Search

Betalains are naturally occurring pigments sourced mainly from Beta vulgaris (beetroot), Hylocereus spp. (dragon fruit), Amaranthus spp., and Opuntia spp. Betalains are widely used for their vibrant colors and health-promoting properties. These nitrogenous, water-soluble pigments are crucial colorants in the food industry, responsible for the red, purple, and yellow plant tissues, predominantly in the order Caryophyllales. They are grouped into betacyanins, with reddish-violet hues, and betaxanthins, yellow to orange. Examples include beetroot stems for betacyanins and yellow pitaya pulp for betaxanthins. Several pharmacological activities were reviewed in the scientific literature, describing their potential implications for human health. In this review, we focused on the main and latest studies on the pharmacological effects and mechanisms of betalains, including antioxidant, anti-inflammatory, antihypertensive, hypolipidemic, antidiabetic, hepatoprotective, neuroprotective, anticancer, and antimicrobial properties, in both in vitro and in vivo studies. Overall, betalain consumption is considered safe, with no major adverse effects or allergic reactions reported. We also approached topics such as the pharmacokinetics, bioavailability, stability, and enhanced stabilization of betalains. This article provides a comprehensive overview of bioactive potential of betalains, highlighting the biochemical mechanisms involved. The current knowledge broadens the clinical applicability of betalains, making them potential sources of nutraceutical compounds that can be used to develop functional foods.

Identification of metabolic engineering targets is a fundamental challenge in strain development programs. While high-throughput (HTP) genetic engineering methodologies capable of generating vast diversity are being developed at a rapid rate, a majority of industrially interesting molecules cannot be screened at sufficient throughput to leverage these techniques. We propose a workflow that couples HTP screening of common precursors (e.g., amino acids) that can be screened either directly or by artificial biosensors, with low-throughput targeted validation of the molecule of interest to uncover nonintuitive beneficial metabolic engineering targets and combinations hereof. Using this workflow, we identified several nonobvious novel targets for improving p -coumaric acid ( p -CA) and l-DOPA production from two large 4k gRNA libraries each deregulating 1000 metabolic genes in the yeast Saccharomyces cerevisiae . We initially screened yeast cells transformed with gRNA library plasmids for individual regulatory targets improving the production of l-tyrosine-derived betaxanthins, identifying 30 targets that increased intracellular betaxanthin content 3.5-5.7 fold. Hereafter, we screened the targets individually in a high-producing p -CA strain, narrowing down the targets to six that increased the secreted titer by up to 15%. To investigate whether any of the six targets could be additively combined to improve p -CA production further, we created a gRNA multiplexing library and subjected it to our proposed coupled workflow. The combination of regulating PYC1 and NTH2 simultaneously resulted in the highest (threefold) improvement of the betaxanthin content, and an additive trend was also observed in the p -CA strain. Lastly, we tested the initial 30 targets in a l-DOPA producing strain, identifying 10 targets that increased the secreted titer by up to 89%, further validating our screening by proxy workflow. This coupled approach is useful for strain development in the absence of direct HTP screening assays for products of interest.

Genetic engineering of flower color provides biotechnological products such as blue carnations or roses by accumulating delphinidin-based anthocyanins not naturally existing in these plant species. Betalains are another class of pigments that in plants are only synthesized in the order Caryophyllales. Although they have been engineered in several plant species, especially red-violet betacyanins, the yellow betaxanthins have yet to be engineered in ornamental plants. We attempted to produce yellow-flowered gentians by genetic engineering of betaxanthin pigments. First, white-flowered gentian lines were produced by knocking out the dihydroflavonol 4-reductase (DFR) gene using CRISPR/Cas9-mediated genome editing. Beta vulgaris BvCYP76AD6 and Mirabilis jalapa MjDOD, driven by gentian petal-specific promoters, flavonoid 3',5'-hydroxylase (F3'5'H) and anthocyanin 5,3'-aromatic acyltransferase (AT), respectively, were transformed into the above DFR-knockout white-flowered line; the resultant gentian plants had vivid yellow flowers. Expression analysis and pigment analysis revealed petal-specific expression and accumulation of seven known betaxanthins in their petals to c. 0.06-0.08 μmol g FW -1 . Genetic engineering of vivid yellow-flowered plants can be achieved by combining genome editing and a suitable expression of betaxanthin-biosynthetic genes in ornamental plants., (© 2023 The Authors. New Phytologist © 2023 New Phytologist Foundation.)

