Molecular mechanism of lycopene cyclases regulating carotenoids ratio in different branches during tea leaf and flower development

Carotenoids are essential components in tea quality, contributing to leaf color and aroma. However, little information about carotenoids in different tea cultivars and their biosynthesis regulation mechanism during leaf development is known. Here we analyzed carotenoids by HPLC in the buds and leaves of 113 tea cultivars harvested on the same day. By profile clustering, carotenoids were divided into five groups. Same group cultivars displayed divergence in the total content of carotenoids but a similar molar ratio. To figure out the molecular mechanisms of this phenomenon, we further characterized all functional lycopene cyclases, which are the branch point of the carotenoid biosynthesis pathway. Two β-lycopene cyclases(CsLCYB1 and CsLCYB2) and one ε-lycopene cyclase(CsLCYE1) were cloned. Subcellular localization analysis showed that all cloned CsLCYs were localized in plastids. Enzyme activity assays in E. coli indicated both CsLCYBs catalyzed lycopene into β-carotene, and CsLCYE1 produced δ-carotene and ε-carotene. We found CsLCYB1 and CsLCYE1 predominantly expressed in leaf, while CsLCYB2was mainly expressed during flowering stages. Suppression by antisense oligonucleotides reduced CsLCYB1 and CsLCYE1 transcripts and led to reduction of both β,β-branch and β,ε-branch carotenoids in leaf. The expression levels of CsLCYB1 showed a significant positive correlation withβ,β-branch carotenoids in leaf. Our study provides carotenoid profiles of different tea cultivars, which can assist tea producers in selecting cultivars of interest. Meanwhile, we proposed the molecular mechanism of carotenoids reflecting the tenderness of tea plant leaf from a metabolic flux perspective, and suggested lycopene cyclase that could be applied to the breeding of tea varieties with different branch carotenoids.

Natural variations and geographical distributions of seed carotenoids and chlorophylls in 1167 Chinese soybean accessions

Understanding the composition and contents of carotenoids in various soybean seed accessions is important for their nutritional assessment.This study investigated the variability in the concentrations of carotenoids and chlorophylls and revealed their associations with other nutritional quality traits in a genetically diverse set of Chinese soybean accessions comprised of cultivars and landraces.Genotype,planting year,accession type,seed cotyledon color,and ecoregion of origin significantly influenced the accumulation of carotenoids and chlorophylls.The mean total carotenoid content was in the range of 8.15–14.72μg g–1 across the ecoregions.The total carotenoid content was 1.2-fold higher in the landraces than in the cultivars.Soybeans with green cotyledons had higher contents of carotenoids and chlorophylls than those with yellow cotyledons.Remarkably,lutein was the most abundant carotenoid in all the germplasms,ranging from 1.35–37.44μg g–1.Carotenoids and chlorophylls showed significant correlations with other quality traits,which will help to set breeding strategies for enhancing soybean carotenoids without affecting the other components.Collectively,our results demonstrate that carotenoids are adequately accumulated in soybean seeds,however,they are strongly influenced by genetic factors,accession type,and germplasm origin.We identified novel germplasms with the highest total carotenoid contents across the various ecoregions of China that could serve as the genetic materials for soybean carotenoid breeding programs,and thereby as the raw materials for food sectors,pharmaceuticals,and the cosmetic industry.

Transcriptomes of Litopenaeus vannamei reveal modulation of antioxidant system induced by dietary archaeal carotenoids

