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The color of the maize (Zea mays L.) grain is determined by the content and composition of carotenoids, including provitamin A, which is a product of the β-β (β-carotene, β-cryptoxanthin) and β-ɛ (α-carotene) branches of carotenogenesis. The ratio of the fluxes of the branches depends on the activity of the lycopene-ɛ-cyclase LcyE, which determines the β-ɛ branch. In this work, we analyzed allelic variants of the LcyE gene, which are potentially effective for increasing the biosynthesis of β-carotene, in 20 maize inbred lines of domestic selection, which differ in grain color. The 5'-UTR regions of the LcyE gene were amplified and sequenced. Fragment analysis showed the presence of allele 2 in four lines and the new allele 5 in 16 lines. The polymorphism of the new allele 5 was characterized—four single nucleotide polymorphisms and two deletions. The comparison of cis-regulatory elements in the analyzed 5'-UTR region in alleles 2 and 5 revealed a difference in binding sites with transcription factors. Expression of the LcyE gene was determined in the leaves of two lines with allele 2 and three lines with allele 5. A direct relationship was shown between the presence of allele 5 and a decrease in gene expression: the level of gene transcription in the case of allele 2 was 10–15 times higher than in the case of the allele 5. It has been suggested that the presence of allele 5 of the LcyE gene in the maize genome correlates with a decrease or suppression of the LcyE expression and, with stable activity of other carotenogenesis enzymes, with grain color. The use of allele 5 donors in combination with a known dark yellow or orange color of the grain can be used in the breeding of maize with increased synthesis of provitamin A in the grain. [ABSTRACT FROM AUTHOR]

The carotenoids available in food are vital dietary micronutrients for human health. Plants synthesize and accumulate different carotenoids in plastids in a tissue-specific manner. The level of β-carotene (provitamin A) and other nutritionally important carotenoids is substantially low in the green tissues such as leaves compared to the fruits and roots. In photosynthetic tissues, chloroplasts can accumulate a moderate level of carotenoids, mainly to facilitate photosynthesis and environmental stress tolerance. However, chromoplasts from the storage tissues such as tomato fruit and carrot root can synthesize and accumulate carotenoids to a substantially higher level. A synthetic biology approach that utilizes a transient expression of bacterial phytoene synthase (crtB) gene in the photosynthetic leaves can induce the transition of chloroplasts into chromoplasts. The plastid-localized heterologous expression of crtB in leaves can induce the overaccumulation of phytoene, triggering the chloroplast-to-chromoplast transition; therefore, enhancing the biosynthesis and accumulation of carotenoids, including provitamin A. The transition of chloroplasts into chromoplasts, however, altered the photosynthetic thylakoids, consequently reducing the photosynthetic efficiency and plant growth. An efficient metabolic engineering strategy is desirable to enhance the production of targeted carotenoids in leaves without perturbing the photosynthetic efficiency and plant growth. Collectively, a synthetic biology strategy that triggers the transformation of chloroplasts into chromoplasts in photosynthetic tissues unfolds new avenues for carotenoid biofortification in the leafy food and vegetable crops, which can increase the dietary intake of carotenoids, therefore, combating the crisis of vitamin A deficiency. [ABSTRACT FROM AUTHOR]

The cognition of original qualitative attributes of native Peruvian chili peppers elucidate their potential industrial exploitation. There is a lack of scientific information about chili pepper physicochemical and nutritional attributes, that hampers their immediate industrial use. Therefore, the aim of the present study was to characterize twenty Peruvian chili pepper landraces in terms of structural and nutritional properties. In particular, the Bostwick consistency index of chili purées, dry matter, dietary fibers and pectin content were determined. In addition, the color and carotenoid profile of each landrace were studied. A wide range of consistency and stability were found, and partially explained by the dry matter, dietary fiber and pectin content. C. annuum landraces displayed the highest dry matter and fiber content. Moreover, different carotenoid profiles were observed, being β-carotene the most abundant in almost all the samples. Capsanthin was present only in red landraces, while yellow ones had low total carotenoid content. High Retinol Activity Equivalent levels were determined for red peppers, while high insoluble dietary fiber level was found in all the samples. Cluster analysis, including all the chili pepper attributes, showed no relation between taxonomical classification and chili peppers structural, sensorial, and nutritional characteristics. Unique features of studied chili pepper landraces presented in this work should be considered during their industrial processing. In this context, these results might be considered a starting point for processing design and for the proper valorization of this raw material. [ABSTRACT FROM AUTHOR]

