Unravelling postharvest quality in microgreens through modulation of preharvest factors

The relatively brief growth cycle required for microgreens to reach harvest maturity renders genotype selection a key component for this expanding new industry. Important compositional differences were presently characterized across microgreens from 13 species and five botanical families. Nitrate hyper-accumulator microgreens were identified that warrants preharvest measures to suppress nitrate content. Across species, K was the most abundant macro-mineral, followed by Ca, P, Mg, S and Na. Genotypic differences in Na, K and S concentrations were wide while variation in P, Ca and Mg was narrower. Antioxidant capacity assayed in vitro was highest in brassicaceous microgreens. The levels of ascorbic acid present in microgreens were higher than corresponding levels in sprouts, plausibly owing to the presence of photosynthetic hexose precursors absent from sprouts. Genotypic variation in pigmentation was also expressed in terms of chlorophyll and carotenoid concentrations. Lamiaceae microgreens exhibited comparatively higher phenolic content, notwithstanding significant varietal differences. Moreover, alternative phenolics-rich species of microgreens, such as coriander from the Apiaceae were for the first time identified. Qualitative and quantitative determination of phenolic profiles demonstrated the predominance of flavonol glycosides, with the O-glycosides of kaempferol showing more species-related distribution. Principal Component Analysis revealed that the clustering of phenolic profiles reflected microgreens' botanical taxonomy with relative consistency. Such information is critical for selecting new species/ varieties of microgreens that satisfy demand for both taste and health. Further to genotype selection, the targeted modulation of microgreens secondary metabolism through select spectral bandwidths was assessed as a tool to produce phytochemically-enriched microgreens of high functional quality and nutritive value. Analytical data on microgreens' response to different light spectra constitutes a valuable resource for designing future crop-specific spectral management systems. Thus, variation in productivity, nutritive and functional quality of novel microgreens (amaranth, cress, mizuna, purslane) was examined in response to select spectral bandwidths (red, blue, blue-red). Growth parameters dependent on primary metabolism were found most favored by blue-red light's efficiency in activating the photosynthetic apparatus. Nitrate accumulation was higher under monochromatic light owing to the dependency of nitrite reductase on the light-driven activity of PSI, most efficiently promoted by blue-red light. Mineral composition was mostly genotype-dependent, however monochromatic red and blue lights increased K and Na and decreased Ca and Mg concentrations. Lutein, β-carotene, and lipophilic antioxidant capacity were generally increased by blue-red light putatively due to the coupling of heightened photosynthetic activity to increased demand for protection against oxidative stress. Finally, the general response to light treatments was a decrease in polyphenolic constituents, particularly flavonol glycosides, and total polyphenols under blue-red light. Notwithstanding that genotype specificity underlies some of the responses to light treatments summarized above, the current work highlights how selection of genetic background combined with effective light management can drive the production of microgreens with superior functional quality. The choice of growth substrate is critical for the production of high-quality microgreens. Therefore, understanding how the physicochemical properties of natural fiber (agave fiber, coconut fiber and peat moss) and synthetic substrates (capillary mat and cellulose sponge) impact the growth and yield attributes, the nutritive and phytochemical composition and the antioxidant potential of select microgreen species (coriander, kohlrabi and pak choi) wan imperative and novel next step in the present line of research. A key finding of this work, which advances our understanding of the current and future literature on microgreens production and potential bioactive value, is that substrates which combine optimal physicochemical properties, such as peat moss, tend to promote faster growth and higher fresh yields that favor high production turnover; however, this is achieved at the expense of reduced phytochemical content, foremost of polyphenols. Therefore, controlled stress applications (e.g., osmotic stress) on microgreens growing on such media warrants investigation as a means of enhancing phytochemical composition without substantial compromise in crop performance and production turnover. Substrates promoting fast growth (e.g., peat moss) also tend to promote nitrate accumulation