HIGHLIGHTS Brachiaria brizantha proved to be a promising biomass for ethanol production. Fermentation was not impaired by the inhibitors furfural and hydroxymethylfurfural.

Abstract Obtaining low cost lignocellulolytic enzymes and efficient biomass pretreatment are key to increase the competitiveness of second-generation ethanol in comparison with fossil fuels. The enzymatic cocktail produced by the Chrysoporthe cubensis fungus as well as the mixture prepared with the cocktails of the Chrysoporthe cubensis and Penicillium pinophilum fungi have already proven to be efficient for hydrolyzing biomass pretreated with alkali. In this study, they were evaluated in saccharification of sugarcane bagasse pretreated with dilute acid or hot water at 121°C using an enzyme loading equal to 8 filter paper units per gram of biomass. The most promising results were obtained from the hydrolysis of biomass pretreated with hot water by the C. cubensis-P. pinophilum enzymes blend. In this condition, the glucose and xylose production were 25.2 g.L-1 and 4.6 g.L-1, respectively, that resulted in the conversion of 68% of glucan and 23% of xylan in only 48 hours. This study shows that the hydrothermal pretreatment is a promising alternative to improve the enzymes performance, produced by the fungi C. cubensis and P. pinophilum, in the sugarcane bagasse hydrolysis without the need of chemical compounds, generally used in the acid and alkali pretreatments. Furthermore, the hydrothermal pretreatment for 60 min allowed all cocktails applied to convert the cellulose efficiently with only 24 h of saccharification, which contributes to the energy savings employed in the process.

ABSTRACT The effects of processing of pelleted diets and their storage time on the stability of the enzyme complex SSF (solid-state fermentation) were evaluated. Two diets were formulated with the same nutritional composition, differing only in the Schizochytrium sp. levels (0 g kg-1 and 50 g kg-1). The samples were collected during the following processing steps: mixing, then pelleting, and then oven. To evaluate the storage time, the diet ready after drying was considered as day 1. On this day, two samples were obtained, being one kept in a room with ambient temperature of 25 °C and another stored in freezer at -18 °C. At 15, 30, 45, and 60 days, sub-samples were taken. All samples from the processing step and storage time were sent to the laboratory and the following enzyme activities were measured: α-galactosidase, endoglucanase (carboxy-methyl-cellulase), xylanase, sucrase, amylase, lipase, and trypsin. There were no significant differences between the mixture and post-oven phase, except for the activity of α-galactosidase, lipase, and sucrase. For the storage temperature of 25 °C, no effect was observed for the activities of endoglucanase, sucrase, xylanase, amylase, and trypsin between the first (after oven) and the 60th day of the trial. As for the storage temperature of -18 °C, no difference was observed, except for the activities of endoglucanase and xylanase between the first and last day. Comparing the two types of storage (25 °C and -18 °C), there was difference only for the activity of galactosidase and trypsin at 60 days. The enzymes of the enzyme complex SSF studied remain stable during the processing of pelleted diet at 55 °C, maintaining activity for at least 60 days when stored at temperatures up to 25 °C.

RESUMO Um dos maiores problemas na comercialização de bananas é o destacamento individual dos frutos das pencas. O presente trabalho teve como objetivo estabelecer a relação entre a concentração de carboidratos e a atividade enzimática na região do pedicelo e a suscetibilidade de bananas ao despencamento natural. Foram utilizadas as cultivares triploides ‘Terra’ (plátano, grupo AAB) e ‘Prata’ (banana, grupo AAB) e a tetraplóide ’Prata Graúda’ (banana, grupo AAAB). O experimento foi conduzido em parcelas subdivididas, com três parcelas (cultivares) e cinco subparcelas (estádios de cor de casca), em um delineamento experimental inteiramente casualizado, com três repetições e três frutos constituindo a unidade amostral. A cultivar Terra mostrou resistência ao despencamento, mesmo estando seus frutos maduros, ao contrário da ‘Prata Graúda’, que, já a partir do estádio 5 (fruto amarelo com pontas verdes), exibiu suscetibilidade à queda. Em todos os estádios de amadurecimento, a cultivar ‘Terra’ teve os maiores teores de matéria seca, apresentando diminuição dos teores de pectinas no decorrer do amadurecimento, enquanto os de amido se mantiveram altos mesmo no fruto maduro. As enzimas pectinametilesterase (PME) e poligalacturonase (PG) são as enzimas-chave na solubilização da parede celular que acompanha o amadurecimento. A alta suscetibilidade da ‘Prata Graúda’ ao despencamento está associada à elevada atividade de PG e PME e ao baixo teor de matéria seca; a maior resistência ao despencamento da cultivar Terra está relacionada com o maior acúmulo de matéria seca e amido no pedicelo.

