Betalain Pigments Research Articles

Aqueous beetroot dye as an alternative to haematoxylin and eosin in the diagnosis of breast tumours

Breast tumours are heterogeneous diseases that result from changes in the cells and connective tissues of the breast. Different histological and histochemical stains have been employed to aid in the detection of these changes in routine diagnosis. The aim of this study was to assess the usefulness of aqueous beetroot dye in the histochemical staining of breast tumours. Thirty breast tumour blocks from the Histopathology Laboratory of the University of Calabar Teaching Hospital, Calabar were retrieved, sectioned and stained with Cole’s haematoxylin and eosin and aqueous beetroot. The histological staining and 3-D image characteristics were analysed. The histological types of breast tumours were 7(23.3%) benign, of which 5(16.7%) were grade II benign lesion, 1(3.3%) was fibrocystic disease and 1(3.3%) was fibroadenoma. Among the 23(76.7%) malignant tumours, 15(56.7%) were grade 3, 3(10%) were B4 carcinomas and 3(10%) were B5 types. The sections stained with the red beetroot dye gave distinct metachromatic staining of the connective tissues of the cell membranes of the epithelial cells and other connective tissue cells, cytoskeleton, cytoplasm, basement membrane, and collagen fibres of the stroma in shades of green, purple, and black. The histological staining and 3-D image characteristics of the beetroot-stained benign and malignant tumours were similar to the haematoxylin and eosin-stained counterparts in terms of nuclear, cytoplasmic and connective tissue staining, 3-D surface plot features, and histological details (χ2=1.200; p=0.549). This finding has shown that betalain pigments in beetroot dye gave good metachromatic staining of epithelial and connective tissue changes in breast tumours. Beetroot dye and 3-D surface plots when combined can serve as a cost effective, health and environment-friendly alternative staining method to routine haematoxylin and eosin in diagnosis of breast tumours in low resource areas.

Effect of Spray Dryer Scale Size on the Properties of Dried Beetroot Juice.

Experiments detailing the spray drying of fruit and vegetable juices are necessary at the experimental scale in order to determine the optimum drying conditions and to select the most appropriate carriers and solution formulations for drying on the industrial scale. In this study, the spray-drying process of beetroot juice concentrate on a maltodextrin carrier was analyzed at different dryer scales: mini-laboratory (ML), semi-technical (ST), small industrial (SI), and large industrial (LI). Selected physicochemical properties of the beetroot powders that were obtained (size and microstructure of the powder particles, loose and tapped bulk density, powder flowability, moisture, water activity, violet betalain, and polyphenol content) and their drying efficiencies were determined. Spray drying with the same process parameters but at a larger scale makes it possible to obtain beetroot powders with a larger particle size, better flowability, a color that is more shifted towards red and blue, and a higher retention of violet betalain pigments and polyphenols. As the size of the spray dryer increases, the efficiency of the process expressed in powder yield also increases. To obtain a drying efficiency >90% on an industrial scale, process conditions should be selected to obtain an efficiency of a min. of 50% at the laboratory scale or 80% at the semi-technical scale. Designing the industrial process for spray dryers with a centrifugal atomization system is definitely more effective at the semi-technical scale with the same atomization system than it is at laboratory scale with a two-fluid nozzle.

Natural or light-induced pigment accumulation in grain amaranths coincides with enhanced resistance against insect herbivory.

Increased resistance to insect herbivory in grain amaranth plants is associated with increased betalain pigmentation, either naturally acquired or accumulated in response to blue-red light irradiation. Betalains are water-soluble pigments characteristic of plants of the Caryophyllales order. Their abiotic stress-induced accumulation is believed to protect against oxidative damage, while their defensive function against biotic aggressors is scarce. A previous observation of induced betalain-biosynthetic gene expression in stressed grain amaranth plants led to the proposal that these pigments play a defensive role against insect herbivory. This study provided further support for this premise. First, a comparison of "green" and "red" Amaranthus cruentus phenotypes showed that the latter suffered less insect herbivory damage. Coincidentally, growth and vitality of Manduca sexta larvae were more severely affected when fed on red-leafed A. cruentus plants or on an artificial diet supplemented with red-leaf pigment extracts. Second, the exposure of A. cruentus and A. caudatus plants, having contrasting pigmentation phenotypes, to light enriched in the blue and red wavelength spectra led to pigment accumulation throughout the plant and to increased resistance to insect herbivory. These events were accompanied by the induced expression of known betalain-biosynthetic genes, including uncharacterized DODA genes believed to participate in this biosynthetic pathway in a still undefined way. Finally, transient co-expression of different combinations of betalain-biosynthetic genes in Nicotiana benthamiana led to detectable accumulation of betalamic acid and betanidin. This outcome supported the participation of certain AhDODA and other genes in the grain amaranth betalain-biosynthetic pathway.

