Pepper Carotenoids Research Articles

Development of an HPLC-PDA Method for the Determination of Capsanthin, Zeaxanthin, Lutein, β-Cryptoxanthin and β-Carotene Simultaneously in Chili Peppers and Products.

For the better standardization and widespread application of the determination method of carotenoids in both chili peppers and their products, this work reports for the first time the simultaneous determination of five main carotenoids, including capsanthin, zeaxanthin, lutein, β-cryptoxanthin and β-carotene in chili peppers and their products, with optimized extraction and the high-performance liquid chromatography (HPLC) method. All parameters in the methodological evaluation were found to be in good stability, recovery and accuracy compliance with the reference values; the R coefficients for the calibration curves were more than 0.998; and the LODs and LOQs varied from 0.020 to 0.063 and from 0.067 to 0.209 mg/L, respectively. The characterization of five carotenoids in chili peppers and their products passed all the required validation criteria. The method was applied in the determination of carotenoids in nine fresh chili peppers and seven chili pepper products.

Nanoencapsulation of Red Bell Pepper Carotenoids: Comparison of Encapsulating Agents in an Emulsion Based System

Red bell pepper extract rich in carotenoids was (RBPE) encapsulated with four different encapsulating agents: calcium caseinate (ECC), bovine gelatin (EBG), whey proteins isolate (EWPI), and concentrate (EWPC), aiming to investigate the most effective material to coat and enable the water dispersibility of pigments. Formulations were obtained by the oil in water (O/W) emulsification technique, followed by freeze-drying. Samples were analyzed by encapsulation efficiency, high-performance liquid chromatography (HPLC), Fourier transform infrared spectroscopy (FTIR), dynamic light scattering (DLS), atomic force microscopy (AFM), thermogravimetric analysis (TGA), dispersion stability, and CIELab. Nanoformulations showed a carotenoid encapsulation efficiency of 54.0% (ECC), 57.6% (EWPI), 56.6 % (EWPC), 64.0 % (EBG). Recovered carotenoid profiles from nanoformulations showed similarity to the RBPE, indicating the efficiency of the encapsulation process. Average particle sizes of approximately 109 nm (ECC), 71 nm (EWPI), 64 nm (EWPC), and 173 nm (EBG) were obtained. AFM revealed that all formulations exhibited spherical forms and a heterogeneous distribution profile. Regarding TGA, formulations presented similar thermal behaviors to and lower decomposition speeds than RBPE, suggesting improved thermal stability. Powder formulations were easily dispersed in water (8 mg/mL) and presented intense color and stability to sedimentation for 48 h. Results indicated that all formulations and the chosen technique efficiently increased carotenoid dispersibility in water, indicating their potential to be applied as natural food pigments.

Capsanthin induces G1/S phase arrest, erlotinib-sensitivity and inhibits tumor progression by suppressing EZH2-mediated epigenetically silencing of p21 in triple-negative breast cancer cells.

Capsanthin is a naturally occurring red pepper carotenoid with possible antitumor activity, but its antitumor mechanisms have yet to be delineated. We tested the anti-proliferative activity of capsanthin with human triple-negative breast cancer (TNBC) and found that cell proliferation was inhibited after 24, 48 and 72 h of treatment. We also investigated the cellular and molecular mechanisms of the antitumor efficacy of capsanthin on TNBC cells and found that capsanthin delayed cell-cycle progression at the G1/S stage, that cyclin A expression was suppressed, and that p21 expression was upregulated. Capsanthin also inhibited the EZH2 expression and EZH2 could binding to the p21 promoter in TNBC cells. We further discovered that capsanthin has synthetic effects when combined with erlotinib (Tarceva). In the animal experiment, we found that the capsanthin-induced inhibition of TNBC cell proliferation decreased the incidence of the initiation and growth of TNBC cell–derived tumors in mice. Our study reveals that capsanthin exerted antitumor effects through delaying cell-cycle progression, induces erlotinib-sensitivity and inhibits tumor progression by inhibiting EZH2/p21 axis, and capsanthin is a potential drug candidate for development of a safe and effective therapy against TNBCs, especially for TNBCs that have developed resistance to targeting therapy.