Abstract Poverty eradication and global food security are among the targets of world leaders, most especially combating the scourge of hidden hunger. Provitamin A carotenoids cannot be synthesized de novo by human and so it must be taken as part of the diet. The deficiency of which is causing almost 6000 sights to be lost daily in most developing countries because of the monotonous starchy diets lacking substantial amount of carotenoid. Conventional breeding as well as genetic engineering have been used to increase the level of carotenoid in many staples including rice, potato, maize and cassava. While products from genetic engineering are still been subjected to strict regulatory measures preventing the delivery of the products to target consumers, some of the products from conventional breeding are already on the table of consumers. Interestingly, both technologies are crucial to tackling micronutrient deficiencies. This review discusses the role of carotenoid in human, the biosynthesis in plant and some of the staple crops that have been modified for increased carotenoid. Some measures expected of the leaders of the countries in need of these products for safe delivery to the target population after two decades is also highlighted. [ABSTRACT FROM AUTHOR]

Vitamin A deficiency is a widely prevalent health disorder among millions of people worldwide. Introgression of crtRB1 and lcyE favourable alleles that enhance concentration of provitamin A in maize endosperm have been employed in maize biofortification programmes. To make marker-assisted selection (MAS) more effective, we have developed rapid and convenient multiplex-polymerase chain reaction (PCR) assay to simultaneously discover the allelic combinations among the segregants. Validation of the multiplex assay was done in two backcross-derived populations developed using elite inbreds viz., HKI193-1 and HKI193-2 carrying unfavourable alleles of crtRB1 (296 bp) and lcyE (300 bp) and HarvestPlus inbreds viz., HP704-22 and HP704-23 possessing favourable alleles of crtRB1 (543 bp) and lcyE (650 bp). We also standardized the uniplex-PCR assays for both the genes that gave robust and reproducible results in sub-tropical populations. Gel profiles of BC1F1, BC2F1 and BC2F2 revealed that these assays identified the backcross progenies homo-or hetero-zygous for the favourable- or unfavourable-alleles. Multiplex-PCR assay also precisely confirmed the results of individual uniplex assays in different backcross generations. Cost and time analyses showed that multiplex-PCR assay has potential to save 41% of cost, and 50% of time compared to two uniplex assays in a MAS programme. It has also saved 50% of the manpower. The multiplex assay possesses significant advantage over uniplex assays and enhances the efficiency of selection. This is the first report of development and validation of multiplex-PCR assay of crtRB1 and lcyE for utilization in maize biofortification programme. [ABSTRACT FROM AUTHOR]

Provitamin A (proVA) carotenoids are converted into retinol (vitamin A) in the human body, are the subject of human nutrition studies, and are targets for biofortification of staple crops. β-Carotene has been the principal target for enhancing levels of proVA. There is recent interest in enhancing the proVA carotenoid β-cryptoxanthin since it has excellent bioavailability, and in maize may be nearly as effective as β-carotene in providing retinol to humans. This study was designed to enhance our understanding of the genetic control of: levels of β-cryptoxanthin, conversion of β-carotene into β-cryptoxanthin and zeaxanthin, conversion of β-cryptoxanthin into zeaxanthin, and flux into and within the β-branch of carotenoid pathway. A biparental population derived from two inbreds with relatively high levels of β-cryptoxanthin and different ratios of β-carotene to β-cryptoxanthin and β-cryptoxanthin to zeaxanthin was studied. Three field replications of this F population were grown, grain analyzed by liquid chromatography (LC), and composite interval mapping (CIM) performed to identify 90 quantitative trait loci (QTL) for carotenoids. We detected QTL for β-carotene/(β-cryptoxanthin + zeaxanthin) and (β-carotene + β-cryptoxanthin)/zeaxanthin ratios that contain candidate gene hydroxylase 4 ( hyd4), which has not been previously associated with QTL for carotenoids in maize grain. Two color assessment methods, visual score and chromameter reading, were used to phenotype one replicate of the population for initial assessment as simple alternative measuring procedures. A common finding for LC and chromameter analysis included QTL on chromosome 5 that contain candidate gene lycopene β cyclase ( lcyβ). [ABSTRACT FROM AUTHOR]