Additional betalain accumulation by genetic engineering leads to a novel flower color in lisianthus (Eustoma grandiflorum).

Betalains, comprising violet betacyanins and yellow betaxanthins, are pigments found in plants belonging to the order Caryophyllales. In this study, we induced the accumulation of betalains in ornamental lisianthus (Eustoma grandiflorum) by genetic engineering. Three betalain biosynthetic genes encoding CYP76AD1, dihydroxyphenylalanine (DOPA) 4,5-dioxygenase (DOD), and cyclo-DOPA 5-O-glucosyltransferase (5GT) were expressed under the control of the cauliflower mosaic virus (CaMV) 35S promoter in lisianthus, in which anthocyanin pigments are responsible for the pink flower color. During the selection process on hygromycin-containing media, some shoots with red leaves were obtained. However, most red-colored shoots were suppressed root induction and incapable of further growth. Only clone #1 successfully acclimatized and bloomed, producing pinkish-red flowers, with a slightly greater intensity of red color than that in wild-type flowers. T1 plants derived from clone #1 segregated into five typical flower color phenotypes: wine red, bright pink, pale pink, pale yellow, and salmon pink. Among these, line #1-1 showed high expression levels of all three transgenes and exhibited a novel wine-red flower color. In the flower petals of line #1-1, abundant betacyanins and low-level betaxanthins were coexistent with anthocyanins. In other lines, differences in the relative accumulation of betalain and anthocyanin pigments resulted in flower color variations, as described above. Thus, this study is the first to successfully produce novel flower color varieties in ornamental plants by controlling betalain accumulation through genetic engineering.

Bioactive Betalain Extracts from Cactus Pear Fruit Pulp, Beetroot Tubers, and Amaranth Leaves.

Natural food items and the additional benefits they provide have received considerable attention in recent years. Betalains are nutritious pigments which have valuable biological properties, e.g., antioxidant and free radical scavenging activity. In this study, aqueous betalain extracts were obtained from different coloured cactus pears (purple, red/pink, and orange), amaranth, and beetroot, with and without the addition of ascorbic acid, microwave-heated, and freeze-dried and subsequently analysed by thin layer chromatography (TLC). Beetroot samples without the addition of ascorbic acid (AA) had lower phenols, flavonoids, and ascorbic acid content than beetroot samples extracted with the addition of AA. Amaranth had significantly higher contents of antioxidants than all the other plants. Results for phenolic compounds showed that there were no significant differences between cactus pear cultivars, however, significant differences were seen between the two beetroot samples (microwave-heated with and without AA) as well as amaranth. For flavonoid compounds, amaranth had significantly higher values than all other samples. The lowest flavonoid content was found in beetroot without AA (0.49 mgCE/g). For ascorbic acid, significant differences were noticed between amaranth (71.71 mg/100 g) and samples from cactus pear and beetroot. TLC results showed that purple and red cactus pear samples had the most vivid colours, a reflection of the high betacyanin and betaxanthin contents in the cultivars. Moreover, extracts from cactus pear, beetroot, and amaranth were classified according to a decision tree which was designed by the Code of Federal Regulations/Food Additives Regulation of the EU. The classification of betalain pigment extracts as colouring foods was achieved through enrichment factor calculations and the colourant decision tree. The results showed that the betalain pigment extraction method used is inexpensive, time-saving, energy-saving, non-toxic, and chemical solvent free and yields high concentrations of betalains.