Chili Pepper Carotenoids: Nutraceutical Properties and Mechanisms of Action.

Chili pepper is a prominent cultivated horticultural crop that is traditionally used for food seasoning and is applied for the treatment and prevention of multiple diseases. Its beneficial health properties are due to its abundance and variety of bioactive components, such as carotenoids, capsaicinoids, and vitamins. In particular, carotenoids have important nutraceutical properties, and several studies have focused on their potential in the prevention and treatment of human diseases. In this article, we reviewed the state of knowledge of general aspects of chili pepper carotenoids (biosynthesis pathway, types and content in Capsicum spp., and the effects of processing on carotenoid content) and recent findings on the effects of carotenoid nutraceuticals, such as antioxidant, cancer preventive, anti-inflammatory, cardiovascular disorder preventive, and anti-obesity effects.

Accumulation characteristics of carotenoids and adaptive fruit color variation in ornamental pepper

To understand better the mechanisms of the pigment accumulation in ornamental pepper, the fruit color, pigment and expression of carotenoid biosynthetic genes were investigated. The test materials were divided into three categories. The first category included cultivars B3, J1, J2 and Q3, the second were M3, M3′, W8 and WU, C2, S1, W3 and W4 belonged to the third category. The pigment changes during ripening were characterized by loss of chlorophyll and anthocyanin, and a rapid accumulation of carotenoids. It was found that GGPS, PSY, CRTZ-2 and CCS genes have abundant transcription during the accumulation of carotenoids in ornamental pepper, which have a more important regulatory function. Correlation analysis were conducted between fruit color, pigment and expression of genes, it showed that there were significant correlation between key genes of carotenoid biosynthetic pathway, among which PSY was highly correlated with CCS and CRTZ-2, CCS was highly correlated with CRTZ-2. Chromaticity values were negatively correlated with anthocyanin content in fruits, and the correlation between a* and carotenoid content was significantly higher. The information will be benefit for exploring the carotenoid regulation mechanism of ornamental pepper, and providing theoretical for fruit color breeding of ornamental pepper.

Genome-wide identification and expression analysis of the carotenoid metabolic pathway genes in pepper (Capsicum annuum L.)

Carotenoids belong to a major family of isoprenoids which play vital roles in plant growth and development. Pepper (Capsicum annuum L.) is not only a vegetable and condiment grown worldwide, but also can be used as a model organism in fruit color research. In this study, 61 genes known to be involved in mevalonate (MVA), plastidial 2-C-methyl-d-erythritol-4-phosphate (MEP) and carotenoid metabolism (biosynthesis and catabolism) pathways were identified and characterized based on the genome of pepper plants. In the present study, it was found that 56 of the 61 carotenoid metabolism genes in pepper were unevenly distributed throughout 12 chromosomes. CaGGPPS2, CaGGPPS4a and CaGGPPS4b were generally clustered into one region. This study identified CaHMGR1/CaHMGR3 and CaHMGR1/CaHMGR2 as segregation duplication events. In addition, the pepper carotenoid metabolism genes had high variations in gene structure. The key carotenoid metabolism genes showed different expression patterns during the pepper fruit development and under environmental stress conditions. The results obtained in this research investigation will contribute insight to the evolution and function of carotenoid metabolic pathways in pepper fruit.

Reversed-Phase High-Performance Liquid Chromatography for the Quantification and Optimization for Extracting 10 Kinds of Carotenoids in Pepper (Capsicum annuum L.) Leaves.

Carotenoids are considered to be crucial elements in many fields and, furthermore, the significant factor in pepper leaves under low light and chilling temperature. However, little literature focused on the method to determinate and extract the contents of carotenoid compositions in pepper leaves. Therefore, a time-saving and highly sensitive reversed-phase high-performance liquid chromatography method for separation and quantification of 10 carotenoids was developed, and an optimized technological process for carotenoid composition extraction in pepper leaves was established for the first time. Our final method concluded that six xanthophylls eluted after about 9-26 min. In contrast, four carotenes showed higher retention times after nearly 28-40 min, which significantly shortened time and improved efficiency. Meanwhile, we suggested that 8 mL of 20% KOH-methanol solution should be added to perform saponification at 60 °C for 30 min. The ratio of solid-liquid was 1:8, and the ultrasound-assisted extraction time was 40 min.