Oxidative stress induced by factors such as ammonia nitrogen has become a major issue in shrimp farming.The effects of carotenoids on the growth and antioxidant capability of Litopenaeus vannamei juveniles were investigated in this study using dietary archaeal carotenoids supplementation.For four weeks,shrimp were given diets containing 0 mg/kg(Ctrl)and 55.98 mg/kg(Car)archaeal carotenoids.Dietary archaeal carotenoids significantly enhanced the astaxanthin content in shrimp muscles and carapaces,as well as the superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)activity(P<0.05).The malonaldehyde(MDA)content in Car group significantly decreased(P<0.05).The transcriptome analysis was conducted to determine the molecular processes in response to archaeal carotenoids supplementation.A total of 1536 differentially expressed genes(DEGs)were detected,including 538 upregulated DEGs and 998 downregulated DEGs.GO functional enrichment analysis between Ctrl and Car indicated that 26 GO terms including extracellular region,metabolic process,and proteolysis were enriched.The KEGG pathway enrichment analysis revealed that the amino sugar and nucleotide sugar metabolism,cysteine and methionine metabolism,glycine serine and threonine metabolism,and amino acid biosynthesis were enriched.Archaeal carotenoids influenced the expression of several important genes involved in reactive oxygen species(ROS)generation,Nrf2 signaling,and antioxidant enzymes.Seven DEGs were chosen to confirm the RNA-Seq data using qRT-PCR.The genes and pathways discovered in this work assist to elucidate the molecular processes through which archaeal carotenoid enhances L.vannamei antioxidative system.

Responses of Chlorophyll Fluorescence and Carotenoids Biosynthesis to High Light Stress in Rice Seedling Leaves at Different Leaf Position

In the present study, we investigated the changes of photosynthesis, chlorophyll fluorescence and the content of carotenoid pigments in rice (Oryza sativa L.) seedling leaves and their responses to high light. The results showed that the rate of photosynthesis, the contents of individual and total carotenoids and the pool size of xanthophyll cycle decreased with age increasing of the leaf. When the leaves were exposed to high light for 2 h, the qN of mature leaf (5th leaf) increased more significantly than that of younger (6th leaf) and older leaves (3rd and 4th leaf). Comparing with the leaves before exposure to high light, the excitation pressure on PSⅡ (1- qP ) increased by 44%, 57%, 19% and 45% in the 3rd, 4th, 5th and 6th leaf under high light, respectively. The highest content of carotenoids and the greatest conversion of violaxanthin to zeaxanthin were found in the 5th leaf, and it was consistent with the 5th leaf exhibiting the strongest resistance to high light. Our results suggested that the ability of rice leaf to resist photoinhibition is related to the level of carotenoids and the ability of carotenoids biosynthesis.

Correlation between Carotenoids Content and Several Quality Traits of Wheat Varieties

A total of 31 wheat varieties（lines） released in different years in Huanghuai Area were selected as materials so as to study the carotenoids content,lipoxygenase activity, and some other quality traits of wholemeal. Results showed that carotenoid content and other quality traits had highly significant differences among varieties. Carotenoids content was significantly positive correlated with watersoluble pentosan and yellowness. Carotenoids content was significantly negative correlated with peak viscosity, hold through, breakdown, final viscosity, unit weight,formation time, and was negative correlated with peak time, protein content and stability time. According to cluster analysis, carotenoid contents in 31 varieties were divided into three major groups： one group had common carotenoid content, which included 19 varieties such as Meng 0318; one group had relatively low carotenoid content, which included 10 varieties such as Luomai23; and the last group had relatively high carotenoid content, which included 2 varieties such as Huaimai22. The average carotenoid contents of the three groups were 1.93, 1.07 and 2.99 mg/kg respectively.

Association of Dietary Carotenoids Intake with Skeletal Fluorosis in the Coal-burning Fluorosis Area of Guizhou Province