In Vitro Antioxidant Capacity of Opuntia spp. Fruits Measured by the LOX-FL Method and its High Sensitivity Towards Betalains

Current in vitro methodologies neglect or subestimate the contribution of betalains to antioxidant capacity in foods because they do not reflect their in vivo biological mechanisms. In this study, we assessed the sensibility of the lipoxygenase-fluorescein (LOX-FL) method towards betalains, phenolic compounds and ascorbic acid from Opuntia spp. fruits; and (ii) the antioxidant capacity of peel and pulp extracts from Opuntia ficus-indica L. Mill (var. Fresa, Colorada and Blanco) and Opuntia stricta var. Dillenii; by comparing the LOX-FL method to traditional antioxidant methods (ORAC and TEAC). The spectrophotometric monitoring of the LOX-FL reaction avoided interference caused by betalain pigments. Indicaxanthin and betanin showed high antiperoxidative and radical scavenging mechanisms in the LOX-FL assay. O. stricta var. Dillenii tissues the highest antioxidant capacity which correlated with betanin content. ORAC and TEAC antioxidant methods were less sensible towards betalain antioxidant activity. To our knowledge, this is the first time the LOX-FL antioxidant method has been used on betalains and betalain-rich foods.Graphical

Betalain Extracts from Beetroot as Food Colorants: Effect of Temperature and UV-Light on Storability.

Beetroot is a good source of natural food colorants given that it contains significant amount of betalain pigments. This study investigates the three-months storage stability of betalain extracts recovered from unsold beetroot by a specific enzymatic mix. The extract storability was evaluated by the kinetic degradation model of betacyanins and betaxanthins, as well as by pigment color attributes (CIELa*b* parameters) under dark and UV-light exposure at different temperatures (4, 25 and 40°C). Considering all the tested storage conditions, betaxanthins appeared to be more stable than betacyanins in terms of degradation rate constant, half-life time and activation energy. The effect of UV-light was negligible if the pigment was not exposed to temperatures higher than the room one, since no relevant differences were found between the parameters studied at 4 and 25°C. Taking into account the visual color, all the investigated color indexes (total color difference, chroma and hue) were mainly affected by temperature, even if mildly high (40°C), and to a lesser extent by UV-light exposure.

Effect of amino acid supplementation, elicitation and LEDs on Hylocereus costaricensis callus culture for the enhancement of betalain pigments

Hylocereus costaricensis more popularly known as the purplish-red pitaya or dragon fruit belongs to the Cactaceae family. The present study analyzes the effect of amino acids, elicitors, and light quality to improve callus proliferation, enhance betalain content and antioxidant profiles through elicitation and amino acid supplementation. The treatments that stimulated the highest total betalain content in red callus cultures were 50 µM tyrosine that produced a 1.6-fold increase (11.28 ± 0.38 mg/g DM) in betalain content and 200 µM leucine that produced 2.56-fold increase (18.13 ± 5.98 mg/g DM) in betalain content. Elicitation of yellow callus cultures with 100 mg/L yeast extract enhanced betalain content by 2-fold without affecting callus biomass. Betalain content could also be improved by 1.23-fold with 20 µM silver nitrate (12.51 ± 0.55 mg/g DM) while callus dry mass and total phenolic content were elevated at 40 µM silver nitrate. The utilization of light was a promising strategy for increased production of secondary metabolites in callus cultures of H. costaricensis. Both yellow and red callus cultures incubated under red LED lights enhanced total betalain content by 3.8- fold (11.64 ± 0.54 mg/g DM) and 4.8-fold (9.42 ± 1.48 mg/g DM), significantly higher (p < 0.01) than cool white LED lights (control treatment). Amino acid supplementation and elicitation strategies in this study achieved betalain content in yellow and red callus that was higher than that in dragon fruit peel (6.31 ± 1.82 mg/g DM), the pulp (3.26 ± 0.26 mg/g DM), and in vitro seedlings (2.01 ± 0.12 mg/g DM). The present investigation reports a simple and efficient protocol for in vitro betalain production from H. costaricensis. This study may enable the future commercial production of betalain compounds under a controlled environment, independent of climate and soil conditions.