ABSTRACT A major problem in post-harvest handling of bananas is the individual detachment of the fruit from the hands. This study aimed to establishing the relationship between carbohydrate concentration and enzyme activity in the pedicel region of three cultivars of bananas, resistant and susceptible to natural dropping, during post-harvest ripening, and the susceptibility of bananas to finger dropping. Cultivars ‘Terra’ (plantain, AAB group) and ‘Prata’ (banana, AAB group) triploids and the ‘Prata Graúda’ (banana, AAAB group) tetraploid were used. The experiment was distributed in split plots, with three plots (cultivars) and five subplots (peel color stages) in a completely randomized design with three replications and three fruits per sample unit. ‘Terra’ showed resistance to dropping, even though the fruit were ripe, unlike ‘Prata Graúda’, which, starting from the fifth stage (yellow fruit with green tips), exhibited high susceptibility to dropping. At all ripening stages, the ‘Terra’ had the highest dry mass levels. In turn, the ‘Prata Graúda’ always maintained the lowest levels. The ‘Terra’ showed decreasing levels of pectins during ripening, whereas starch remained high even in the ripe fruit. About the enzymes studied, the results confirmed the increased resistance of the ‘Terra’ to dropping, allowing to conclude that polygalacturonase (PG) and pectinametylesterase (PME) are the key enzymes for the solubilization of the cell wall that accompanies ripening, thus playing a critical role in inducing natural dropping. The high susceptibility of the ’Prata Graúda’ to dropping is associated with the high activity of PG and PME and the low levels of dry mass; the greater resistance of the ‘Terra’ to dropping is related to higher accumulation of dry mass and starch in the pedicel.

Níquel é um micronutriente envolvido no metabolismo de nitrogênio e é componente da molécula da urease. Foram avaliados o crescimento de plantas e a atividade da urease em alface (Lactuca sativa L.) cultivada em solo em vasos, em experimento fatorial 2 x 8, com duas fontes de nitrogênio (N) e oito doses de Ni, com cinco repetições. O N foi aplicado ao solo (200 mg dm-3), incorporando-se metade no transplantio das mudas e a outra metade, mais tarde, na superfície do solo, utilizando-se nitrato de amônio (NH4NO3) e ureia [CO(NH2)2]. Os tratamentos com Ni (0, 2, 4, 8, 12, 16, 24 e 32 mg dm-3) foram aplicados, usando-se cloreto de níquel (NiCl2). Produção de matéria seca, atividade da urease e do Ni nas folhas de alface e Ni no solo (Mehlich-3 e DTPA) foram determinados. A produção de matéria seca nas plantas supridas com NH4NO3 foi maior do que nas supridas com CO(NH2)2. Não houve diferença na produção de matéria seca das folhas em resposta à aplicação de Ni ao solo. A atividade da urease aumentou com o incremento das doses de Ni, mas não se alterou em razão da fonte de N utilizada. As extrações com Mehlich-3 e DTPA foram eficientes para avaliar a disponibilidade de Ni para alface no Latossolo Vermelho-Amarelo.