The yellow pitahaya peels generated as by-products during the consumption and processing of the fresh fruit are a rich and underutilized source of betaxanthins (natural yellow-orange pigment with antioxidant activity) and mucilage (structuring material used in the spray-drying process), molecules of high interest for the food industry. In this work, the betaxanthin-rich extract (BRE) obtained from this by-product was microencapsulated by spray drying (SD) using pitahaya peel mucilage (MPP) and maltodextrin (MD) as wall materials. Both types of microencapsulates (i.e., SD-MPP and SD-MD) retained high betaxanthin content (as measured by UV-vis) and antioxidant activity (ORAC). These microencapsulates were characterized structurally (FTIR and zeta potential), morphologically (SEM and particle size/polydispersity index), and thermally (DSC/TGA). The powdered microencapsulates were incorporated into the formulation of candy gummies as a food model, which were subjected to an in vitro gastrointestinal digestion process. The characterization study (FTIR and antioxidant activity) of the microcapsules showed that the fruit peel mucilage favors the retention of betaxanthins, while the SEM analysis revealed a particle size of multimodal distribution and heterogeneous morphology. The addition of SD-MPP microcapsules in the candy gummy formulation favored the total dietary fiber content as well as the gumminess and chewiness of the food matrix; however, the inhibition of AAPH • (%) was affected. The stability of the yellow color in the gummies after 30 days of storage indicates its suitability for storage. Consequently, the microencapsulation of betaxanthins with pitahaya peel mucilage can be used as a food additive colorant in the food industry, replacing synthetic colorants, to develop products with beneficial qualities for health that can satisfy the growing demand of consumers.

Effects of tray rotation speeds (TRS: 0, 20, 40 rpm), temperatures (50, 60, 70 °C) and wavelength spectra (mid and near-infrared) were comparatively evaluated on improving drying kinetics, physicochemical properties and bioactive content of red dragon fruits. Results indicated that successive increases in TRS and temperature led to significant reductions in drying time and increases in drying rates and moisture diffusivity. High TRS (40 rpm) and lower temperatures (50, 60 °C) also improved colour, total soluble solids, rehydration ratio, total phenolics and flavonoid contents, betalain content and antioxidant activity. Meanwhile, NIR drying presented a more energy-efficient approach, but with substantial reductions in quality properties compared with MIR drying. Overall, the results suggested the importance of wavelength absorption properties of plant tissues and potential avoidance of localized overheating for enhanced efficiency during infrared drying and prompted the development of suitable approaches and optimization studies for improving efficiency., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

There is a growing interest in establishing the methylotrophic yeast Pichia pastoris as microbial cell factories for producing fuels, chemicals, and natural products, particularly with methanol as the feedstock. Although CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) based genome editing technology has been established for the integration of multigene biosynthetic pathways, long (500-1000 bp) homology arms are generally required, probably due to low homologous recombination (HR) efficiency in P. pastoris . To achieve efficient genome integration of heterologous genes with short homology arms, we aimed to enhance HR efficiency by introducing the recombination machinery from Saccharomyces cerevisiae . First, we overexpressed HR related genes, including RAD52 , RAD59 , MRE11 , and SAE2 , and evaluated their effects on genome integration efficiency. Then, we constructed HR efficiency enhanced P. pastoris , which enabled single-, two-, and three-loci integration of heterologous gene expression cassettes with ∼40 bp homology arms with efficiencies as high as 100%, ∼98%, and ∼81%, respectively. Finally, we demonstrated the construction of β-carotene producing strain and the optimization of betaxanthin producing strain in a single step. The HR efficiency enhanced P. pastoris strains can be used for the construction of robust cell factories, and our machinery engineering strategy can be employed for the modification of other nonconventional yeasts.