Beetroot Microencapsulation with Pea Protein Using Spray Drying: Physicochemical, Structural and Functional Properties

Beetroot is a root vegetable with carotenoids, phenols, vitamins, minerals, and water-soluble betalain pigments such as betacyanins (red-violet color) and betaxanthins (yellow-orange color), which have many nutritional and health benefits. Its use in the food industry is mainly as a powdered natural dye. This study aims to investigate the effect of adding pea protein to beetroot juice as an encapsulating agent, and the spray-dried temperature on the physicochemical, structural, and functional properties of the powder. The spray drying was conducted at 125 and 150 °C with 3.5% and 7% pea protein used in the mixtures with the beetroot juice. The water content, bulk density, porosity, hygroscopicity, water solubility, water absorption index, color, and microstructure of the obtained powder were determined. In addition, betacyanin, total phenols, antioxidant capacity, and powder encapsulate efficiency were analyzed. Using pea protein in the spray drying of beetroot juice had shown high yields of spray drying and good characteristics of the powdered product. Beetroot powder with 7% of pea protein was more porous and luminous, and less hygroscopic than beetroot powder with 3.5% of pea protein. However, the use of 7% of pea protein increased the amount of water immobilized by the samples and reduced the soluble solids present in the product compared to beetroot powder with 3.5% of pea protein. The use of 7% of pea protein protected beetroot bioactive compound higher than the use of 3.5%. Higher spray-drying temperature (150 °C) significantly decreased phenols content and antioxidant capacity of the beetroot powders (p &lt; 0.05). Results showed using 7% pea protein mixed with beetroot juice and a 125 °C spray-drying temperature gave the most content of the studied bioactive compounds and antioxidant capacity. Moreover, the proposal gives more stable powders from a functionality viewpoint because it showed the higher encapsulate efficiency.

MycoRed: Betalain pigments enable in vivo real-time visualisation of arbuscular mycorrhizal colonisation.

Arbuscular mycorrhiza (AM) are mutualistic interactions formed between soil fungi and plant roots. AM symbiosis is a fundamental and widespread trait in plants with the potential to sustainably enhance future crop yields. However, improving AM fungal association in crop species requires a fundamental understanding of host colonisation dynamics across varying agronomic and ecological contexts. To this end, we demonstrate the use of betalain pigments as in vivo visual markers for the occurrence and distribution of AM fungal colonisation by Rhizophagus irregularis in Medicago truncatula and Nicotiana benthamiana roots. Using established and novel AM-responsive promoters, we assembled multigene reporter constructs that enable the AM-controlled expression of the core betalain synthesis genes. We show that betalain colouration is specifically induced in root tissues and cells where fungal colonisation has occurred. In a rhizotron setup, we also demonstrate that betalain staining allows for the noninvasive tracing of fungal colonisation along the root system over time. We present MycoRed, a useful innovative method that will expand and complement currently used fungal visualisation techniques to advance knowledge in the field of AM symbiosis.

Optimization of Betalain Pigments Extraction Using Beetroot by-Products as a Valuable Source

(1) Background: This study is designed to extract the bioactive compounds from beetroot peel for future use in the food industry. (2) Methods: Spectrophotometry techniques analyzed the effect of conventional solvent extraction on betalains and polyphenolic compounds from beetroot peels. Several treatments by varying for factors (ethanol and citric acid concentration, temperature, and time) were applied to the beetroot peel samples. A Central Composite Design (CCD) has been used to investigate the effect of the extraction parameters on the extraction steps and optimize the betalains and total polyphenols extraction from beetroot. A quadratic model was suggested for all the parameters analyzed and used. (3) Results: The maximum and minimum variables investigated in the experimental plan in the coded form are citric acid concentration (0.10–1.5%), ethanol concentration (10–50%), operating temperature (20–60 °C), and extraction time (15–50 min). The experimental design revealed variation in betalain content ranging from 0.29 to 1.44 mg/g DW, and the yield of polyphenolic varied from 1.64 to 2.74 mg/g DW. The optimized conditions for the maximum recovery of betalains and phenols were citric acid concentration 1.5%, ethanol concentration 50%, temperature 52.52 °C, and extraction time 49.9 min. (4) Conclusions: Overall, it can be noted that the extraction process can be improved by adjusting operating variables in order to maximize the model responses.

