S. nigrescens, R. obtusifolius, R. pompana, Y. aloifolia but mainly by the extract of R. obtusifolius (IC₅₀, 38.36 µg mL⁻¹) (Figure 8a). This extract was fivefold more effective than that of S. edule (IC₅₀, 213.6 µg mL⁻¹). Interestingly, the ethanolic extract from Y. aloifolia had a similar inhibitory potency than that of R. obtusifolius (IC₅₀, 43.76 µg mL⁻¹). PL is visualized as the best target for controlling obesity since its inhibition by different molecules seems to have less side effects than those observed by commercial drugs such as sibutramine [19]. Currently, there is enough evidence that several molecules from plants exert inhibitory properties on this enzyme [12,19]. No inhibitory activity on PL was exerted by the ethanol extracts of A. hybridus and P. auritum (Figure 8a).

The extracts of Y. aloifolia caused an evident inhibition of both AG (IC₅₀, 60.04 µg mL⁻¹) and AA (IC₅₀, 79.62 µg mL⁻¹), but the most effective extract on AA was that of S. edule (IC₅₀, 47.4 µg mL⁻¹) (Figure 8b,c). The same extract and that of R. obtusifolius had acceptable inhibitory activity on AG, whereas the extracts of R. obtusifolius, A. hybridus and P. auritum were effective against AA (Figure 8b,c). The ethanolic extract from the berries of S. nigrescens has no effect on both enzymes. The IC₅₀ of these extracts was comparable to that reported for the essential oil from D. foliolosa on AG and AA [17]. The function of AG and AA is to hydrolyze assimilable sugars, which results in a critical increase in the postprandial glucose levels [38]. Then, the inhibition of these two enzymes can contribute to controlling postprandial hyperglycemia, and reduce the risk of developing diabetes [38]. Glycosidic bonds α-D-(1,4) are broken by AA to produce oligosaccharides, which are further converted into glucose by AG. Thus, the natural inhibitors of these enzymes can delay the increase in blood glucose after consuming a carbohydrate-rich food diet [38].

In contrast, the ethanolic extract from S. edule showed the best inhibitory effects on HMG-CoA reductase (IC₅₀, 33.6 µg mL⁻¹), whereas that of R. pompana was less effective (Figure 8d). The anti-HMG CoA reductase from S. edule was better than that observed for B. nelumbiifolia, which is a common quelite consumed in the geographical region [11]. No activity was observed for R. obtusifolius and P. auritum on HMG-CoA reductase (Figure 8d). The potential of medicinal or edible plants for controlling hypercholesterolemia is still largely unexplored [39]. Hopefully, further investigation into edible plants may reveal safer and more effective antihypercholesterolemic drugs [35].

ODC is a promising target for cancer research [40]. It is known that some plant flavonoids and fatty acid derivatives from common fruits can inhibit the activity of this enzyme [13,40]. Nevertheless, there has been little exploration of natural molecules as natural inhibitors of ODC. According to our results, the ethanolic extract from R. pompana produced a strong inhibition on this enzyme (IC₅₀, 35.36 µg mL⁻¹), followed by that of Y. aloifolia (IC₅₀, 53.28 µg mL⁻¹) and S. nigrescens (IC₅₀, 180.22 µg mL⁻¹). S. edule, R. obtusifolius, A. hybridus, and P. auritum had no effect on this enzyme. Further chromatographic approaches performed with the ethanolic extracts from R. pompana and Y. aloifolia should help to identify their bioactive anti-ODC compounds.

3.7. Antimicrobial Activity

The antimicrobial strength of aqueous and ethanolic extracts from the edible organs of the seven quelites studied are summarized in

Table 2. Inhibitory activity of aqueous and hydroalcoholic extracts from seven edible plants grown in the northern highlands of Puebla.

