The antioxidant activities of the volatile and the nonvolatile fractions from Satureja montana obtained by supercritical fluid extraction (SFE) and by conventional techniques, hydrodistillation (HD) and soxhlet extraction (SE), were compared. A good agreement between DPPH and rancimat methods was obtained showing that the extracts were able to scavenge free radicals and to inhibit lipid oxidation. The volatile oil (obtained by SFE at 90 bar/40 degrees C) was the most effective extract, presenting the lowest EC(50) (0.06 g/L) and the highest protector factor (PF = 2.03). These results demonstrated the advantages of SFE over conventional techniques by avoiding thermodegradation and hydrolysis reactions. Furthermore, volatile oil is 15 times richer in thymoquinone than the essential oil (HD). This compound is of great importance due to its antioxidant, neuroprotective, and anti-cancer activities. The combination of carvacrol + thymol + thymoquinone in volatile oil may be responsible for the increase in the antioxidant activity when compared to HD, which demonstrates that, in this case, SFE improves value to the final product.

Plant cryopreservation technologies are used within gene banks for the long‐term preservation of vegetatively propagated collections. Surface‐sterilized plant tissues grown in the field, greenhouse/screenhouse, growth chamber, or in vitro are the source of shoot tips subjected to vitrification‐based cryopreservation methods. Here, we describe the methods used to minimize microbial contamination during the tissue culture initiation process. We also discuss the occurrence and possible elimination of endophytes after extended in vitro culture and during recovery after liquid nitrogen exposure. We describe two case studies in which bacterial endophytes were observed in Citrus gene bank accessions during recovery after cryopreservation. These were identified using the MinION Oxford Nanopore system and Kirby–Bauer disc diffusion assays to examine the bacterial responses to antibiotic exposure. The methods used in this case study could be applied to identify endophytes to better target antimicrobial treatments of plant tissue collections.

Drought is the most limiting environmental factor to crop productivity and presents a great variability in the degree of tolerance among and within species, among varieties. The aim of this study was to characterize sugarcane accessions regarding tolerance to water stress during in vitro cultivation based on changes in biometric, physiological and biochemical characteristics, within species and among species, to support future breeding programs. Adventitious shoots of five sugarcane accessions: Saccharum robustum, Saccharum spontaneum and Saccharum officinarum species, cultivated in Murashige and Skoog medium supplemented with 2% sucrose and 4 g/l Phytagel were used in five water potentials, 0, -0.3, -0.6, -0.9, -1.2 MPa, induced by mannitol. Survival, length of shoots and roots, number of shoots and roots, biomass, proline content in leaves and activity of antioxidant enzymes were analyzed. There is difference among species, and also, within the same sugarcane species when submitted to in vitro drought stress, and S. officinarum was shown to be the most tolerant. Proline can be used as a biochemical indicator of response to drought in sugarcane accessions and its accumulation was intensified in S. robustum and S. spontaneum accessions. Catalase activity remained unchanged with increased drought in sugarcane accessions evaluated.

Hancornia speciosa Gomes, popularly known as mangaba tree, is a fruit tree native to Brazil, with natural occurrence in several regions. However, some factors have contributed to the reduction of natural populations of this species, in addition to the recalcitrant characteristic of its seeds, which hinders their storage for conservation purposes. The application of plant tissue culture techniques is a complementary strategy to the conservation of the existing genetic variability and allows Original Research Article

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Azadirachta indica A. Juss, popularly known as neem, is a species native to India, belonging to family Meliaceae, considered the most important plant species with insecticidal action. The aim of this study was to evaluate the influence of growth regulators on induction and growth of neem callus and to observe their viability for embryogenesis through morpho-histological characteristics. In vitro germinated plants were used for excision of nodal explants. These segments were inoculated in Murashige and Skoog culture medium containing 1.0 mg/l 2,4-D (2,4-dichlorophenoxyacetic) combined with BAP (6-benzylaminopurine) at the following concentrations: 0.0, 0.5, 1.0 and 2.0 mg/l (T1, T2, T3 and T4 respectively), for callus induction. At 0 (mass of nodal segments without callus), 20, 40 and 60 days of culture, the percentage of callus formation was observed and the callus weight was measured for each treatment and at the end of the 60 days, consistency, color, and cell histology were evaluated. There was callus formation in all treatments tested. The highest induction of Azadirachta indica A. Juss callus is observed in the presence of 1.0 mg/l 2,4-D + 2.0 mg/l BAP, with callus showing light brown color, friable consistency and rounded cells with intense cell division, typical of cells with potential embryogenic capacity.