Vitamin A deficiency caused by insufficient intake of β-carotene has emerged as one of the most important problems afflicting people worldwide. Traditional yellow maize is predominated by lutein and zeaxanthin; but contains low β-carotene par below the daily requirement for humans. A natural variant of crtRB1 allele increases β-carotene in kernel by blocking its conversion to zeaxanthin. In the present study, genetic diversity analyses were carried out among 24 diverse maize inbreds possessing rare allele of crtRB1. The mean β-carotene among the inbreds was 9.3 μg/g, with 14 inbreds having 9.1-18.8 μg/g. Among the inbreds of exotic origin, HP704-23, HP704-22, HP467-15, HP465-41 and HP467-20 were promising with high β-carotene. In case of inbreds developed in India, VQL-2-PV, VQL1-PV, V345-PV, V335-PV, HKI161-PV, MGU-PV-2, HKI1105-PV, HKI323-PV and MGU-PV-3 were identified to possess high kernel β-carotene. These inbreds with rare allele of crtRB1 are unique germplasm and therefore holds immense promise in the biofortification programme worldwide. Molecular profiling of these inbreds using 65 SSRs distributed throughout the genome generated 268 alleles with a mean of 4.12 alleles per locus. The polymorphism information content varied from 0.21 to 0.82 with a mean of 0.58. The study detected 12 each of unique and rare alleles. Genetic dissimilarity ranged from 0.40 to 0.94 with an average of 0.79. Cluster analyses grouped 24 genotypes into four major clusters, and principal coordinate analysis depicted the diverse nature of the genotypes consistent with their pedigree. The study identified potential hybrid combinations for higher β-carotene that can be directly utilized in the biofortification programme. [ABSTRACT FROM AUTHOR]

Quality protein maize (QPM; Zea mays L.) has effectively enhanced levels of the amino acids, lysine, and tryptophan, over normal maize and provided balanced dietary protein for the health and development of monogastric animals and humans. However, as in normal maize, QPM varieties are low in provitamin A (ProVA), a precursor of vitamin A, which can lead to vitamin A deficiency in humans when maize is a significant part of their diet. In this study, maize inbred Hp321-1 carrying the favorable alleles crtRB1- 5′TE- 2 and crtRB1- 3′TE- 1 that can enhance levels of ProVA, was used as donor for improving ProVA in QPM inbred lines CML161 and CML171. Functional markers for identifying the favorable alleles crtRB1- 5′TE- 2 and crtRB1- 3′TE- 1 were used in foreground selection, and simple sequence repeat markers were used in background selection for the BCF, BCF, and BCF generations. The background recovery rates were 77.4 and 84.5 % for CML161 and CML171 populations, respectively, in the BCF generation, and 89.9 and 92.1 % in the BCF generation. With foreground and background selection, the mean ProVA concentration has been improved from 1.60 µg g in the parent of CML161 to 5.25 µg g in its BCF offspring, from 1.80 µg g in the parent of CML171 to 8.14 µg g in its BCF offspring while maintaining similar QPM characteristics of the recurrent parents. The success from this study offers maize breeders a procedure for increasing ProVA in QPM lines, which will greatly mitigate vitamin A deficiency and protein-energy malnutrition in developing countries. [ABSTRACT FROM AUTHOR]

