Application of metabolic engineering to enhance the content of alkaloids in medicinal plants.

Plants are a rich source of bioactive compounds, many of which have been exploited for cosmetic, nutritional, and medicinal purposes. Through the characterization of metabolic pathways, as well as the mechanisms responsible for the accumulation of secondary metabolites, researchers have been able to increase the production of bioactive compounds in different plant species for research and commercial applications. The intent of the current review is to describe the metabolic engineering methods that have been used to transform in vitro or field-grown medicinal plants over the last decade and to identify the most effective approaches to increase the production of alkaloids. The articles summarized were categorized into six groups: endogenous enzyme overexpression, foreign enzyme overexpression, transcription factor overexpression, gene silencing, genome editing, and co-overexpression. We conclude that, because of the complex and multi-step nature of biosynthetic pathways, the approach that has been most commonly used to increase the biosynthesis of alkaloids, and the most effective in terms of fold increase, is the co-overexpression of two or more rate-limiting enzymes followed by the manipulation of regulatory genes.

Effects of saline elicitors on saponin production in Agave salmiana plants grown in vitro.

Agave plants are natives of Mexico and have an important role in the functional food industry. Agave salmiana grows in dry and desert soils, which are high in salt content; however, little is known about its response to saline conditions. In this study, A. salmiana plants grown in vitro were exposed to 0.1, 0.5, and 1.0 mM of salt elicitors, including AlCl3, NaCl, and CoCl2, and saponin synthesis and morphological characteristics were examined. Saponins were identified and quantified in ethanolic extracts using HPLC-ELSD. Root length and number, leaf length and number, and plant fresh weight were evaluated to determine the phenological condition of the plant. The presence of salts at various concentrations did not affect the physiological characteristics of the plant. Moreover, 0.5 mM NaCl induced a higher production of total saponin. Chlorogenin glycoside 1 (CG1) and hecogenin glycoside 1 (HG1) content remained unchanged across treatments. By contrast, CG2 and HG2 concentrations tended to decrease in response to increased concentrations of AlCl3, NaCl, or CoCl2. In vitro salt elicitors could be a feasible tool in the synthesis of specific saponins, without compromising on plant biomass. Our findings can be used in further generation of low saponin agave plants in field for the improvement of fermentation yield.

Current Methods for the Discovery of New Active Ingredients from Natural Products for Cosmeceutical Applications.

Cosmeceuticals are designed to serve a dual purpose: to provide desired esthetical effects and to treat dermatological conditions. Natural products derived from plants and marine organisms are a novel source of potential cosmeceutical active ingredients for incorporation into new formulations due to consumer demands. Contrary to common perceptions, most regulatory agencies do not view cosmeceuticals as being a separate category from cosmetics; thus, these products are not regulated accordingly, thereby forcing the consumer to rely on the self-regulatory policies of the cosmetics industry. Cosmeceuticals are advertised as having capabilities that include anti-aging, anti-acne, solar-protective, wound healing, and skin whitening. Such traits normally comprise several biological activities. In order to ensure the safety and efficacy of these products, active ingredients employed in the formulations must undergo a series of tests. In this review, in vitro (enzymatic and cellular) and in vivo tests employed to evaluate the potential of new cosmeceutical active ingredients are discussed, and new trends that are being explored by the cosmeceutical industry are described.

Steroidal Saponin and Flavonol Content and Antioxidant Activity during Sporophyte Development of Maguey (Agave salmiana).

Agave salmiana Otto ex Salm-Dyck has traditionally been used for the production of fermented beverage known as "pulque" that has recently gained acceptance as a functional food. However, the plant requires up to 10 years to be used as raw material. The objective of this work was to evaluate the antioxidant and bioactive principles of Agave salmiana during different stages of development. Wild grown plants from Coahuila, Mexico, were identified based on leaf and spine traits to obtain a representative sample from six different stages of development (I-VI) from 1 to 7 years old. Total phenolic content (TPC), antioxidant activity (AOX), as well as composition and content of flavonols and saponins by HPLC-MS-TOF and HPLC-ELSD-PDA were evaluated. Concentrations of TPC were found to be between 5 to 13 mg gallic acid equivalents/g, reaching a maximum at stage II. The AOX presented a negative tendency from stage I to stage VI (from 148 to 50 µmol Trolox equivalents/g respectively). Kaempferol, quercetin and five saponins were identified. Similar to AOX, flavonols presented a negative concentration tendency with a reduction of 65% between the stage I and VI. Plants of stage III and IV presented the highest content of saponins, particularly chlorogenin glycoside, containing 3.19 and 2.90 mg protodioscin equivalents/g, respectively. These data suggest that plants from stages I to IV may be used as a source of antioxidant and bioactive principles, and that the content of these metabolites could be used as a marker to determine the developmental stage of the plant.