Objective To explore whether the intake of dietary carotenoids could protect against skeletal fluorosis in Guizhou province in which coal-burning fluorosis is endemic. Methods A case-control study of 196 patients with skeletal fluorosis and 196 age and gender-matched controls was conducted in Zhijin, Guizhou Province. Face-to-face interviews were conducted to assess habitual dietary intake using a 75-item food frequency questionnaire and various covariates with structured questionnaires. Urinary fluoride was measured using an ion-selective electrode method. The genotype of superoxide dismutase 2（SOD2） rs11968525 was detected by Taq Man method. Results We observed significant dose-dependent inverse associations of skeletal fluorosis with intake of β-carotene, lutein/zeaxanthin, lycopene, and total carotenoids（P-trend = 0.002 to 0.018）, whereas α-carotene and β-cryptoxanthin intakes were not found to be related to skeletal fluorosis, after adjustment for potential confounders. The adjusted ORs and 95% CI of skeletal fluorosis for the highest versus lowest quartile were 0.30（0.10, 0.86） for β-carotene, 0.23（0.08, 0.66） for lycopene, 0.26（0.10, 0.75） for lutein/zeaxanthin and 0.34（0.14, 0.74） for total carotenoids（all P-trend ＆lt; 0.05）. Stratified analyses showed that the protective effects of lutein/zeaxanthin and total carotenoids on skeletal fluorosis were more evident for individuals with the AG＋AA genotypes of SOD2（rs11968525）. Conclusion Increased intakes of β-carotene, lutein/zeaxanthin, lycopene, and total carotenoids are independently associated with a lower risk of coal-burning skeletal fluorosis. SOD2（rs11968525） polymorphisms might modify the inverse associations between dietary carotenoids and skeletal fluorosis.

Evaluation of antigenotoxic effects of carotenoids from green algae Chlorococcum humicola using human lymphocytes

Objective:To identify the available phytochemicals and carotenoids in the selected green algae and evaluate the potential genotoxic/antigenotoxic effect using lymphocytes.Methods:Organic,solvent extracts of Chlorococcum humicola(C.humicola)were used for the phytochemical analysis.The available carotenoids were assessed by HPLC,and LC-MS analysis.The genotoxicity was induced by the benzo(a)pyrene in the lymphocyte culture,the genotoxic and antigenotoxic effects of algal carotenoids with and without genotoxic inducer were evaluated by chromosomal aberration(CA),sister chromatid exchange(SCE)and micronucleus assay(MN).Results:The results of the analysis showed that the algae were rich in carotenoids and fatty acids.In the total carotenoids lutein,β-carotene andα-carotene were found to be present in higher concentration.The frequency of CA and SCE increased by benzo(a)pyrene were significantly decreased by the carotenoids(P<0.05 for CA,P<0.001 for SCE).The MN frequencies of the cells were significantly decreased by the treatment with carotenoids when compared with the positive controls(P<0.05).Conclusions:The findings of the present study demonstrate that,the green algae C.humicola is a rich source of bioactive compounds especially carotenoids which effectively fight against environmental genotoxic agents,the carotenoids itself is not a genotoxic substance and should be further considered for its beneficial effects.

Study on Extraction Method of Carotenoids from Citrus Leaves

In order to explore the extraction method of carotenoids from citrus leaves,the extraction effect of carotenoids from leaves of Honey pomelo by ultrasonic-assisted extraction was investigated by single factor experiment and orthogonal experiment.The results showed that the optimum extraction condition was:the ration of ethanol and acetone 1∶1(v/v,containing 0.1%BHT),solid-liquid ratio 1∶30(g/mL),extraction time 7 min,ultrasonic power 240 W,extraction frequency 3 times.Under the above condition,the extraction rate of carotenoids was 85.98%.

Quantitative Structure-activity Relationship Study on the Antioxidant Activity of Carotenoids

Carotenoids are a family of effective active oxygen scavengers, which can reduce the danger of occurrence of chronic diseases such as cardiovascular disease, cataract, cancer, and so on. The quantitative structure-activity relationship （QSAR） equation between carotenoids and antioxidant activity was established by quantum chemistry AM1, molecular mechanism （MM＋） and stepwise regression analysis methods, and the model was evaluated by leave-one-out approach. The results showed that the significant molecular descriptors related to the antioxidant activity of carotenoids were the energy difference （E_HL） between the lowest unoccupied molecular orbital （LUMO） and the highest occupied molecular orbital （HOMO） and ionization energy （Eiso）. The model showed a good predictive ability （Q^2 〉 0.5）.