A New Nonsense Mutation in Capsanthin/Capsorubin Synthase Controlling Orange Pepper Fruit

Carotenoids are plant pigments that play a major role in conferring fruit color. Carotenoid content is often controlled by genetic variation in the biosynthetic genes. The color of mature pepper fruit is mainly classified as red, orange, and yellow. Orange and yellow fruit colors are determined by mutations in phytoene synthase (Psy) and capsanthin-capsorubin synthase (Ccs), respectively. In contrast to the current fruit color model, we hypothesized that genetic variation in Ccs also controls orange fruit color in pepper. Ripe fruit of Capsicum annuum ‘K146465’ is orange, and its carotenoid profile obtained by HPLC analysis showed a lack of the major pepper carotenoid capsanthin but an abundance of lutein, zeaxanthin, and β-carotene compared to red pepper. cDNA cloning and sequencing analysis detected a new nonsense mutation due to a T1026 insertion in the coding region of Ccs, but no DNA sequence variation in Psy. We developed a derived cleaved amplified polymorphic sequence (dCAPS) marker to distinguish the nonsense mutation in Ccs. Genetic analysis of the F2 population derived from C. annuum ‘Sweet Banana’ (red fruit color) × C. annuum ‘K146465’ revealed that orange fruit color is determined by a single recessive gene. The nonsense mutation in Ccs distinguished by the dCAPS marker co-segregated with orange fruit color in the F2 population. This germplasm, coupled with the dCAPS marker and carotenoid profiling, will facilitate marker-assisted breeding to select orange fruit color and improve lutein, zeaxanthin, and β-carotene levels in pepper.

Increasing solubility of red bell pepper carotenoids by complexation with 2-hydroxypropyl-β-cyclodextrin

Red bell pepper carotenoids were complexed with 2-hydroxypropyl-β-cyclodextrin (2-HPβCD) in different mass ratios (1:4, 1:6, 1:8 and 1:10) through ultrasonic homogenization in order to increase carotenoid solubility and their use as natural pigment in food. Inclusion complexes, red bell pepper extract and physical mixtures were analyzed by DSC, FT-IR, 1H NMR and DLS. Solubility assay was performed to identify the effect of complexation on the solubility of carotenoids. From characterization assays, results showed that inclusion process occurred for all tested ratios. Results for water solubility assays demonstrated clear differences between solubility index of inclusion complexes (8.06±2.59–16.55±4.40mg/mL) and physical mixtures (3.53±1.44–7.3±1.88mg/mL), while carotenoid extract was no water soluble, as expected. These results indicated that molecular inclusion of carotenoids in 2-HPβCD was efficient to enhance their solubility in water, enabling application of red bell pepper carotenoid as natural pigment and/or bioactive substances in food.

Comparison of a compatible and an incompatible pepper-tobamovirus interaction by biochemical and non-invasive techniques: Chlorophyll a fluorescence, isothermal calorimetry and FT-Raman spectroscopy

Leaves of a pepper cultivar harboring the L3 resistance gene were inoculated with Obuda pepper virus (ObPV), which led to the appearance of hypersensitive necrotic lesions approx. 72 h post-inoculation (hpi) (incompatible interaction), or with Pepper mild mottle virus (PMMoV) that caused no visible symptoms on the inoculated leaves (compatible interaction). ObPV inoculation of leaves resulted in ion leakage already 18 hpi, up-regulation of a pepper carotenoid cleavage dioxygenase (CCD) gene from 24 hpi, heat emission and declining chlorophyll a content from 48 hpi, and partial desiccation from 72 hpi. After the appearance of necrotic lesions a strong inhibition of photochemical energy conversion was observed, which led to photochemically inactive leaf areas 96 hpi. However, leaf tissues adjacent to these inactive areas showed elevated ΦPSII and Fv/Fm values proving the advantage of chlorophyll a imaging technique. PMMoV inoculation also led to a significant rise of ion leakage and heat emission, to the up-regulation of the pepper CCD gene as well as to decreased PSII efficiency, but these responses were much weaker than in the case of ObPV inoculation. Chlorophyll b and total carotenoid contents as measured by spectrophotometric methods were not significantly influenced by any virus inoculations when these pigment contents were calculated on leaf surface basis. On the other hand, near-infrared FT-Raman spectroscopy showed an increase of carotenoid content in ObPV-inoculated leaves suggesting that the two techniques detect different sets of compounds.

