Caracterización química y cuantificación del rendimiento de extracción de pigmento en siete accesiones mexicanas de Bixa orellana

Achiote (Bixa orellana) is a plant used for obtaining a natural dye rich on carotenoids (mainly bixin and norbixin); it is also the plant species with the highest content of tocotrienols in nature. In the present work, the pigment extraction yield of seven Mexican accessions of Bixa orellana was quantified. Also color parameters and content of tocotrienols, tocopherols, norbixin, bixin, total phenolic compounds and antioxidant capacity were evaluated in the corresponding annatto extracts. The highest percentage of pigment extraction yield was obtained with KOH (4.84%). Accessions 43 (L*= 4.01 ± 0.79, C*= 7.33 ± 1.07, h= 25.76 ± 6.35) and 50 (L*= 3.17 ± 0.64, C*= 6.81 ± 0.53, h= 26.41 ± 4.41) had the lowest color values, meaning these accessions had a darker and redder color. Four accessions showed the highest content of bixin: accession 48 (3.1%), 45 (2.6%) 43 (2.4%) and 47 (2.2%). Accession 50 had showed the highest content of total phenolic compounds and of tocotrienols (T3), mainly the isoform δ-T3 (5.03 ± 0.64 mg g−1 Seed Dry Weight), as well as the highest antioxidant capacity.El achiote (Bixa orellana) es una planta utilizada para obtener un colorante natural rico en carotenoides (principalmente bixina y norbixina); además, es la especie vegetal con el mayor contenido de tocotrienoles. En este trabajo, se determinó el rendimiento de extracción de pigmento de siete accesiones mexicanas de Bixa orellana. También se evaluaron los parámetros de color y el contenido de tocotrienoles, tocoferoles, norbixina, bixina, compuestos fenólicos totales y la capacidad antioxidante en extractos de annato. El mayor porcentaje de rendimiento de extracción de pigmento fue obtenido con KOH (4.847905%). Las accesiones 43 (L*= 4.01 ± 0.79, C*= 7.33 ± 1.07, h= 25.76 ± 6.35) y 50 (L*= 3.17 ± 0.64, C*= 6.81 ± 0.53, h= 26.41 ± 4.41) presentaron los valores más bajos de los párametros de color, lo que significa que estas accesiones tuvieron un color más oscuro y más rojo. Cuatro accesiones mostraron el mayor contenido de bixina: accesión 48 (3.1%), 45 (2.6%) 43 (2.4%) and 47 (2.2%). La accession 50 mostró el mayor contenido de compuestos fenólicos totales y de tocotrienoles (T3), principalmente la isoforma δ-T3 (5.03 ± 0.64 mg g-1 Peso Seco), así como también la mayor capacidad antioxidante.Fil: Raddatz Mota, D.. Universidad Autónoma Metropolitana; MéxicoFil: Pérez Flores, L. J.. Universidad Autónoma Metropolitana; MéxicoFil: Carrari, Fernando Oscar. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Insani, Ester Marina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Asis, Ramón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; ArgentinaFil: Mendoza Espinoza, J. A.. Universidad Nacional Autónoma de México; MéxicoFil: Díaz de León Sánchez, F.. Universidad Autónoma Metropolitana; MéxicoFil: Rivera Cabrera, F.. Universidad Autónoma Metropolitana; Méxic

De novo transcriptome sequencing in Bixa orellana to identify genes involved in methylerythritol phosphate, carotenoid and bixin biosynthesis.

BackgroundBixin or annatto is a commercially important natural orange-red pigment derived from lycopene that is produced and stored in seeds of Bixa orellana L. An enzymatic pathway for bixin biosynthesis was inferred from homology of putative proteins encoded by differentially expressed seed cDNAs. Some activities were later validated in a heterologous system. Nevertheless, much of the pathway remains to be clarified. For example, it is essential to identify the methylerythritol phosphate (MEP) and carotenoid pathways genes.ResultsIn order to investigate the MEP, carotenoid, and bixin pathways genes, total RNA from young leaves and two different developmental stages of seeds from B. orellana were used for the construction of indexed mRNA libraries, sequenced on the Illumina HiSeq 2500 platform and assembled de novo using Velvet, CLC Genomics Workbench and CAP3 software. A total of 52,549 contigs were obtained with average length of 1,924 bp. Two phylogenetic analyses of inferred proteins, in one case encoded by thirteen general, single-copy cDNAs, in the other from carotenoid and MEP cDNAs, indicated that B. orellana is closely related to sister Malvales species cacao and cotton. Using homology, we identified 7 and 14 core gene products from the MEP and carotenoid pathways, respectively. Surprisingly, previously defined bixin pathway cDNAs were not present in our transcriptome. Here we propose a new set of gene products involved in bixin pathway.ConclusionThe identification and qRT-PCR quantification of cDNAs involved in annatto production suggest a hypothetical model for bixin biosynthesis that involve coordinated activation of some MEP, carotenoid and bixin pathway genes. These findings provide a better understanding of the mechanisms regulating these pathways and will facilitate the genetic improvement of B. orellana