	S. nigrescen		S. edule		R. obtusifolius		A. hybridus		P. auritum		R. pompana		Y. aloifolia
Strain	AE	EE	AE	EE	AE	EE	AE	EE	AE	EE	AE	EE	AE	EE
Escherichia coli ATCC 25922	>500	>500	457.2	361.5	472.8	341.7	497.2	398.5	>500	>500	>500	346.9	376.2	246.8
Escherichia coli ATCC 11303	>500	>500	379.8	216.7	227.1	187.3	>500	>500	>500	>500	>500	>500	444.6	385.1
Staphylococcus aureus ATCC 25923	251.3	132.6	>500	>500	189.4	111.6	267.8	211.7	>500	>500	376.8	281.5	265.1	116.3
Enterococcus faecalis ATCC 29212	333.7	258.1	>500	>500	236.9	91.3	>500	>500	>500	>500	255.3	459.1	>500	>500
Salmonella typhi ATCC 6539	>500	>500	>500	>500	>500	>500	461.8	345.9	>500	>500	>500	>500	473.9	399.7
Candida albicans ATCC 90028	125.8	86.4	365.8	244.3	156.9	78.3	>500	>500	471.4	358.2	211.5	184.2	198.2	101.3

MIC—Minimum Inhibitory Concentration expressed in µg mL⁻¹. AE—Aqueous extract, EE—Ethanolic extract.

.

The ethanolic extracts of these plants were more effective than those based in pure water. Moderate antibacterial activity was observed for the ethanolic extract of Y. aloifolia on Escherichia coli ATCC 25922 (MIC, 246.8 µg mL⁻¹),whereas those of S. edule (MIC, 246.8 µg mL⁻¹) and R. obtusifolius (MIC, 187.3 µg mL⁻¹) were more effective on Escherichia coli ATCC 11303 (Table 2). Interestingly, the viability of Staphylococcus aureus ATCC 25923 was strongly inhibited by the ethanolic extracts of S. nigrescens (MIC, 132.6 µg mL⁻¹), R. obtusifolius (MIC, 111.6 µg mL⁻¹), A. hybridus (MIC, 211.7 µg mL⁻¹), and Y. aloifolia (MIC, 116.3 µg mL⁻¹). Intense inhibitory activity of the ethanolic extract of R. obtusifolius was observed on Enterococcus faecalis ATCC 29212 (MIC, 91.3 µg mL⁻¹) (Table 2). Conversely, moderate antibacterial activity was exerted by the ethanolic extract of A. hybridus on Salmonella typhi ATCC 6539, whereas weak inhibitory activity was noted in the ethanolic extracts from other plants assayed (Table 2). The ethanolic extracts from the leaves of R. obtusifolius (78.3 µg mL⁻¹) and those from the berries of S. nigrescens (86.4 µg mL⁻¹) were the most effective against Candida albicans ATCC 90028.

Plant extracts are a complex mixture of compounds with therapeutic potential which may cause fewer side effects compared to synthetic drugs [41]. These natural preparations may also have low probabilities to induce microbial resistance [41]. The results obtained in our microbiological assessment suggest the studied plants exert different levels of antimicrobial activity on microorganisms associated with recurrent infections of the gastrointestinal tract. However, more investigation is required to learn of the possible variations and their biological activity in terms of seasonal exploration and geographical location.

4. Conclusions

Seven edible plants considered quelites in the northern highlands of Puebla-México were studied to learn more about their chemical and biochemical properties. These plants showed differences in their contents of essential nutrients and nutraceuticals. The hydroalcoholic extracts of these plants showed clear inhibitory properties, such as acting as regulators of key enzymes involved in carbohydrate and lipid absorption, synthesis of cholesterol, and cancer development. Among the studied species, Yucca aloifolia showed higher contents of nutrients and nutraceuticals. All the studied plants had potential properties as regulators of lipid and carbohydrate metabolism; however, the edible inflorescences of Y. aloifolia showed the best activity in all the assayed biological activities.