Metabolite Profiling of Rosemary Cell Lines with Antiproliferative Potential against Human HT-29 Colon Cancer Cells.

Rosemary (Rosmarinus officinalis) is a culinary and medicinal plant used in food and pharmaceutical industry. The wide range of biological activities is mainly related to phenolic and terpenic compounds; like carnosic acid (CA), carnosol (CS) and rosmarinic acid (RA), mainly reported in rosemary leaf extracts, and recently described in rosemary callus extracts. The aim of this work was to investigate the chemical profile of rosemary cell lines and evaluate their antiproliferative potential against human HT-29 colorectal cancer cell lines. For this purpose, rosemary leaf explants were dedifferentiated on MS medium and added with 2, 4-D (2, 4-dichlorophenoxyacetic acid; 2 mg/L) and BAP (6-benzylaminopurine; 2 mg/L). Cell aggregates were separated according to colour and three rosemary cell lines cultures were established: green (RoG), yellow (RoY) and white (RoW). The chemical profile of rosemary cell lines extracts was characterized by combining HPLC and GC platforms coupled to HR-MS/MS. The antiproliferative activity against HT-29 cell line was analyzed with MTT assay. A total of 71 compounds, including hydroxycinnamic acid and hydroxybenzoic acid derivatives, flavonoids, phenolic di- and triterpenes, as well as relevant unsaturated fatty acids and their esters, phytosterols, and carotenoids were tentatively identified in the extract of the target cell lines. The antiproliferative activity test against HT-29 cell using the MTT assay revealed that the viability of HT-29 colon cancer cells was affected after treatment with the RoW extract (IC50 of 49.63 µg/mL) at 48 h. These results showed that rosemary cell lines can also accumulate other bioactive phytochemicals with demonstrated antiproliferative potential.

Chemical analysis of callus extracts from toxic and non-toxic varieties of Jatropha curcas L.

Jatropha curcas L. belongs to Euphorbiaceae family, and it synthesizes flavonoid and diterpene compounds that have showed antioxidant, anti-inflammatory, anticancer, antiviral, antimicrobial, antifungal and insecticide activity. Seeds of this plant accumulate phorbol esters, which are tigliane type diterpenes, reported as toxic and, depending on its concentration, toxic and non-toxic varieties has been identified. The aim of this work was to characterize the chemical profile of the extracts from seeds, leaves and callus of both varieties (toxic and non-toxic) of Jatropha curcas, to verify the presence of important compounds in dedifferentiated cells and consider the possibility of using these cultures for the massive production of metabolites. Callus induction was obtained using NAA (1.5 mg L-1) and BAP (1.5 mg L-1) after 21 d for both varieties. Thin layer chromatography analysis showed differences in compounds accumulation in callus from non-toxic variety throughout the time of culture, diterpenes showed an increase along the time, in contrast with flavonoids which decreased. Based on the results obtained through microQTOF-QII spectrometer it is suggested a higher accumulation of phorbol esters, derived from 12-deoxy-16-hydroxy-phorbol (m/z 365 [M+H]+), in callus of 38 d than those of 14 d culture, from both varieties. Unlike flavonoids accumulation, the MS chromatograms analysis allowed to suggest lower accumulation of flavonoids as the culture time progresses, in callus from both varieties. The presence of six glycosylated flavonoids is also suggested in leaf and callus extracts derived from both varieties (toxic and non-toxic), including: apigenin 6-C-α-L-arabinopyranosyl-8-C-ß -D-xylopyranoside (m/z 535 [M+H]+), apigenin 4'-O-rhamnoside (m/z 417 [M+H]+), vitexin (m/z 433 [M+H]+), vitexin 4'-O-glucoside-2″-O-rhamnoside (m/z 741 [M+H]+), vicenin-2 (m/z 595 [M+H]+), and vicenin-2,6″-O-glucoside (m/z 757 [M+H]+).

Somatic embryogenesis from leaf explants of Tagetes erecta L.

Tagetes erecta is an asteraceous plant of industrial, ornamental and medicinal importance; its inflorescences have been used as a pigment source for food coloring, mainly for poultry skin and eggs. Nevertheless, there are few reports on plant regeneration or micropropagation, because unsuccesfull results in the plant's reaction to the growth regulators, developing embryogenesis on Tagetes erecta. In this study, somatic embryogenesis was induced and plantlets of Tagetes erecta were regenerated. For induction of globular structures MS medium supplemented with 2,4-D (4.5 µM) and BAP (8.8 µM) was used; globular structures were transferred to MS medium with 45 g l-1 sucrose until the embryos maturation. Transmission electron microscopy showed characteristic subcellular structures of embryogenic callus. Somatic embryos were transferred to MS medium without plant growth regulators and whole plantlets were obtained. In vitro plants were successfully transplanted into a mixture of peat moss and vermiculite (1 : 1 v/v) under greenhouse conditions. In this study, somatic embryogenesis and plant regeneration system from foliar explants were established, an important requirement for performing genetic transformation events on Tagetes erecta.