Ability of Yeast Metabolic Activity to Reduce Sugars and Stabilize Betalains in Red Beet Juice

To lower the risk of obesity, diabetes, and other related diseases, the WHO recommends that consumers reduce their consumption of sugars. Here, we propose a microbiological method to reduce the sugar content in red beet juice, while incurring only slight losses in the betalain content and maintaining the correct proportion of the other beet juice components. Several yeast strains with different metabolic activities were investigated for their ability to reduce the sugar content in red beet juice, which resulted in a decrease in the extract level corresponding to sugar content from 49.7% to 58.2%. This strategy was found to have the additional advantage of increasing the chemical and microbial stability of the red beet juice. Only slight losses of betalain pigments were noted, to final concentrations of 5.11% w/v and 2.56% w/v for the red and yellow fractions, respectively.

The Influence of Different Pretreatment Methods on Color and Pigment Change in Beetroot Products.

Vegetable processing pomace contains valuable substances such as natural colors that can be reused as functional ingredients. Due to a large amount of water, they are an unstable material. The aim of our research was to assess how the pretreatment method (thermal or nonthermal) affects the properties of powders obtained from beet juice and pomace after the freeze-drying process. The raw material was steamed or sonicated for 10 or 15 min, and then squeezed into juice and pomace. Both squeezed products were freeze-dried. The content of dry substance; L*, a*, and b* color parameters; and the content of betalain pigments were analyzed. Pretreatments increased the proportion of red and yellow in the juices. Steam and ultrasound caused a significant reduction in parameter b* in the dried pomace. A significant increase in betanin in lyophilizates was observed after pretreatment with ultrasound and steam for 15 min. As a result of all experiments, dried juices and pomaces can also be used as a colorant source. However, there is higher potential with pomaces due to their additional internal substances as well as better storage properties. After a few hours, juice was sticky and not ready to use.

Natural coloring red beetroot under effects of gamma radiation

As color is an important attribute related to the visual appeal and quality of food products, the reduction of pigment losses during food processing is a basic concern for the industry. Based on this, the objective of this work was to evaluate the resistance of betalain from beet against the use of gamma radiation. Analyzes were carried out on pigments belonging to the betalain class on beets exposed to different doses of gamma radiation (Co60) using doses of 5, 10 and 15 kGy on beets, in addition to anon-irradiated sample for control. Color parameters were evaluated using the CIE L, a*, b * methodology, as well as the quantification of betalain in the samples. By of results the color parameters were affected in until 50% in relation the control. And the quantity betalain activity in the sample irradiated with doses of gamma radiation was more less 65% in relation the control. Therefore, it can be concluded that there is a potential resistance of the betalain pigment in the face of high doses of radiation, which returns interest to red beetroot dyes since most pigments are fragile to thermal treatments or with ionizing radiation.

Sustainable betalain pigments as eco-friendly film coating over aluminium surface

Sustainable betalain pigments extracted from beetroots were used as eco-friendly film coating over aluminium surface. The film coating was examined and verified by corrosive 1.0 M HCl solution. Characterization of chemical structure of the betalain pigments was done using spectroscopic attenuated total reflectance Fourier transform infrared (ATR-FTIR) and UV–Vis technique. Scanning electron microscope technique was used to demonstrate the eco-friendly film coating before and after been examined via corrosive 1.0 M HCl solution. Various operating conditions were studied to meet highest inhibition efficiency values such as betalain concentration, operating temperature and turbulent flow, and maximum inhibition efficiency = 98.4% was displayed. Multiple adsorption isotherms like Langmuir, Temkin, and Freundlich were used effectively to determine equilibrium constants and adsorption parameters. Dynamic investigations revealed a decline of rate of corrosion (RC) and rise of activation energy values (Ea) at higher betalain concentrations. Thermodynamic investigations revealed physisorption process, exothermic enthalpy, and low entropic values of betalain on aluminium surface. The noticeable specifications of low cost, abundance and sustainability, environmentally friendly, and high technical feasibility of betalain pigments adapt them to be the futuristic outstanding sustainable film coating for metal surfaces.