in microgreens, especially in brassicaceous species that are known nitrate hyperaccumulators. Therefore, nitrate deprivation practices should be considered for microgreens grown on such substrates in order to minimize consumer exposure to nitrates. Although microgreens have become acclaimed as novel gastronomic ingredients that combine visual, kinesthetic and bioactive qualities, the definition of the optimal developmental stage for their harvesting remains fluid. The ontogenetic stages for harvesting microgreens range from the cotyledonary stage to the emergence of the second true leaf. Their superior phytochemical content against their mature counterparts fueled the subsequent work hypothesis that significant changes in their compositional profile likely take place during the brief interval of ontogeny from the appearance of the first (S1) to the second true leaf (S2). Elucidating this hypothesis will contribute towards the standardization of harvest maturity for the microgreens industry. Microgreens of four brassicaceous genotypes (Komatsuna, Mibuna, Mizuna and Pak Choi) thus grown under controlled conditions, harvested at S1 and S2. They were appraised for yield traits and subsequently examined for mineral, volatile organic compounds, polyphenols, ascorbate as well as hydrophilic and lipophilic pigment concentrations. Analysis of compositional profiles revealed genotype as the principal source of variation for all constituents. The absence of significant growth stage effect on many of the phenolic components identified is consistent with previous findings that post-germination differences in phenolic composition between S1 microgreens and baby leaves are minimal. The response of mineral and phytochemical composition and of antioxidant capacity to growth stage was also limited and largely genotype-dependent. It is, therefore, questionable whether delaying harvest from S1 to S2 would significantly improve the bioactive value of microgreens while the cost-benefit analysis for this decision must be genotype-specific. In terms of yield, the lower-yielding genotypes (Mizuna and Pak Choi) registered higher relative increase in fresh yield between S1 and S2, compared to the faster-growing and higher-yielding genotypes. Although the optimal harvest stage for specific genotypes must be determined considering the increase in yield against reduction in crop turnover, harvesting at S2 seems advisable for the lower-yielding genotypes. As reiterated above, microgreens constitute rudimentary leafy greens that impart gastronomic novelty and sensory delight, but are also packed with nutrients and phytochemicals. As such, they comprise an upcoming class of functional foods. However, apart from bioactive secondary metabolites, microgreens also accumulate antinutritive agents such as nitrate, especially under conducive protected cultivation conditions. As stated above, commercially favorable substrates such as peat moss promote fast growth but also tend to promote nitrate accumulation in microgreens, warranting nitrate deprivation practices in order to minimize consumer exposure to nitrates. In this perspective, nutrient deprivation before harvest (DBH) was examined as a plausible strategy, applied by replacing nutrient solution with osmotic water for six and twelve days, on different species (lettuce, mustard and rocket) of microgreens. DBH impact on major constituents of the secondary metabolome, mineral content, colorimetric and yield traits was appraised. Nutrient deprivation was found effective in reducing nitrate content, however effective treatment duration differed between species with decline being more precipitous in nitrate hyperaccumulating species such as rocket. DBH interacted with species for phenolic constituents. It increased the phenolic content of lettuce, decreased that of rocket and did not affect mustard. Further research to link changes in phenolic composition to the sensory and in vivo bioactive profile of microgreens might be warranted. However, it may be safely concluded that brief (≤ 6 days) DBH can be applied across species with moderate or no impact on the phenolic, carotenoid and mineral composition of microgreens. Such brief nutrient deprivation applications also have limited impact on microgreens' yield and colorimetric traits hence on the commercial value of the product. They can therefore be applied for reducing microgreen nitrate levels without significantly impacting key secondary metabolic constituents and their potential bioactive role. Through step-wise examination and appraisal of critical preharvest factors – ranging from genotype and substrate selection, to spectral management, ontogenetic stage at harvest and nutrient deprivation schemes – the current project contributes to the advancement of our understanding on the role and potential utility of these factors in configuring microgreens' yield, sensory, safety, nutritive and bioactive profile