Callus culture development of two varieties of Tagetes erecta and carotenoid production

Background The properties of natural pigments, such as antioxidants, functional, medical, and nutraceutical, have demonstrated the advantages of these natural compounds over synthetic ones. Some products are accepted only when they are pigmented with natural, food-quality colorants: for example poultry products (manly marigold flower extracts). Carotenoids such as β-carotene, β-criptoxanthin and lutein are very attractive as natural food colorants due to their antioxidant and pro-vitamin activities which provide additional value to the target products. Marigold (Tagetes erecta) is an Asteraceous ornamental plant native to Mexico, and it is also important as a carotenoid source for industrial and medicinal purposes but nowadays its production is destined mainly for ornamental purposes. Results Friable callus of T. erecta yellow flower (YF) and white flower (WF) varieties was induced from leaf explants on Murashige and Skoog (MS) medium supplemented with 9.0 µM 4-dichlorophenoxyacetic acid (2,4-D) and 8.8 µM benzyladenine (BA). Calluses developed from both varieties were different in pigmentation. Extract characterization from callus cultures was carried out by high-performance liquid chromatography (HPLC). This analytical process detected several carotenoids; the main pigments in extracts from YF callus were lutein and zeaxanthin, whereas in the extracts of the WF callus the main pigments were lutein, zeaxanthin, β-cryptoxanthin and β-carotene. Callus cultures of T. erecta accumulated pigments even after several rounds of subculture. Conclusions WF callus appeared to be a suitable candidate as a source of different carotenoids, and tested varieties could represent an alternative for further studies about in vitro pigment production.

Digital image analysis of diverse Mexican rice cultivars.

BACKGROUND: Digital image analysis has an important role in geographical provenance of grains, as it can provide parameters of size, shape and color, which are important quality parameters for the design of engineering processes such as drying and milling of grains. In this study, digital image analysis was used to classify nine rice cultivars based on different morphometric parameters using the three sides of the grain (lateral, ventral and axial), Feret diameter, and 10 different form factors and color parameters (CIE L*, a* and b*). RESULTS: Result of principal component analyisis was an equation with seven variables (area, perimeter, length, width, thickness, sphericity and color), which was useful for distinguishing between nine different cultivars. The morphometric and color parameters for the Mor A-98 and Mor A-92 varieties showed they had 88% similarity. The variability was expressed with a confidence of 95%. CONCLUSION: Multivariate analysis indicated that the lateral side is the most sensitive for the classification of Mexican rice grains because of its color and morphometric characteristics. These results showed the application of image analysis for the future classifications of grains.

Morphostructural characterization of rice grain (Oryza sativa L.) variety Morelos A-98 during filling stages.

The morphostructure of grain rice Morelos A-98 was characterized in five stages of physiological maturation, in order to generate morphometric information during the filling process. Micrographic images from optical and scanning electron microscopy coupled to a digital capture system were used. Images were digitally processed to measure different descriptors such as shape, fractal dimension, and surface texture. Results showed that, two weeks after anthesis, an accelerated grain filling was observed, particularly on those grains positioned in the distal panicle zone, compared to those located in the base of this one. As deposition of assimilates in the grain increased, the area and perimeter of the transversal cut of the grains also increased (P ≤ 0.05); meanwhile, the rounded shape factor tended to increase as well (P ≤ 0.05), while the elliptic shape factor decreased. As the dehydrated endosperm passed from "milky" to "doughy" stages, values of fractal dimension area and endosperm perimeter as well as surface texture values showed that grain borders tended to become smoother and that there was a greater structured endosperm area (P ≤ 0.05).

Estudio del desarrollo, calidad de planta y acumulación de pigmentos en lígulas de tagetes erecta l: expuestas a dos intensidades de luz / Study of development, quality of plant and accumulation of pigments in ligules of tagetes erecta l: exposed to two light intensities

The carotenoids are photosensitive pigments during photosynthesis. The objective of this work was to study the effect on development and accumulation of carotenoids in ligules of Tagetes erecta exposed under two different lighting ambient (with mesh and without mesh of 50 percent). The plant development was evaluated measuring the height of the plant, number of floral buds, the ligules diameter. In adition, the quantification and identification of carotenoids from ligules was done by HPLC. The results showed significant differences (p<0.05) in the height of the plant, number of floral buds and ligules diameter of T. erecta. The group grown without mesh received greater UV radiation and different temperature, that under a mesh. The first conditions lead to a reduction of the ligules diameter and total content of xanthophylls (lutein and zeaxanthin). The plastids ultrastructure in the cells of T. erecta developed with mesh showed the greatest amount of thylakoid membranes and more conspicuous starch granules.

Los carotenoides son pigmentos fotosensibles frente a un exceso de intensidad luminosa durante el proceso de fotosíntesis. El objetivo de este trabajo fue el estudio del efecto en el desarrollo de la planta y la acumulación de carotenoides por la exposición a dos diferentes intensidades lumínicas (con y sin malla de sombra al 50 por ciento). Se evaluó el desarrollo de T. erecta en cuanto a la altura de la planta, número de botones florales y el diámetro de las lígulas. Adicionalmente, en las lígulas se cuantificaron e identificaron los carotenoides por HPLC. Los resultados mostraron diferencias significativas (p<0.05) en cuanto al desarrollo de las plantas expuestas a mayor radiación UV y temperatura, presentaron reducción del diámetro de las lígulas y disminución en el contenido de Xantófilas totales ( luteína y zeaxantina) con respecto a las cultivadas con malla,. La ultraestructura de los plastidios mostró mayor cantidad de membranas tilacoidales y gránulos de almidón más conspicuos en las células de las plantas de T erecta desarrolladas con malla.

Marigold regeneration and molecular analysis of carotenogenic genes.

Marigold (Tagetes erecta) is an Asteraceous plant of industrial, ornamental and medicinal importance. Tagetes erecta species, popularly known as marigold, is grown as ornamental plant and is adapted to several agro climates. Inflorescences have been utilized as pigment source for food coloring, mainly of poultry skin and eggs. Lutein is the main pigment in marigold flowers. Some carotenoids are well known as provitamin A compounds. There are many reports on carotenoids and their effect on the prevention of certain ocular diseases, ischemic heart disease, strokes, photoprotection, immune response, aging and cancer. Marigold flowers are certainly a good source of carotenoids; they show very different pigmentation levels. This chapter describes the establishment of techniques for plant regeneration, gene expression, pigment extraction and pigment deposition in specific structures of marigold.