Shade improves growth, photosynthetic performance, production and postharvest quality in red pitahaya (Hylocereus costaricensis)

High solar radiation impairs the successful cultivation of Hylocereus (Cactaceae) species. This investigation aimed to evaluate shading as a strategy for coping with high solar radiation in red pitahaya (Hylocereus costaricensis), through a long-term field experiment. To this end, we studied growth, physiological performance, productivity and postharvest quality of red pitahaya grown under four shade levels, namely, 35, 50, 65 and 80%, compared to plants under full sunlight (control). Red pitahaya showed superior performance under shading compared to the control, especially plants under about 35% shade. Shading improved initial plant growth followed by high physiological performance, i.e., increased maximum quantum efficiency of photosystem II (Fv/Fm) along with the accumulation of total chlorophyll content, with no photoinhibition and sunburn. Shaded plants showed improved yields along with production precocity, as seen through comparison with the local cultivation under full sunlight. In the second year, a maximum estimated yield of 18.7 t ha−1 was observed under about 35% shade, which is a production increase of 70% vs plants under full sunlight (10.9 t ha−1). Shading improved fruit quality through increased fresh weight, pulp yield, pulp firmness and reddish pulp coloring, indicating the accumulation of betalain pigments. Our findings support shading as a valuable strategy for coping with stress induced by extremely high solar radiation in field plantations, thus providing increased productivity by preventing bleaching and death of Hylocereus, especially in drylands.

Effect of growth regulators on betalain profile in callus culture of Celosia sp.

Red and yellow inflorescence of Celosia indicated the content of betalain pigment, betacyanin and betaxantin. In vitro system through callus culture is one of the strategies for producing plant bioactive compounds including betalain. Therefore, this study aims to analyse the content and profile of betalain pigments in Celosia callus in vitro induced by a combination of auxin and cytokinin. Callus was induced from in vitro cotyledonary and hypocotyl sprouts at 1 week after germination. Callus induction medium was MS + BAP + 2,4-D or NAA. Subsequently, the profile and content of betalain pigments in each type of pigmented callus and in vitro hypocotyl and leaves were analysed by HPLC method. Repeated subcultures every two weeks resulted three types of pigmented callus: red, yellow and greenish white. HPLC analysis of in vitro hypocotyl, leaf and callus tissue derived from red and yellow Celosia inflorescence detected six types of compounds, namely 1) amaranthin, 2) isoamaranthin, 3) betalamic acid, 4) miraxanthin V, 5) 3-methoxytyramine betaxanthine and 6) (S)-tryptophan betaxanthine. Callus from red Celosia contains a small amount of betaxanthin which was higher in yellow Celosia. BAP:2,4-D combination produced higher total content of betalain pigments.

Production of Betacyanins in Transgenic Nicotiana tabacum Increases Tolerance to Salinity.

Although red betalain pigments (betacyanins) have been associated with salinity tolerance in some halophytes like Disphyma australe, efforts to determine whether they have a causal role and the underlying mechanisms have been hampered by a lack of a model system. To address this, we engineered betalain-producing Nicotiana tabacum, by the introduction of three betalain biosynthetic genes. The plants were violet-red due to the accumulation of three betacyanins: betanin, isobetanin, and betanidin. Under salt stress, betacyanic seedlings had increased survivability and leaves of mature plants had higher photochemical quantum yields of photosystem II (Fv/Fm) and faster photosynthetic recovery after saturating light treatment. Under salt stress, compared to controls betacyanic leaf disks had no loss of carotenoids, a slower rate of chlorophyll degradation, and higher Fv/Fm values. Furthermore, simulation of betacyanin pigmentation by using a red filter cover improved Fv/Fm value of green tissue under salt stress. Our results confirm a direct causal role of betacyanins in plant salinity tolerance and indicate a key mechanism is photoprotection. A role in delaying leaf senescence was also indicated, and the enhanced antioxidant capability of the betacyanic leaves suggested a potential contribution to scavenging reactive oxygen species. The study can inform the development of novel biotechnological approaches to improving agricultural productivity in saline-affected areas.