Main conclusion: Down-regulation of lipoxygenase enzyme activity reduces degradation of carotenoids of bio-fortified rice seeds which would be an effective tool to reduce huge post-harvest and economic losses of bio-fortified rice seeds during storage. Bio-fortified provitamin A-enriched rice line (golden rice) expressing higher amounts of β-carotene in the rice endosperm provides vitamin A for human health. However, it is already reported that degradation of carotenoids during storage is a major problem. The gene responsible for degradation of carotenoids during storage has remained largely unexplored till now. In our previous study, it has been shown that r9- LOX1 gene is responsible for rice seed quality deterioration. In the present study, we attempted to investigate if r9- LOX1 gene has any role in degradation of carotenoids in rice seeds during storage. To establish our hypothesis, the endogenous lipoxygenase (LOX) activity of high-carotenoid golden indica rice seed was silenced by RNAi technology using aleurone layer and embryo-specific Oleosin-18 promoter. To check the storage stability, LOX enzyme down-regulated high-carotenoid T transgenic rice seeds were subjected to artificial aging treatment. The results obtained from biochemical assays (MDA, ROS) also indicated that after artificial aging, the deterioration of LOX-RNAi lines was considerably lower compared to β-carotene-enriched transgenic rice which had higher LOX activity in comparison to LOX-RNAi lines. Furthermore, it was also observed by HPLC analysis that down-regulation of LOX gene activity decreases co-oxidation of β-carotene in LOX-RNAi golden rice seeds as compared to the β-carotene-enriched transgenic rice, after artificial aging treatment. Therefore, our study substantially establishes and verifies that LOX is a key enzyme for catalyzing co-oxidation of β-carotene and has a significant role in deterioration of β-carotene levels in the carotenoid-enriched golden rice. [ABSTRACT FROM AUTHOR]

Provitamin A-biofortified maize may contribute to alleviating vitamin A deficiency (VAD), in developing countries. However, processing the maize into food products may reduce its provitamin A content. The aims of this study were to determine the composition of provitamin A carotenoids in biofortified maize varieties as well as to assess their retention during processing of popular maize foods consumed in KwaZulu-Natal, South Africa. The non-provitamin A carotenoid, zeaxanthin and the provitamin A carotenoids, β-cryptoxanthin, and trans and cis isomers of β-carotene, and other unidentified trans and cis isomers of β-carotene were detected in varying concentrations in the maize. Milling provitamin A-biofortified maize into mealie meal resulted in a higher retention of carotenoids compared to milling into samp. The highest retention of provitamin A carotenoids was observed in cooked phutu and cooked samp, whilst cooking into thin porridge resulted in the lowest retention of provitamin A carotenoids. In phutu, 96.6 ± 20.3% β-cryptoxanthin and 95.5 ± 13.6% of the β-carotene were retained after cooking. In samp, 91.9 ± 12.0% β-cryptoxanthin and 100.1 ± 8.8% β-carotene; and in thin porridge, 65.8 ± 4.6% β-cryptoxanthin and 74.7 ± 3.0% β-carotene were retained after cooking. This study demonstrates that provitamin A retention in maize is affected by the cooking method (and hence cooked food form) and therefore cooking methods that result in a good retention of provitamin A need to be identified and recommended. [ABSTRACT FROM AUTHOR]

Carotenoid β-hydroxylases attach hydroxyl groups to the β-ionone rings (β-rings) of carotenoid substrates, resulting in modified structures and functions of carotenoid molecules. We cloned and characterized two genes (each with three homeologs), HYD1 and HYD2, which encode β-hydroxylases in wheat. The results from bioinformatic and nested degenerate PCR analyses collectively suggest that HYD1 and HYD2 may represent the entire complement of non-heme di-iron β-hydroxylases in wheat. The homeologs of wheat HYDs exhibited major β-ring and minor ε-ring hydroxylation activities in carotenoid-accumulating E. coli strains. Distinct expression patterns were observed for different HYD genes and homeologs in vegetative tissues and developing grains of tetraploid and hexaploid wheat, suggesting their functional divergence and differential regulatory control in tissue-, grain development-, and ploidy-specific manners. An intriguing observation was that the expression of HYD1, particularly HYD- B1, reached highest levels at the last stage of tetraploid and hexaploid grain development, suggesting that carotenoids (at least xanthophylls) were still actively synthesized in mature grains. This result challenges the common perception that carotenoids are simply being turned over during wheat grain development after their initial biosynthesis at the early grain development stages. Overall, this improved understanding of carotenoid biosynthetic gene expression and carotenoid metabolism in wheat grains will contribute to the improvement of the nutritional value of wheat grains for human consumption. [ABSTRACT FROM AUTHOR]