This study evaluated the effect of vitrification solutions and exposure time on the cryopreservation of Brazilian green dwarf coconut plumules (BGD) using the droplet vitrification technique. Explants were excised from BGD mature fruits from the Active Germplasm Bank of Embrapa Tabuleiros Costeiros, Sergipe, Brazil. Firstly, embryos were disinfected, and after excision, plumules were pre-cultivated for 72 hours in Y3 + 0.6 M sucrose + 2.2 g L -1 Gelrite ® culture medium. Plumules were exposed to PVS2 and PVS3 solutions for 15 and 30 minutes and rapidly immersed in liquid nitrogen (-196 ºC). After cryopreservation, they were thawed in culture medium solution (Y3 + 1.2 M sucrose) and cultured in regeneration medium. The experimental design was completely randomized in a 2x2 factorial scheme (vitrification solutions per exposure times), with five replicates per treatment. Data were compared by the Tukey's test at 5% probability. Significant differences were observed in the callogenesis percentage for the solutions x exposure time interaction for non-cryopreserved cultures (-NL) and for exposure time after cryopreservation (+NL). PVS2 and PVS3 combined with 15 minutes of exposure promoted the highest callus formation (70 and 100%, respectively) in control cultures. The exposure time of 30 min, regardless of vitrification solution, resulted in 30% embryogenic callus formation after cryopreservation. These results contributed to the long-term conservation of coconut palm.RESUMO: O objetivo desse estudo foi avaliar o efeito das soluções de vitrificação e do tempo de exposição na criopreservação de plúmulas de coqueiro anão verde do Brasil de Jiqui (BGD), pela técnica de vitrificação em gotas. Os explantes foram excisados de frutos maduros oriundos do Banco de Germoplasma Ativo de Embrapa Tabuleiros Costeiros, Sergipe, Brasil. Os embriões foram desinfestados e as plúmulas, após a excisão, pré-cultivadas durante 72 horas em meio de cultura Y3 suplementado com sacarose 0,6 e 2,2 g L -1 Gelrite ® . As plúmulas foram expostas em soluções de PVS2 e PVS3 durante 15 e 30 minutos, e rapidamente imersas em nitrogênio líquido (-196 ºC). Após a criopreservação, foram descongeladas na solução de meio de cultura Y3 com 1,2 M de sacarose, e cultivadas em meio de regeneração. O delineamento experimental foi inteiramente casualizado em esquema fatorial 2x2 (soluções de vitrificação x tempos de exposição), com cinco repetições por tratamento. Os dados foram comparados pelo teste de Tukey à probabilidade de 5%. Observaram-se diferenças significativas na porcentagem de calogênese para a interação entre soluções e tempo de exposição para as culturas não criopreservadas (-NL), e para o tempo de exposição após a criopreservação (+NL). O PVS2 e o PVS3 combinados com 15 minutos promoveram a maior formação de calo (70 e 100%, respectivamente) nas culturas de controle. O tempo de exposição de 30 min, independente da solução de vitrificação, promoveu 30% da formação de calos embriogênicos após a criopreservação. Estes resultados contribuem para ...

The objective of this research was to evaluate the proline synthesis and physiological response of cassava genotypes which were micro propagated and induced to salinity stress in vitro. Micro cuttings of approximately 1.0cm long with a single bud of genotypes TBRS Tapioqueira, BRS Verdinha and Lagoão which were previously established in vitro were inoculated in a MS medium containing different concentrations of NaCl (0; 25; 50; 75; 100mM) and were analyzed after 90th day for: number of roots, number of leaves and shoot dry mass. The proline content of BRS Tapioqueira and Lagoão was assessed at 30th, 60th and 90th day. There was no analysis of proline of the variety Verdinha because of the contamination of the explants. The experimental design was completely randomized in double factorial scheme (3 genotypes x 5 salt treatments), with seven repetitions for growth variables. For comparing proline content, completely randomized design was used in a plot subdivided in time, with genotype and NaCl factors in plot and time in subplot, with two repetitions. For r time and genotypes Tukey test (P<0,05) was used and for NaCl levels regression test (P<0,05). Salinity affected the growth of all varieties; although, BRS Tapioqueira and BRS Verdinha were less affected by induced salt stress. There was an increase in the accumulation of proline from the salt increment, this synthesis of proline being a biochemical indicator of salt stress in cassava plants cultivated in vitro.