Applicability of optimized hyperspectral indices for estimating Betalain content in Suaeda salsa

ABSTRACT Betalains (Bt) are a collective group of natural, edible, red-yellow plant pigments, and are an important biochemical parameter in vegetable growth. Betalains are believed to have fungicidal activity and their presence in plants is a key to explaining the physiological response and resistance of plants caused by different environmental stress factors or seasonal fluctuations. Hyperspectral remote sensing has proven potential to understand many biochemical processes widely used in investigating vegetation growth condition. To better understand vegetation containing betalain pigments, for growth monitoring and agronomic decision-making, a novel betalain source, Suaeda salsa (S. salsa) was studied by extracting betalain at different phenological periods, and registering the results with time-course measurements of canopy spectral reflectance via hyperspectral remote imaging. Partial least squares regression (PLSR) analysis was undertaken with three spectral transformation methods (the raw hyperspectral reflectance (R), first derivative reflectance (FDR) and second derivative reflectance (SDR). The variable importance in projection (VIP) score resulting from PLSR model was used to determine the key spectral wavelengths and reduce the dimensionality of the hyperspectral reflectance data matrix. The study results demonstrated that Bt content had a significant correlation with the effective wavelength (FD-R 436nm, R 847nm, R 848nm, R 860nm, R 871nm, R 971nm and SD-R 437nm, R 496nm, R 499nm, R 508nm, R 740nm, R 772nm, R 774nm, R 780nm) and optimized spectral indices (simple normalized difference spectral indices (NDSI(802nm,408nm), NDSI(781nm,776nm)), normalized polarization indices (NPDI(781nm,775nm), NPDI(781nm,776nm)), ratio spectral indices (RSI(781nm,776nm), RSI(781nm,775nm)), chlorophyll indices(CI(781nm,776nm), CI(781nm, 775nm)) derived from first derivative spectral reflectance. The PLSR-5 (constructed with the spectral indices) model resulted in an R 2 val (determination coefficient) of 0.79 and root mean square error (RMSEval) was 0.81 μg cm−2. Compared to PLSR-3 model built by sensitive bands, the PLSR-5 model estimation accuracy increased 15%. These results suggest the possibility of estimating Bt content using hyperspectral indices, which contain sufficient information for a successful estimation. Study findings will help researchers in deciding suitable Bt indices for S. salsa growth status and stress assessment, and will facilitate detection growth condition of the vegetables with betalain pigments.

Elucidation of the core betalain biosynthesis pathway in Amaranthus tricolor

Amaranthus tricolor L., a vegetable Amaranthus species, is an economically important crop containing large amounts of betalains. Betalains are natural antioxidants and can be classified into betacyanins and betaxanthins, with red and yellow colors, respectively. A. tricolor cultivars with varying betalain contents, leading to striking red to green coloration, have been commercially produced. However, the molecular differences underlying betalain biosynthesis in various cultivars of A. tricolor remain largely unknown. In this study, A. tricolor cultivars with different colors were chosen for comparative transcriptome analysis. The elevated expression of AmCYP76AD1 in a red-leaf cultivar of A. tricolor was proposed to play a key role in producing red betalain pigments. The functions of AmCYP76AD1, AmDODAα1, AmDODAα2, and AmcDOPA5GT were also characterized through the heterologous engineering of betalain pigments in Nicotiana benthamiana. Moreover, high and low L-DOPA 4,5-dioxygenase activities of AmDODAα1 and AmDODAα2, respectively, were confirmed through in vitro enzymatic assays. Thus, comparative transcriptome analysis combined with functional and enzymatic studies allowed the construction of a core betalain biosynthesis pathway of A. tricolor. These results not only provide novel insights into betalain biosynthesis and evolution in A. tricolor but also provide a basal framework for examining genes related to betalain biosynthesis among different species of Amaranthaceae.