Adequate consumption of carotenoids including lycopene, β-carotene, lutein, zeaxanthin, and astaxanthin have many benefits for human health. In plants, carotenoids are derived from isoprenoid precursors from the 2- C-methyl- d-erythritol 4-phosphate (MEP) pathway located in plastids. The MEP pathway is also required for the biosynthesis of chlorophyll, terpenoids, plant hormones, and other metabolites. Despite its complexity and difficulty, various strategies have been successfully used to improve the carotenoid biosynthesis in plants through metabolic engineering. Here, these metabolic engineering strategies are reviewed. In addition, the development of gene stacking technologies for carotenoid biosynthesis is evaluated. These technologies will expedite our efforts to bring the health benefits of carotenoids and other nutritional compounds to our diet. [ABSTRACT FROM AUTHOR]

Echinenone has been used as an edible orange pigment, antioxidant and provitamin A. An echinenone-accumulating strain, BOT-20, of Botryococcus braunii was isolated from freshwater environments in Japan. The B. braunii BOT-20 strain is different from other strains of B. braunii, as it appeared dark red during its growth in the laboratory culture as opposed to green. The biomass of the strain was 1.9 g L at 1 month after cultivation. The n-hexane/acetone (3:1, v/v) extract of the strain was 45.5% of the dry biomass weight and consisted of carotenoids (92%, of which 73% was echinenone) and hydrocarbons (8%). The echinenone content was 30.5% of the dry biomass weight, and production was 630 mg L. Hydrocarbons comprised only 3.7% of the total dry biomass weight. The main component of hydrocarbon was an analogue of botryococene by H and C NMR. With high values of echinenone content and production, the B. braunii strain BOT-20 is expected to be a new bioresource for the commercial production of echinenone. [ABSTRACT FROM AUTHOR]

Vitamin A deficiency causes xerophthalmia in preschool-aged children worldwide. The objective of this study was to estimate the genetic parameters that would be useful in selecting parent plants for developing productive hybrids with higher levels of provitamin A in the maize kernel. A complete 7 × 7 diallel mating scheme was used to generate 21 single-cross hybrids. The F crosses and check hybrids were evaluated in complete block design across three different Brazilian environments, and carotenoid content was analyzed by high performance liquid chromatography. General combining ability effects were significant for all traits except α-carotene This result indicates that the contribution of the additive effect was more important for provitamin A and other carotenoids and, consequently that there is high chance of improving this trait through recurrent selection methods. Line 3 produced the highest level of kernel provitamin A among the inbred lines evaluated and also demonstrated the potential to contribute to the development of genetic materials with a good performance for provitamin A. Inbred lines 1, 6, and 7 showed a higher concentration of favorable alleles for grain yield, and inbred lines 3 and 6 exhibited a higher concentration of favorable alleles for β-carotene. Hybrid 1 × 3 performed well in terms of provitamin A and grain yield and combination 2 × 3 was the best performer in terms of lutein content. [ABSTRACT FROM AUTHOR]

Rice the major staple food crop which feeds more than half of the world's population but, lacks pathway to synthesize and accumulate provitamin A in endosperm therefore rice eaters particularly children, and pregnant women suffer due to vitamin A deficiency. The pathway for provitamin A synthesis in rice endosperm has been engineered and transgenic rice lines have yellow endosperm, called 'Golden Rice'. The present study aimed at studying the inheritance of transgene(s) in six transgenic events of 'Golden Rice' and transfer of provitamin A trait from transgenic lines to a widely grown mega rice variety Swarna. The events E1, R1 and W1 showed normal Mendelian inheritance in F, BCF and BCF generations. The event W1 was studied in BCF as well and showed normal Mendelian inheritance of 3:1. The inheritance pattern in L1 event in BCF and BCF showed normal Mendelian inheritance following expected ratio 1:1 and 3:1 respectively. The two events G1 and T1 showed distorted segregation in BCF and BCF respectively in Swarna genetic background. In G1 event, transgene inheritance showed segregation distortion in BCF in favour of transgene negative plants. In T1 event, inheritance followed expected Mendelian segregation in BCF, BCF and BCF, generations. However, when tested against co-dominant inheritance 1:2:1 pattern in BCF, segregation distortion was observed with less than the expected transgene homozygotes. While against 3:1 ratio, it showed the expected segregation pattern in BCF generation. Segregation distortion probably due to differential transmission of transgene positive/negative gametes through either/both parents which needs further study. [ABSTRACT FROM AUTHOR]