Mapping the primary and secondary metabolomes of carob (Ceratonia siliqua l.) fruit and its postharvest antioxidant potential at critical stages of ripening

Six critical stages corresponding to major morphophysiological events in carob fruit ripening were defined, and changes in the primary and secondary metabolome and in vitro antioxidant capacity were examined in two genotypes collected at low (15 m) and high (510 m) altitudes from genetically identified and georeferenced trees. Soluble carbohydrates were analyzed by HPLC-RI, macro-minerals by ion chromatography coupled to conductivity detection and polyphenols by UHPLC-Q-Orbitrap-HRMS. spectroscopy facilitated assays for condensed tannins and in vitro free-radical scavenging capacity of 1,1-diphenyl-2-picrylhydrazyl (DPPH) and ferric-reducing antioxidant power (FRAP). The fruit respiration rate and moisture content declined sharply during the transition from the breaker to green pedicel stage. Sugar accumulation spiked at the onset of fruit coloration and culminated at 498.7 ± 8.4 mg g−1 dry weight (dw) in the late ripe stage, while the ratio of reducing sugars to sucrose decreased from 3.45 ± 0.32 to 0.41 ± 0.02. The total phenolic compounds and condensed tannins declined with ripening, particularly during the transition from the breaker to green pedicel stage. Eighteen polyphenols were identified and quantitated, with catechins and hydrolyzable tannins being dominant until the onset of fruit coloration. The transition to the green pedicel stage signaled a precipitous decline (90.9%) in catechins, hydrolyzable tannins (60.2%) and flavonol glycosides (52.1%) concomitant to the rise in gallic acid, which was putatively fueled by the enzymatic hydrolysis of gallotannins in immature fruit. Catechins, hydrolyzable tannins and flavone glycosides were more abundant at higher altitudes and gallic acid at lower altitudes. An antioxidant capacity was also favored by higher elevations and declined with ripening, particularly after the breaker stage. Correlations with FRAP and DPPH assays were significant for the total phenolic content, condensed tannins, catechins and hydrolyzable tannins. The highest correlation factors were obtained for epigallocatechin-gallate (r = 0.920 and r = 0.900; p < 0.01). Although the sharp drop in hydrolyzable and nonhydrolyzable tannins and catechins compromised the in vitro antioxidant capacity at physiological maturity, it also reduced the astringency and configured a palatable organoleptic fruit profile. These changes unraveled significant episodes in the ripening-related secondary metabolism of the carob fruit. They further highlighted the value of immature carob as a potent source of gallotannins, with putative in vivo anti-inflammatory action, and of catechins beneficial in preventing and protecting against diseases caused by oxidative stress

Study of the agricultural effectiveness of a last-generation ultrapure mycorrizal gel inoculum

Las micorrizas arbusculares son el objeto de estudio de los científicos desde hace más de un siglo. Hasta hace relativamente poco tiempo eran grandes desconocidos para nosotros, sin embargo, presentan una enorme importancia para la agricultura y el medio ambiente. Manejar correctamente las micorrizas arbusculares podría proporcionar enormes beneficios para la humanidad ya sea aumentando la productividad de cultivos, equilibrando el uso de abonos, mejorando las resistencias de las plantas a plagas, patógenos y diferentes situaciones de estrés; o ayudando a preservar suelos, aumentar la biodiversidad de diferentes biomas y microbiomas, secuestrar el dióxido de carbono de la atmósfera y luchar contra la polución de suelos, aguas, atmósfera y contra el cambio climático. Desde hace varias decenas de años se han diseñado diferentes inoculantes comerciales basados en micorrizas arbusculares, ahora conocidos como “inoculantes micorrícicos convencionales”. Dichos productos solían presentar diversas desventajas para su uso en la agricultura y en la revegetación de territorios degradados, entre ellos su formato sólido insoluble o poco soluble (tierras, limas, arcillas expandidas, polvos micronizados etc.), la presencia únicamente de esporas micorrícicas (propágulos de lenta germinación), su baja densidad de propágulos y la presencia de otros microorganismos (a veces patogénicos). Todas estas características limitaban su uso en la agricultura, la jardinería, la revegetación y otras aplicaciones, frecuentemente generando resultados muy modestos de su efecto en las plantas que ponían en duda el uso rentable de micorrizas. Los agricultores necesitaban inóculos micorrícicos arbusculares aptos para su aplicación vía riego, con buena rapidez de colonización de raíces de las plantas, con alta concentración de