Cytochrome P450 enzymes (P450s) are a superfamily of heme-thiolate proteins widely existing in various organisms and play a key role in the metabolic network and secondary metabolism. However, the low expression levels and activities have become the biggest challenge for P450s studies. To improve the functional expression of P450s in Saccharomyces cerevisiae , an Arabidopsis thaliana cDNA library was expressed in the betaxanthin-producing yeast strain, which functioned as a biosensor for high throughput screening. Three new target genes AtGRP7 , AtMSBP1 , and AtCOL4 were identified to improve the functional expression of CYP76AD1 in yeast, with accordingly the accumulation of betaxanthin increased for 1.32-, 1.86-, and 1.10-fold, respectively. In addition, these three targets worked synergistically/additively to improve the production of betaxanthin, representing a total of 2.36-fold improvement when compared with the parent strain. More importantly, these genes were also determined to effectively increase the activity of another P450 enzyme (CYP736A167), catalyzing the hydroxylation of α -santalene to produce Z-α-santalol. Simultaneous overexpression of AtGRP7 , AtMSBP1 , and AtCOL4 increased α -santalene to Z-α-santalol conversion rate for more than 2.97-fold. The present study reported a novel strategy to improve the functional expression of P450s in S. cerevisiae and promises the construction of platform yeast strains for the production of natural products., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Jiang, Dong, Liu, Shi, Wang, Tao, Liang and Lian.)

Patients with cancer are prone to several debilitating side effects including fatigue, insomnia, depression and cognitive disturbances. Beetroot ( Beta vulgaris L.) as a health promoting functional food may be potentially beneficial in cancer. As a source of polyphenols, flavonoids, dietary nitrates and other useful nutrients, beetroot supplementation may provide a holistic means to prevent cancer and manage undesired effects associated with chemotherapy. The main aim of this narrative review is to discuss beetroot's nutrient composition, current studies on its potential utility in chemoprevention and cancer-related fatigue or treatment-related side effects such as cardiotoxicity. This review aims to provide the current status of knowledge and to identify the related research gaps in this area. The flavonoids and polyphenolic components present in abundance in beetroot support its significant antioxidant and anti-inflammatory capacities. Most in vitro and in vivo studies have shown promising results; however, the molecular mechanisms underlying chemopreventive and chemoprotective effects of beetroot have not been completely elucidated. Although recent clinical trials have shown that beetroot supplementation improves human performance, translational studies on beetroot and its functional benefits in managing fatigue or other symptoms in patients with cancer are still lacking., Competing Interests: CONFLICTS OF INTEREST No potential conflicts of interest were disclosed., (Copyright © 2021 Korean Society of Cancer Prevention.)

Betalains are emerging natural pigments with high tinctorial strength and stability, physiological activities, and fluorescent properties for potential application in food, cosmetic, and pharmaceutical industries. Betalains including yellow betaxanthins and red betacyanins are mainly restricted in the Caryophyllales plants. To expand the availability of individual betaxanthins, here, we constructed an Escherichia coli BTA6 for de novo biosynthesis of betalamic acid. Using this strain as a monoculture platform, 14 yellow and 2 red betaxanthins were produced by feeding amino acids and amines. Furthermore, we constructed an l-histidine overproducing strain using chromosome engineering to deattenuate regulation and established a coculture system. After optimization of the initial inoculation ratios and fermentation conditions, the compatible and robust coculture system produced 287.69 mg/L of histidine-betaxanthin. This is the first report on de novo production of betaxanthins in engineered E. coli using glucose as a carbon source. Our work highlights the feasibility of microbial cell factories to produce individual betalains.

In this study, we developed a simple and economical method for the green synthesis of Cu 2+ sensors based on betaxanthin pigments. Aminoisophthalic acid-betaxanthin was synthesized by coupling 2-aminoisophthalic acid and betalamic acid produced from DOPA-extradiol-4,5-dioxygenase in situ and in vitro. The resulting 2-aminoterephthalic acid-betaxanthin (2-AIPA-BX) presented a satisfying fluorescence quantum yield in water and a high degree of selectivity for Cu 2+ over interfering metal ions. The bioproduction process of 2-AIPA-BX was scaled up from test tubes to 1 L-flasks, indicating the robustness and reproducibility of this method. Additionally, we successfully incorporated 2-AIPA-BX into paper-based analytical devices to facilitate simple, inexpensive, and portable setup with lower sample consumption for onsite monitoring of environmental and biological samples.

A biosensor screening assay based on the synthesis of betaxanthin was applied to relatively high throughput screening of the L-tyrosine mutant library. In the assays, fluorescence output showed a linear relationship between extracellular L-tyrosine content and yellow pigment formation. In addition, the yellow pigment accumulation of the L-tyrosine high-yield strain can be easily distinguished with the naked eye compared with the wild-type strain. As a result, numerous mutants that exhibited significantly increased coloration, were screened out after random mutagenesis, and p-coumaric acid production in mutants NK-A3 and NK-B4, were remarkably improved by 4-fold more than that of the wild-type strain. In general, this study provides a novel strategy for screening mutant libraries in the search for highly L-tyrosine-producing strains.