Red pepper (Capsicum annuum) carotenoids as a source of natural food colors: analysis and stability-a review.

Carotenoids are increasingly drawing the attention of researchers as a major natural food color due to their inherent nutritional characteristics and the implicated possible role in prevention and protection against degenerative diseases. In this report, we review the role of red pepper as a source for natural carotenoids. The composition of the carotenoids in red pepper and the application of different methodologies for their analysis were discussed in this report. The stability of red pepper carotenoids during post-harvest processing and storage is also reviewed. This review highlights the potential of red pepper carotenoids as a source of natural food colors and also discusses the need for a standardized approach for the analysis and reporting of composition of carotenoids in plant products and designing model systems for stability studies.

Biochemistry and Molecular Biology of Carotenoid Biosynthesis in Chili Peppers (Capsicum spp.)

Capsicum species produce fruits that synthesize and accumulate carotenoid pigments, which are responsible for the fruits’ yellow, orange and red colors. Chili peppers have been used as an experimental model for studying the biochemical and molecular aspects of carotenoid biosynthesis. Most reports refer to the characterization of carotenoids and content determination in chili pepper fruits from different species, cultivars, varieties or genotypes. The types and levels of carotenoids differ between different chili pepper fruits, and they are also influenced by environmental conditions. Yellow-orange colors of chili pepper fruits are mainly due to the accumulation of α- and β-carotene, zeaxanthin, lutein and β-cryptoxanthin. Carotenoids such as capsanthin, capsorubin and capsanthin-5,6-epoxide confer the red colors. Chromoplasts are the sites of carotenoid pigment synthesis and storage. According to the most accepted theory, the synthesis of carotenoids in chili peppers is controlled by three loci: c1, c2 and y. Several enzymes participating in carotenoid biosynthesis in chili pepper fruits have been isolated and characterized, and the corresponding gene sequences have been reported. However, there is currently limited information on the molecular mechanisms that regulate this biosynthetic pathway. Approaches to gain more knowledge of the regulation of carotenoid biosynthesis are discussed.

Chile Pepper Carotenoids: The Link Between Color and Sub-Cellular Morphology

Extended abstract of a paper presented at Microscopy and Microanalysis 2013 in Indianapolis, Indiana, USA, August 4 – August 8, 2013.

In vitro investigation of the bioaccessibility of carotenoids from raw, frozen and boiled red chili peppers (Capsicum annuum)

Carotenoid-rich foods are associated with antioxidant activity and the ability to alleviate chronic diseases. The present study investigated the effect of processing on the content and bioaccessibility of carotenoids from 13 cultivars of red chili pepper (Capsicum annuum). Carotenoids in chili peppers were analyzed before an in vitro digestion process. The portion of carotenoid transferred to the micelle fraction (bioaccessibility) was also quantified. β-Carotene, β-cryptoxanthin, capsanthin and antheraxanthin were the most abundant carotenoids. Zeaxanthin, violaxanthin, neoxanthin and lutein were detected at lower concentrations. In general, freezing and boiling reduced carotenoid contents. Capsanthin and zeaxanthin had the highest bioaccessibility at an average value from 36 to 40%, followed by antheraxanthin (26%). Bioaccessibility of β-cryptoxanthin, violaxanthin and β-carotene was lower, averaging 6.1, 4.8 and 4.0%, respectively. Neoxanthin and lutein were not detected in micelles. Freezing increased the bioaccessibility of capsanthin, zeaxanthin, antheraxanthin, β-cryptoxanthin and violaxanthin; β-cryptoxanthin bioaccessibility increased and capsanthin and zeaxanthin bioaccessibility decreased following boiling. Differences in the contents and bioaccessibility of carotenoids in 13 C. annuum cultivars and between the processed methods were herein evidenced.