propágulos para un manejo fácil, y, muy importante, libres de cualquier tipo de microorganismos indeseados asociados. En 2005 un producto innovador patentado por dos científicos de CSIC (Alberto Bago y Custodia Cano) supuso una auténtica revolución en el mundo de los inoculantes micorrícicos arbusculares, marcando inicio de nueva era en su aplicación a diferentes ámbitos del cultivo de las plantas. El producto consiste en un gel soluble en agua, ultrapuro (sin presencia de otros microorganismos), con muy alta concentración de diferentes tipos de propágulos (esporas, hifas extrarradicales y trozos de raíces micorrizadas) que permiten una colonización rápida de las raíces de plantas. Actualmente este producto se está comercializando en decenas de los países con excelentes resultados. La presente Tesis Doctoral es un compendio de ensayos realizados a lo largo de más de 10 años, en muy diversos cultivos, localizaciones y condiciones ambientales con ese gel inoculante micorrícico ultrapuro (GIMU) de última generación. Los resultados obtenidos ofrecen por vez primera una visión científica de los efectos del gel en condiciones agronómicas reales, lejos de la propaganda comercial tan habitual en este campo tan competitivo, a la vez que fundamental y fascinante. Los veinte ensayos de la Tesis fueron realizados en tres tipos de cultivos: leñosos (olivo, pistachero, almendro, cerezo), hortícolas (tomate, pimiento, pepino) y extensivos (maíz, soja, girasol). Las condiciones de ensayos fueron de campo o de invernadero las más cercanas a la producción típica de la industria agrícolas. En todos los ensayos fue aplicado el GIMU en diferentes dosis y frecuencias de aplicaciones comparándolo con un control (manejo típico del cultivo sin aplicación del GIMU) o con un control positivo (manejo típico del cultivo sin aplicación del GIMU pero con aplicación adicional de un enraizante industrial o un abonado adicional en las primeras dos semanas del cultivo). En cuatro ensayos se realizó la aplicación conjunta del GIMU y del inoculo basado en las rizobacterias promotoras de crecimiento de las plantas Azospirillum brasilense. Se evaluaron los niveles de micorrización de las raíce, el crecimiento de las plantas, la productividad, la calidad de cosechas, los parámetros económicos (gastos, valor de cosecha, beneficios netos, rentabilidad, retorno de la inversión). Se realizó un análisis estadístico ANOVA con la posterior comparación las media de todas pares según la prueba de Fisher LSD a nivel de significación de 95% (α = 0,05). Los resultados de ensayos de la presente Tesis Doctoral demostraron que la aplicación del GIMU en condiciones agronómicas reales induce importantes efectos positivos sobre el vigor, la productividad, la calidad de las cosechas y los beneficios económicos en diversos cultivos agrícolas: leñosos, hortícolas y extensivos. El GIMU funciona como estimulante natural del enraizamiento, tanto aplicado solo como en combinación con enraizantes comerciales. Además, mejora el desarrollo vegetativo, la supervivencia y el éxito de injerto de los plantones de olivo y pistachero. La aplicación del GIMU aumenta en gran medida la resistencia de las plantas al estrés térmico al aumentar su vigor, productividad y calidad de la fruta, al mismo tiempo que supone una práctica agrícola más sostenible y duradera. La aplicación conjunta del GIMU e inoculo bacteriano de Azospirillum brasilense muestra efectos sinérgicos superiores a la aplicación de estos inoculantes biológicos por separado, teniendo gran potencial de uso para mejorar el vigor de las plantas, la productividad, la calidad de cosecha y los beneficios económicos obtenidos. La aplicación del GIMU tiene efectos positivos sobre el vigor, la productividad, la calidad de cosecha y los beneficios económicos independientemente del tipo de producción agrícola: convencional, integrada o ecológica. La detección de la micorriza arbuscular en las raíces de las plantas no debería considerarse como una prueba fehaciente de funcionamiento del GIMU en las plantas tratadas. Los resultados y los cálculos económicos, relacionados con las aplicaciones del GIMU tanto a nivel local de un cultivo concreto, como a nivel de un país (España) y a nivel global, demuestran que el uso de esta tecnología de última generación puede ser económicamente muy beneficioso para los agricultores, haciendo la agricultura más sostenible. En general, el uso de del GIMU de última generación con HMA Rhizophagus irregularis es capaz de amoldarse a las exigencias y condiciones de la agricultura moderna, y debería ser recomendado tanto para los 10 cultivos estudiados (olivo, pistachero, almendro, cerezo, tomate, pimiento, pepino, maíz, soja, girasol) como para todos los demás cultivos micorrizables con el objetivo de incrementar los beneficios productivos, medioambientales y económicos del sector agrícola