Hydrothermal treatment of maize: Changes in physical, chemical, and functional properties.

The objective of this work was to assess the effects of a traditional parboiling treatment on physical, chemical and functional properties of yellow maize kernels. For this, maize kernels were subjected to the three main stages of a traditional parboiling process (soaking, steaming, and drying) at different moisture contents (15%, 25%, or 35%), and different pressure steaming times (0, 15, or 30 min). Kernels were evaluated for physical and chemical changes, while manually generated endosperm fractions were further evaluated for nutritional and functional changes. The parboiling process negatively altered the maize kernels properties by increasing the number of kernels with burst pericarp and decreasing the total carotenoid content in the endosperm by 42%. However, the most intense conditions (35% moisture and 30 min steam) lowered the number of broken kernels by 41%, and the number of stress cracks by 36%. Results also demonstrated that soaking enhanced the nutritional value of soaked yellow maize by increasing the thiamine content and the bound phenolic content in the endosperm fraction up to 102%. The proper implementation of this hydrothermal treatment could lead to significant enhancements in nutritional and functionality of maize products.

Micropropagation of Agave salmiana: Means to Production of Antioxidant and Bioactive Principles.

Maguey, Agave salmiana, is an important plant for the "pulque" beverage and functional food industries; however, it has several constraints for elite and homogeneous plant production. In this study, a micropropagation process was established to generate in vitro plants. The effect of the method on metabolite content and antioxidant (AOX) activity in regenerated plants was evaluated. Young germinated plantlets were micropropagated from axillary shoots using Murashige and Skoog medium supplemented with L2 vitamins, 0.04 mg/L 2,4-dichlorophenoxyacetic acid and 10 mg/L 6-benzylaminopurine. Total soluble sugars from the aqueous fraction and total phenolic acids, total saponins, and AOX activity of the methanol fraction were determined in wild-type (WT) plants, in in vitro (IN) plants, and ex vitro acclimated plants (EN). The results showed that IN plants have a 50% lower soluble sugar content compared to WT, and EN. The total phenolic acids content was at least 30% higher in micropropagated (IN) and regenerated (EN) plants compared to WT. The total saponin content in IN, and EN plants was 36 and 25 times higher compared to WT. The AOX capacity of IN plants was on average three times higher compared to other treatments. However, no correlation was found between the AOX activity and total phenolic acids or total saponins. A negative and significant correlation (r = -0.927; p = 0.003) was found between the AOX activity and the total soluble sugars content. Micropropagated plants of A. salmiana have a different phytochemical content and bioactivity after the in vitro process compared to WT plants. The micropropagation process could be used as a platform for phytochemical enhancement of Agave plants.

Preventive and therapeutic potential of peptides from cereals against cancer.

Epidemiological studies have shown that regular consumption of food based on whole-grain cereals and their products is associated with reduced risks of various types of degenerative chronic diseases. Food proteins are considered an important source of nutraceutical peptides and amino acids that can exert biological functions to promote health and prevent disease, including cancer. There have been several reports on peptides with anti-tumour activity in recent years. Plant-derived peptides, such as rapeseed, amaranth and soybean lunasin have received main attention. In this review, we extend this vision to analyse the evidence of current advances in peptides in cereals such as wheat, maize, rice, barley, rye and pseudocereals compared with soybean. We also show evidence of several mechanisms through which bioactive peptide exerts anti-tumour activity. Finally, we report the current status of major strategies for the fractionation, isolation and characterisation of bioactive peptides in cereals. BIOLOGICAL SIGNIFICANCE: In recent reports, it has been shown that peptides are an interesting alternative in the search for new treatments for cancer. One of the most studied sources of these peptides is food proteins; however, a review that includes more recent findings for cereals as a potential source of bioactive peptides in the treatment of cancer, the techniques for their isolation and characterisation and the assays used to prove their bioactivity is not available. This review can be used as a tool in the search for new sources of anti-cancer peptides. The authors have no conflicts of interest, financial or otherwise.