Antioxidant, Antinociceptive, and Anti-Inflammatory Effects of Carotenoids Extracted from Dried Pepper (Capsicum annuumL.)

Carotenoids extracted from dried peppers were evaluated for their antioxidant, analgesic, and anti-inflammatory activities. Peppers had a substantial carotenoid content: guajillo 3406 ± 4 μg/g, pasilla 2933 ± 1 μg/g, and ancho 1437 ± 6 μg/g of sample in dry weight basis. A complex mixture of carotenoids was discovered in each pepper extract. The TLC analysis revealed the presence of chlorophylls in the pigment extract from pasilla and ancho peppers. Guajillo pepper carotenoid extracts exhibited good antioxidant activity and had the best scavenging capacity for the DPPH+ cation (24.2%). They also exhibited significant peripheral analgesic activity at 5, 20, and 80 mg/kg and induced central analgesia at 80 mg/kg. The results suggest that the carotenoids in dried guajillo peppers have significant analgesic and anti-inflammatory benefits and could be useful for pain and inflammation relief.

Effect of heat treatment on the content of some bioactive compounds and free radical-scavenging activity in pungent and non-pungent peppers

Raw and heat-treated (boiled and grilled) pungent and non-pungent peppers (poblano, bell, chilaca, caribe, jalapeño, serrano, habanero and manzano) were evaluated for their content of some bioactive compounds and free radical-scavenging activity. Boiling and grilling were performed under household conditions. Ascorbic acid content in raw peppers varied from 306 to 3438μg/g. This content was reduced 15–87% by heat treatments. Total carotenoids content in raw peppers ranged between 1 and 156μg/g. β-Carotene represented 3–78% of total carotenoids in raw peppers. β-Carotene content was reduced (1–45%) by heat treatments. Free radical-scavenging activity varied widely (7–101μmol TE/g) in raw peppers. Boiling and grilling reduced (6–93%) sequentially the antiradical activity of pungent peppers. In contrast, gradual increases of antiradical activity in non-pungent peppers were induced by boiling and grilling. Household heat treatment altered highly the content of bioactive/antioxidant compounds in tested peppers.

Carotenoid Composition and Vitamin A Value in Ají (Capsicum baccatum L.) and Rocoto (C. pubescens R. & P.), 2 Pepper Species from the Andean Region

The carotenoid patterns of fully ripe fruits from 12 Bolivian accessions of the Andean peppers Capsicum baccatum (ají) and C. pubescens (rocoto) were determined by high-performance liquid chromatography (HPLC)-photodiode array detector (PDA)-mass spectrometry (MS). We include 2 California Wonder cultivars as C. annuum controls. A total of 16 carotenoids were identified and differences among species were mostly found at the quantitative level. Among red-fruited genotypes, capsanthin was the main carotenoid in the 3 species (25% to 50% contribution to carotenoid fraction), although ajíes contained the lowest contribution of this carotenoid. In addition, the contribution of capsanthin 5,6-epoxide to total carotenoids in this species was high (11% to 27%) in comparison to rocotos and red C. annuum. Antheraxanthin and violaxanthin were, in general, the next most relevant carotenoids in the red Andean peppers (6.1% to 10.6%). Violaxanthin was the major carotenoid in yellow-/orange-fruited genotypes of the 3 species (37% to 68% total carotenoids), although yellow rocotos were characterized by lower levels (<45%). Cis-violaxanthin, antheraxanthin, and lutein were the next most relevant carotenoids in the yellow/orange Andean peppers (5% to 14%). As a whole, rocotos showed the highest contributions of provitamin A carotenoids to the carotenoid fraction. In terms of nutritional contribution, both ajíes and rocotos provide a remarkable provitamin A activity, with several accessions showing a content in retinol equivalents higher than California Wonder controls. Furthermore, levels of lutein in yellow/orange ajíes and rocotos were clearly higher than California Wonder pepper (≥1000 μg·100/g). Finally, the Andean peppers, particularly red ajíes, can be also considered as a noticeable source of capsanthin, the most powerful antioxidant compound among pepper carotenoids. Practical Application: Capsicum peppers are known for their content in carotenoids, although there is no information about 2 species with Andean origin: ajíes and rocotos. Due to their relevance for the Andean cuisine and increasing importance in ethnic restaurants in Europe, we studied their carotenoid pattern and vitamin A contribution.