We started a cell suspension culture from magenta coloured calli of cockscomb to study the effect of biotic and abiotic elicitors on the biosynthesis of betalain pigments. The cultures were grown in a flask containing 30 ml MS media fortified with 13.5 μM 2,4-D and 0.44 μM BAP. These cultures were elicited during its log-phase of growth using fungal elicitors (prepared from mycelia of Fusarium oxysporum) , yeast extract, copper sulphate and cobalt chloride. The elicitation reduced the cell count, cell viability and percent pigmented cell in the suspension culture. Similarly, it also resulted in reduced betalain content by all the elicitors except 0.125 × 10 -3 % fungal elicitor. Rather, fungal elicitor at this concentration significantly enhanced the amaranthin, betanin, betalamic acid and betaxanthin content in the culture. Besides this, copper sulphate doubled the pigment contribution (ratio of particular pigment content to total pigment content) of betaxanthin at all the concentrations. Therefore, we conclude that fungal elicitor can further be investigated to enhance the content of betalain pigments in suspension culture at a larger scale., Competing Interests: Compliance with ethical standardsThe authors declare that they have no conflict of interest.

Betalains are a family of natural pigments found exclusively in the plant order Caryophyllales. All members of this chemical family are biosynthesized through the common intermediate betalamic acid, which is capable of spontaneously condensing with various primary and secondary amines to produce betalains. Of particular interest is the red-violet betanin, most commonly obtained from Beta vulgaris (beet) as a natural food dye. We demonstrate the first complete microbial production of betanin in Saccharomyces cerevisiae from glucose, an early step towards a fermentation process enabling rapid, on-demand production of this natural dye. A titer of 17mg/L was achieved, corresponding to a color intensity obtained from 10g/L of beetroot extract. Further, we expanded the spectrum of betalain colors by condensing betalamic acid with various amines fed to an engineered strain of S. cerevisiae. Our work establishes a platform for microbial production of betalains of various colors as a potential alternative to land- and resource-intensive agricultural production., (Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.)

The search for natural pigments has been driven by growing evidence indicating that synthetic colorants can cause deleterious health effects. Betalains, in addition to anthocyanins, have been proposed as an alternative to address this need. However, the incorporation of natural pigments poses some challenges to the food industry, such as reduced stability in comparison to their synthetic counterparts. Moreover, betalains are not well studied in comparison to anthocyanins and information about the effects of processing on their physicochemical properties and stability is scattered. Thus, this review will provide an overview of the recent research on the extraction and processing of betalains from natural sources, and comparison of their colorant and physicochemical properties with anthocyanins., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Betalains are important pigments for the food, pharmaceutical, and cosmetics industry. In the yellow Stenocereus pruinosus fruits (pitayas), total betalain concentration, Folin-Ciocalteu reduction capacity, and antiradical capacity per dry weight were 2345.9μgg -1 , 7.3mg gallic acid equivalentsg -1 , and 48.8μmol Trolox equivalentg -1 , respectively. The stability of betaxanthins, which represent 89% of total betalains in yellow pitayas, was evaluated over a range of pH, temperature, as well as in the presence of food additives. Maximum stability was observed at pH6.6, and addition of ascorbic acid increased the half-life 1.8 times. Thermal stability at pH6.48±0.05 was also evaluated from 50°C to 80°C, over which the activation energy for betaxanthin degradation was determined to be 66.2kJmol -1 . Model gelatin gummies and beverages were then prepared with pitaya juice or pulp, and pigment retention and color parameters were investigated during storage under various conditions. To match the yellow color of commercial products, gummies were supplemented with 4.6% w/w juice or pulp, and beverages were supplemented with 5% w/v juice, achieving H° values of 69.0-86.2° and 64.6-87.1°, respectively. Results indicate that betaxanthins were more stable in gummies than in beverages, and that pigment retention increased when products were stored in the dark or at low temperatures. Also, different changes in color during storage were observed between gummies and beverages., (Copyright © 2016 Elsevier Ltd. All rights reserved.)