Identification, heterologous expression and characterization of a dye-decolorizing peroxidase of Pleurotus sapidus

The coding sequence of a peroxidase from the secretome of Pleurotus sapidus was cloned from a cDNA library. Bioinformatic analyses revealed an open reading frame of 1551 bp corresponding to a primary translation product of 516 amino acids. The DyP-type peroxidase was heterologously produced in Trichoderma reesei with an activity of 55,000 U L-1. The enzyme was purified from the culture supernatant, biochemically characterized and the kinetic parameters were determined. The enzyme has an N-terminal signal peptide composed of 62 amino acids. Analysis by Blue Native PAGE and activity staining with ABTS, as well as gel filtration chromatography showed the native dimeric state of the enzyme (115 kDa). Analysis of the substrate range revealed that the recombinant enzyme catalyzes, in addition to the conversion of some classic peroxidase substrates such as 2,2-azino-bis(3-ethylthiazoline-6-sulfonate) and substituted phenols like 2,6–dimethoxyphenol, also the decolorization of the anthraquinonic dye Reactive Blue 5. The enzyme also catalyzes bleaching of natural colorants such as ß-carotene and annatto. Surprisingly, ß-carotene was transformed in the presence and absence of H2O2 by rPsaDyP, however enzyme activity was increased by the addition of H2O2. This indicates that the rPsaDyP has an oxidase function in addition to a peroxidase activity. As a consequence of the high affinity to the characteristic substrate Reactive Blue 5 the rPsaDyP belongs functionally to the dyp-type peroxidase family

Efficacy of different oils used for the extraction of annatto colour from the seeds of Bixa orellana L.

Annatto, a naturally obtained colour is a potential and valuable Non Wood Forest Product (NWFP). It is obtained from the seeds of plant species Bixa orellana L. The present experiment was carried out to find the efficacy of different oils used for annatto colour extraction. Experiment was started with the pretreatment of seeds of B. orellana with soaking in water at 0%, 50%, 100% and 150% (v/w) for 24 hrs at room temperature and colour extraction with castor oil at 100°C showed that maximum bixin (%) was found from the pretreated seeds with 50% (v/w) water soaking i.e., 0.46% bixin. Similarly for oil soluble annatto extraction method, the maximum extraction of bixin (0.99 %) was found with castor oil at 120°C for 1 hr and least bixin (0.03 %) was recorded with mustard oil at 60oC temperature for 1 hr. Sesame oil and ground nut oil were also showed better extractability. Out of different treatments, castor oil at 120°C for 1 hr gave better result with respect to bixin extraction as compared to others

The New Diverse Accession to the List of the Monospecific Genera Bixa orellana L., a Natural Food Dye Plant from Bangalore, Karnataka

Bixa orellana L., is an evergreen shrub or a small tree, measuring 2-8 m and is a native of Brazil. The tree is Cultivated for its Non-Toxic, Edible Food dye Bixin also called popularly as Annatto having Orange- red color. Bixa orellana L., is one of the 13 natural food dye yielding plants. The pigment bixin extracted from its seeds are economically important next to caramel and saffron .Bixin also replaces the most expensive saffron as a colouring agent in various food preparations as it is not all that expensive as saffron. Bixa orellana L., being Monospecific genera, there are no concrete evidences in the literature to classify the various species in this genus. Our earlier studies reported the existence of Intraspecific diversity by identifying the 5 diverse accessions out of 20 accessions of Bixa orellana L., identified and studied from various areas of Bangalore, Karnataka showing Morphological, Physiological and Genetic Variations and also variations in the dye-Bixin content among the above mentioned 5 diverse accessions of the same. The present study reported the 6th diverse accession of Bixa orellana L., identified from Bangalore itself in confirmation of our earlier reports of the existence of intraspecific diversity in the s

Extraction of bixin from annatto seeds using supercritical carbon dioxide

The solubility of 93% pure bixin in supercritical carbon dioxide (SC-CO2) and of the bixin present in annatto seeds (Bixa orellana L.) was measured. For the seeds, the measurements were made in a temperature range from 30 to 50ºC and pressure between 10 and 35 MPa and for the pure bixin, at 40ºC from 10 to 35 MPa. The main pigments of annatto seeds are bixin and norbixin, but the extracts only showed the presence of cis and trans-bixin, indicating that norbixin is not soluble in SC-CO2. The annatto seeds used in the experiments contained about 2.7% bixin and 3.1% oil. In the seeds, the crossover point of solubility was at about 28 MPa and values for solubility were about ten times higher than those of the pure bixin, giving evidence that the oil acted as a co-solvent with the CO2.FAPES