Simultaneous Determination of Sudan Dyes and Carotenoids in Red Pepper and Tomato Products by HPLC

To simultaneously detect Sudan dyes and carotenoids in red pepper and tomato products, high-performance liquid chromatography (HPLC) methods with photodiode-array detection are developed and validated. The methods include the use of end-capped and nonend-capped adsorbents with a gradient elution system starting with water containing methanol. Water content of 9% in the starting mobile phase is found to be necessary to ensure sufficient separation of Sudan dyes and to avoid overlapping or interference with the carotenoids of considerable content. The data of the validation reveal the accuracy and precision of the developed methods. A limit of Sudan dyes detection of 1-5 microg/g in red pepper or tomato sauce could be approached. The methods provide excellent separation of the carotenoids from the unsaponified extracts of red pepper and the tomato products.

Impact of genetic and environmental variation on development of flavonoids and carotenoids in pepper ( Capsicum spp.)

Peppers ( Capsicum spp.) were grown for phytochemical analyses at three different locations including a greenhouse at College Station and field plots at Uvalde and Weslaco, Texas. Cultivar effects were significant at each location for all compounds. The best sources of β-carotene were mature greenhouse-grown fruit of Fidel (23.7 μg/g) and C 127 (22.3 μg/g). Mature greenhouse fruit of Tropic Bell (10.1 μg/g) and PI 357509 (9.2 μg/g) had high lutein, but Uvalde field-grown mature fruit of these lines were low in this compound, (1.4 and 0.5 μg/g, respectively). MJ 201 fruit had the highest zeaxanthin levels (10 μg/g) at both College Station and Uvalde. The best sources of quercetin over all locations were the yellow wax types, Banana Supreme (186 μg/g), PI 357509 (86 μg/g) and Rio Grande Gold (26 μg/g). Fidel (37 μg/g) and Banana Supreme (21.5 μg/g) were the best sources of luteolin. Immature fruit generally contained lower levels of lutein and xeaxanthin than mature, colored fruit. These differences were not always statistically significant. Greenhouse-grown peppers at College Station contained more carotenoids than the field-grown peppers in Uvalde and Weslaco, but there were no significant differences among locations for flavonoid concentrations. Several good candidate parents were identified for the breeding program to develop novel pepper varieties with increased health benefits. Families of these varieties are currently being examined to assess the impact of specific environmental factors and identify genes involved in regulating synthesis of these beneficial phytochemicals.

Composition of Main Carotenoids in Korean Red Pepper (Capsicum annuum, L) and Changes of Pigment Stability During the Drying and Storage Process

ABSTRACT: This study was conducted to identify main carotenoids in red pepper and to investigate pigment stability during drying and storage. The main pigments in saponified extracts were capsanthin, zeaxanthin, β‐cryptoxanthin, β‐carotene, and an unknown peak, which was considered to be capsorubin by spectrophotometry and liquid chromatography/mass spectrometry (LC/MS). Myristoylcapsanthin, lauroylmyristoylcapsanthin, and myristoylpalmitoylcapsanthin were identified from unsaponified extracts analyzing fragment ions by LC/MS. In capsanthin stability, the difference in drying method had the most influence until 2‐mo storage, whereas after 2‐mo storage, temperature became the major factor. Myristoylpalmitoylcapsanthin was more stable than myristoylcapsanthin during storage.