Some Factors Affecting the Production of Carotenoids by <i>Rhodotorula glutinis</i>var. <i>glutinis</i>

A new yeast strain isolated from pin cushion flower (Scabiosa atropurpura) in our laboratory was selected from 200 yeast isolates as carotenoids producer and identified as Rhodotorula glutinis var. glutinis. The selected isolate was grown in synthetic medium to study the effect of carbon to nitrogen ratio, sources of nitrogen and carbon, mineral salts and incubation temperature on carotenoids production. The results indicated the following optimal conditions: carbon to nitrogen ratio of 5, ammonium sulphate as nitrogen source, sucrose as carbon source, presence of zinc sulphate in the medium and cultivation temperature of 25?C. The studied factors affected the dry biomass as well as the proportion of carotenoids and consequently the colour of pellets of the yeast. The yeast strain was grown under the optimal conditions to study the changes occurring in the medium and the pellets during carotenoids production for 6 days. Carotenoids production started after the first day of incubation and most of the carotenoids content in the yeast cells was produced during stationary phase. The highest cellular (861 μg?g–1) and volumetric (1.9 mg?L–1) carotenoids content were ob- tained after 5 days of growth.

Genetic dissection of carotenoids in maize kernels using high-density single nucleotide polymorphism markers in a recombinant inbred line population

Carotenoids are antioxidants and vitamin A precursors that have important roles in human health. Hence, improving the carotenoid contents in maize kernels is a priority objective for breeders in order to obtain nutritional biofortification outcomes. In the current study, the genetic architecture of carotenoids in maize kernels was explored using a recombinant inbred line(RIL) population derived from a cross between inbred lines By804 and B73. A total of 81 QTLs were detected by using a high-density bin map and a simple sequence repeat(SSR)-based linkage map, with one to seven QTLs for each trait explaining 4.21%–47.53% of the phenotypic variation. A comparison of the QTL mapping efficiency between the two linkage maps revealed that the high-density bin map had higher resolution. In the current study 46 additional QTLs were identified, with 16 being common with previous studies and14 newly identified. Among the results, 29.6%(24/81) of QTLs explained > 10% of the phenotypic variation in the RIL population, and 70.4%(57/81) explained ≤ 10%. These results suggest that a few large-effect QTLs, together with a variable number of minor-effect QTLs,contributed to most of the genetic components of carotenoids in maize kernels.

Cloning and Characterization of the Phytoene Desaturase(pds) Gene-a Key Enzyme for Carotenoids Synthesis in Dunaliella(Chlorophyta)

The unicellular green alga Dunaliella is outstanding for its ability of massive accumulation of carotenoids. To elucidate the carotenoids synthesis pathway in this alga,phytoene desaturase(pds) gene cDNA together with its DNA sequences were isolated and their structures and functions analyzed. The full-length pds cDNA of 2290 bp(GenBank Accession No. DQ243892) was de-duced from RACE results,including untranslated 21 bp 5'-and 520 bp 3'-flanking regions and an open reading frame of 582 amino acids,coding a protein of 64.196 kDa. The DNA sequence of 2908 bp(GenBank Accession No. DQ845248) including five introns was obtained. The fifth intron was uncompleted and complex,including two bases' perfect repeats(GT) 10 and large different-sized repeats within the last 400 bp. The Southern blot hybridization result demonstrated that this gene occurred as a single copy in this species,and the quantitative RT-PCR result showed that the transcription of this gene was constitutive. The evolutional significance of pds was discussed.

Optimization of solvent extraction conditions for total carotenoids in rapeseed using response surface methodology

The optimum total carotenoids (TC) extraction from rapeseed with solvent extraction method by UV-visible spectrophotometer determination was investigated by using response surface methodology (RSM). Extraction duration, re-peated extraction cycles, solvent-solid ratio and extraction temperature were assumed to be the most important factors affecting solvent extrac-tion for the determination of TC. Optimum sol-vent extraction conditions for maximizing the determination of TC were: extraction duration 7.3h, repeated extraction three times, ratio of solvent-solid (v/w, mL/mg) 29:1, extraction temperature 42&amp;#176;C. Under the optimal conditions, the yield of TC was up to 4.79 mg /100g. The model had a satisfactory coefficient of R2 (= 0.912) and verified experimentally. The results showed that the conditions were mild and use-ful for maximizing a quantitative spectropho-tometer determination of TC in rapeseed.