Micropropagation effect on the anti-carcinogenic activitiy of polyphenolics from Mexican oregano (Poliomintha glabrescens Gray) in human colon cancer cells HT-29.

Phenolic extracts obtained from spices are known to have anti-carcinogenic activities but little is known about the effect of micropropagation on these beneficial effects. The main objective of this study was to evaluate the cytotoxic activity of flavonoid-enriched extracts (FEE) from the leaves of wild (WT), in vitro (IN), and ex vitro (EX) grown oregano plants in colon cancer cells HT-29 and the non-cancer cells CCD-18Co. Cell proliferation of HT-29 cells was reduced to 50 % by WT, IN, and EX at concentrations of 4.01, 1.32, and 4.84 mg of gallic acid equivalents (GAE)/L, respectively. In contrast, in CCD-18Co cells, higher concentrations were required for the same cytotoxic effect. At 6 mg GAE/L, WT and IN reduced the production of reactive oxygen species (ROS) of lipopolysaccharides (LPS)-stimulated control cells to 59.89 and 59.43 %, respectively, and EX to 73.89 %. The mRNA of Caspase-3 was increased 1.53-fold when cells were treated with 4 mg GAE/L of IN extract, and tumor necrosis factor receptor superfamily, member 6 (FAS), and BCL2-associated X protein (BAX) mRNA increased 2.55 and 1.53 fold, respectively. Results on protein expression corroborated the apoptotic effects with a significant decrease of B-cell lymphoma 2 (BCL2) expression for all treatments but more remarkable for EX that also showed the most intense signal of BAX. Overall, FEE extracts derived from micropropagation had increased pro-apoptotic effects, however extracts from the in vitro plants produced more efficacy at the transcriptional level while extracts from the ex vitro plant were superior at the traductional level.

Chemopreventive effects of free and bound phenolics associated to steep waters (nejayote) obtained after nixtamalization of different maize types.

Free and bound phenolics extracts from nejayote solids were obtained after optimally lime-cooking blue, normal white, red, normal yellow, high-carotenoid and quality protein maize types. The extraction yield ranged from 4.47 to 10.05%. Bound phenolics extracts had higher content of total phenolics, antioxidant activity and ferulic acid compared to the free phenolics extracts. In general, free phenolics extracts were less cytotoxic than the bound phenolics counterparts. Bound phenolics extracts had higher induction of quinone reductase (QR) and particularly the normal yellow nejayote exerted the highest chemopreventive index tested in Hepa1c1c7 cells. When tested for monofunctional phase 2 induction capacity in BPrc1 cells, the bound phenolics extracts of blue, normal white and quality protein nejayotes were better inducers than the normal yellow counterpart. Particularly, the free phenolics extract of the white maize nejayote induced BPrc1 cells QR and exerted a higher chemopreventive index compared to the bound phenolics extract. Therefore, the nejayote of the normal white maize was the best source of monofunctional phase 2 enzyme inducers.

Grain and tortilla quality in landraces and improved maize grown in the highlands of Mexico.

The maize produced in the highlands of Mexico (>2,400 masl) is generally not accepted by the flour and masa and tortilla industry. The objective of this work was to evaluate the grain quality and tortilla properties of maize landraces commonly grown in the highlands of Mexico and compare them with improved germplasm (hybrids). Germplasm analysis included 11 landraces, 32 white hybrids, and six yellow hybrids. Grain quality was analyzed for a range of physical and chemical factors, as well as for alkaline cooking quality. Landrace grains tended to be heterogeneous in terms of size, hardness and color. All landraces had soft-intermediate grains with an average flotation index (FI) of 61%. In contrast, hybrid grains were homogenous in size and color, and harder than landrace grains, with a FI of 38%. Protein, free sugars, oil and phenolic content in landraces were higher than in the hybrids. Significant correlations were found between phenolic content and tortilla color (r= -0.60; p<0.001). Three landraces were identified as appropriate for the masa and tortilla industry, while all the hybrids evaluated fulfilled the requirements of this industry.