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A total of 206 residents (76 males and 130 females) of a rural area of Hokkaido, Japan, attending a health check in August, 1997, were studied to assess the relationship between serum carotenoids and atrophic gastritis (AG). Of the participants, 91 had AG as indicated by their serum levels of pepsinogen I and pepsinogen II. Logistic regression analysis, after adjusting for gender and age, revealed that the odds ratios for serum carotenoid levels were lower for subjects with high serum levels of α-carotene (odds ratio, 0.41; 95% C.I., 0.19–0.88) and β-carotene (odds ratio, 0.41; 95% C.I., 0.18–0.91) than for those with low serum carotenoid levels. In addition, the odds ratios of subjects with high serum levels of β-cryptoxanthin (odds ratio, 0.60; 95% C.I., 0.28–1.31), provitamin A (odds ratio, 0.38; 95% C.I., 0.17–0.85), and retinol (odds ratio, 0.67; 95% C.I., 0.31–1.48) were found to be lower than the odds ratios for those with low serum levels. Odds ratios for subjects with high serum zeaxanthin/lutein levels were higher than odds ratios for those with low serum levels. These results suggest that frequent intake of foods ric, in carotenoids with provitamin A activity may reduce the risk of AG. [ABSTRACT FROM AUTHOR]

Some locally available fruits and vegetables of the arid zone of Rajasthanwere analyzed for their β-carotene content; the selected fruits and vegetables were: Dansra ( Rhus myserensis), Kachri ( Cucumis collosus), fruit Kachri ( Cucumis collosus), Kair ( Capparisdecidua) and Sanghri ( Prosopis cineriria). The β-carotene contents of the samples were estimated in fresh and various processed states (blanched, dried and roasted). The results showed the heat lability of β-carotene. Among the samples, kair was found to be the richest source of β-carotene. Variety, too, had an impact on the content of this fat soluble vitamin. [ABSTRACT FROM AUTHOR]

Maize possesses natural variations for crtRB1 (β-carotene hydroxylase) and lcyE (lycopene-ε-cyclase) which significantly enhances provitamin A (proA) concentration. Enhancement of kernel proA carotenoids in QPM genetic background is of greater value. In this study, allelic variability for proA carotenoids was analyzed to identify promising β-carotene donor to introgress into QPM inbreds employing molecular breeding. Maize inbred MGU23379 (6.31 μg/g of β-carotene) was used as β-carotene donor to cross with recurrent parents (RPs), CB6-36 (CBML6) and CB7-28 (CBML7). In conversion program, F1, BC1F1, BC2F1, BC2F2, BC2F3, and BC2F4 materials were generated. In each generation, foreground selection was carried out with crtRB1-3′TE and umc1066. The crtRB1-3′TE segregated as per the expectation. Selection for modifiers, recombinants, and background genome of RPs was carried out in BC populations. The individuals with high recovery of recurrent parent genome were retained. Tryptophan/lysine content in introgressed progenies was on par, but β-carotene content was significantly high (6.25and 6.80 μg/g) compared to original inbreds (0.71 and 1.29 μg/g). Phenotypic data recorded for different traits in the BC2F3 populations did not show any significant difference between the converted BC2F3 families and their RPs. Also, grain yield of converted inbreds (PVCBML6 and PVCBML7) was on par with their original lines. In PVCBML6, stem anthocyanin pigmentation was reduced and silk color was changed to dark pink; whereas in PVCBML7, tassel was more erect and sparer; and silk color was changed to light pink compared to original inbreds. Converted inbreds provides an ideal platform for stacking number of nutritionally important traits. [ABSTRACT FROM AUTHOR]