Quantitative Structure-activity Relationship Studies on the Antioxidant Activity and Gap Junctional Communication of Carotenoids

The antioxidant and gap junctional communication（GJC） activities of carotenoids are known to be the two main anticancer mechanisms.Quantitative structure-activity relationship（QSAR） models of the two activities were developed using stepwise regression and multilayer perceptron neural network based on the calculated descriptors of quantum chemistry.The results showed that the significant molecular descriptor related to the antioxidant activity of carotenoids was the HOMO-LUMO energy gap（EHL） and the molecular descriptor related to the GJC was the lowest unoccupied molecular orbital energy（ELUMO）.The two models of antioxidant activity both showed good predictive power,but the predictive power of the neural network QSAR model of antioxidant activity was better.In addition,the two GJC models have similar,moderate predictive power.The possible mechanisms of antioxidant activity and GJC of carotenoids were discussed.

Determination of Major Carotenoids in Processed Tropical Leafy Vegetables Indigenous to Africa

Tropical leafy-vegetables (n = 21) indigenous to Cameroon, Africa, were collected, processed, and analyzed for carotenoids by HPLC. The processing techniques used were oven drying;sun-drying;squeeze-washing and boiling;and a combination of boiling in alkaline salt and squeeze-washing. Carotenoids included lutein, α-carotene, zeaxanthin, β-cryptoxanthin, and β-carotene (all-trans, 13-cis, and 9-cis), which varied by species (P 40% of total carotenoids. Traditional preparation and processing procedures led to significant losses of carotenoids and β-carotene was most affected during sun-drying with a maximum of 73.8% loss observed in A. acanthochiton.

Extraction of Carotenoids and Fatty Acids from Microalgae Using Supercritical Technology

The work described here is based on a comparative study of carotenoids and fatty acids extracted from Synechococcus sp. with (1) pure supercritical CO2, (2) CO2 with 5% (v/v) ethanol as cosolvent and (3) ultrasound-assisted extraction using N, N-dimethylformamide (DMF). The effects of extraction conditions on supercritical CO2 extraction with and within cosolvent were analyzed at different temperatures (40℃, 50℃ and 60℃) and pressures (200, 300 and 400 bars). SFE with CO2 proved to be the most selective method for the extraction of β-carotene, but under these conditions the contents of zeaxanthin and fatty acids were only comparable to or lower than those obtained with techniques that use SFE cosolvent. The SFE technique with CO2 and ethanol simultaneously extracted β-carotene and zeaxanthin and not only increased the concentrations of fatty acids obtained, but also helped to remove fatty acids (palmitoleic and linolenic acid) that were not obtained with pure CO2. Comparison of the supercritical technology with the ultrasound-assisted extraction (UAE) shows that the former technique is the most appropriate due to the fact that ethanol is generally regarded as a safe solvent in comparison to DMF.

Enhanced Production of Natural Carotenoids from Genetically Engineered Rhodobacter sphaeroides Overexpressing CrtA

Carotenoids act as precursors of vitamin A,antioxidants,enhancers of immunity,and are thus widely used in food and pharmaceutical industry.Microbial fermentation is one of the most important solutions for production of natural carotenoids.Rhodobacter sphaeroides is one of most promising bacteria employed for large scale production of carotenoids.In the present study,crtA located in the carotenoids biosynthesis pathway in R.sphaeroides was amplified by PCR.The overexpression vector pRKcrtA was constructed and subsequently transferred into R.sphaeroides,producing the genetically engineered strain R.sphaeroides 2.4.1/pRKcrtA overexpressing crtA.The carotenoid production from the genetically engineered strain was significantly increased.Fermentation procedure was optimized for further enhanced carotenoids production.

Polyphenols,Ascorbic Acid and Carotenoids Contents and Antioxidant Properties of Habanero Pepper(Capsicum chinense)Fruit

Their high bioactive compounds content and importance as dietary antioxidants has increased interest in Capsicum fruit. The fruit of seven Capsicum chinense Jacq. var. habanero genotypes grown in Yucatan, Mexico, were analyzed to quantify their phenolic compounds, carotenoids and ascorbic acid contents, and to measure their free radical scavenging (ABTS assay) and antioxidant activities (β-carotene/linoleic acid assay). Phenolics (20.54 to 20.75 mg/100 g sample), carotenoids (1.00 to 1.26 mg/100 g sample) and ascorbic acid contents (187.24 to 281.73 mg/100 g sample) varied between genotypes. Trolox equivalent antioxidant capacity (TEAC) ranged from 1.55 to 3.23 mM/mg sample. During the 120-min decolorization trial, antioxidant capacity decreased over time in the studied genotypes. Values ranged from 36%to 57% β-carotene bleaching during the first 30 minutes. Fruit from all seven studied genotypes are good antioxidant sources and hold promise as natural ingredients in functional foods.

Valuable Biomolecules from Nine North Atlantic Red Macroalgae: Amino Acids, Fatty Acids, Carotenoids, Minerals and Metals

In modern society, novel marine resources are scrutinized pursuing compounds of use in the medical, pharmaceutical, biotech, food or feed industry. Few of the numerous marine macroalgae are currently exploited. In this study, the contents of nutritional compounds from nine common North Atlantic red macroalgae were compared: the lipid content was low and constant among the species, whereas the fatty acid profiles indicated that these species constitute interesting sources of polyunsaturated fatty acids (PUFA). The dominating essential and non-essential amino acids were lysine and leucine, aspartic acid, glutamic acid, and arginine, respectively. The amino acid score of the nine algae varied from 44% to 92%, the most commonly first limiting amino acid being histidine. Lutein, β-carotene, and zeaxanthin were the identified carotenoids. Contents of all macro and trace minerals, with the exception of phosphorus, were higher than those described for conventional food. Low sodium/potassium ratios (0.08 - 2.54) suggested a potential for using the ash fraction for sodium salt replacement. The algae constituted rich sources of carbohydrates (40% to 71% of DM) which show their potential for a broader commercial exploitation. In some species, the concentrations of arsenic, cadmium, and lead exceeded limit values for application in food or feed. In conclusion, the nine algae represent promising potential sources of health promoting additives for human and animal diets, in whole or in a biorefinery concept.

Genetics of carotenoids for provitamin A biofortification in tropical-adapted maize

Yellow maize contains high levels of β-carotene(βC), making it an important crop for combating vitamin A deficiency through biofortification. In this study, nine maize inbred lines were selected at random from 31 provitamin A(PVA) maize inbred lines and crossed in a partial diallel mating design to develop 36 crosses. The crosses were evaluated in the field in two locations(Samaru and Kerawa) and their seed carotenoid content were determined by high-performance liquid chromatography. The modes of gene action, heritability, and correlations between agronomic traits and carotenoid content were estimated. Additive genetic variances(σ~2a) were lower than non-additive genetic variances(σ~2d) for all the carotenoids, plant height(PH), and grain yield(GY), suggesting a preponderance of non-additive gene action. Broad-sense heritability(H^2) was high(H^2> 60%) for zeaxanthin,days to anthesis, and PH, moderate(30% < H^2< 60%) for lutein and GY, and low(H^2< 30%)for alpha carotene, beta cryptoxanthin, βC, and PVA. Genetic advance as a percentage of mean, considered with H^2, also suggests a preponderance of non-additive gene action for PVA carotenoids. Hybrid variety development is thus an appropriate approach to improving grain yield and PVA. GY showed no significant genotypic correlations with carotenoid content, suggesting that these traits can be improved concurrently. Thus, there is ample scope for improvement of PVA and GY in the sample of tropical-adapted maize.