Nickel is a micronutrient involved in nitrogen metabolism and a constituent of the urease molecule. Plant growth and urease activity were evaluated in lettuce (Lactuca sativa L.) grown in soil-filled pots in a 2 x 8 factorial design with two nitrogen (N) sources and eight Ni rates, with five replications. Nitrogen was applied at 200 mg dm-3 (half the dose incorporated into the soil at seedling transplanting and half top-dressed later) using the sources NH4NO3 (AN) and CO(NH2)2 (Ur). The Ni treatments (0, 2, 4, 8, 12, 16, 24 and 32 mg dm-3) were applied as NiCl2. The shoot dry-matter yield, leaf urease activity, Ni levels in the lettuce leaves and Ni levels extracted from soil with Mehlich-3 (M-3) and DTPA were determined. In the plants supplied with AN, the shoot dry-matter yield was higher than in those supplied with Ur. There was no difference in shoot dry matter in response to soil-applied Ni. The leaf urease activity increased with Ni application, regardless of the N source. The extractions with M-3 and DTPA were efficient to evaluate Ni availability for lettuce in the Red-Yellow Latosol.

The effects of enzyme complex SSF (solid state fermentation) on growth performance and the availability of sucrose and monosaccharides in the chyme of Nile were involved. The study included 360 fish (70g±4.43) in a completely randomized design with six dietary treatments (0, 50, 100, 150, 200 and 250 ppm of SSF) arranged in six replicates, with 10 fish per replicate. Every 15 days, one tilapia of each experimental unit was sacrificed for analyses of carbohydrate in the chyme. On day 60 of the experiment, the performance parameters were measured. There was a linear effect according to treatment for final weight and weight gain. For the other performance parameters, there were no differences. There was quadratic effect for sucrose and glucose in function of the treatment, whereas the fructose levels increased linearly. The addition of 150 ppm of the enzyme complex SSF in the feed improves the performance of Nile tilapia and increases the availability of sucrose and monosaccharides in the chyme.

Este trabalho teve como objetivos a quantificação de proteínas e da atividade da enzima α-galactosidase, no eixo embrionário e nos cotilédones, de sementes de Dalbergia nigra (jacarandá-da-bahia) durante a germinação. As sementes foram colocadas para embeber em água por sete dias, sendo retiradas amostras para a avaliação bioquímica e cinética da enzima. A atividade da enzima α-galactosidase aumenta com a embebição das sementes nos dois compartimentos, embora não esteja presente no eixo embrionário de sementes secas. A diferença na atividade da enzima entre os cotilédones e o eixo embrionário foi significativa. O pH 5,5 foi o de máxima atividade para as enzimas de ambos os compartimentos. A temperatura que mais estimulou a atividade da enzima nos cotilédones foi 50 ºC e de 50 a 60 ºC no eixo embrionário. A atividade da α-galactosidase foi inibida por β-mercaptoetanol e cobre, em ambos os compartimentos, enquanto a lactose e o cloreto de sódio estimularam a atividade tanto nos cotilédones como no eixo embrionário. Os valores de K M para enzimas do eixo embrionário e dos cotilédones foram de 0,239 e 0,228 mM, respectivamente.

This work aimed to quantify the protein content and the α-galactosidase activity in the embryonic axis and in the cotyledons of Dalbergia nigra seeds during the imbibition period. The seeds were submitted to water imbibition during seven days. Biochemical and kinetic characterization of the enzyme were done from samples taken during the imbibition period. The activity of the α-galactosidase increased in the two compartments with the soaking of the seeds, although, the enzyme activity was not detected in the embryonic axis of dry seeds. The difference in the activity of the enzyme between cotyledons and embryonic axis was significant. The pH of maximum activity was 5.5 for the enzyme of both compartments. In the cotyledons the higher activity of the enzyme was obtained at 50 ºC. In the embryonic axis the activity was higher at temperatures ranging from 50 to 60 ºC. The activity of the α-galactosidase was inhibited by β-mercaptoethanol and CuSO4 in both compartments, while lactose and sodium chloride stimulated the activity in the cotyledons and in the embryonic axis. The values of K M for the enzymes of the embryonic axis and cotyledons were respectively 0.239 and 0.228 mM.

Cyperus difformis L. é uma planta daninha ocorrente em lavouras de arroz irrigado, que tem apresentado dificuldade de controle devido à resistência a herbicidas inibidores da enzima ALS. Os objetivos deste trabalho foram investigar características cinéticas da enzima ALS de biótipos de C. difformis e determinar as bases bioquímicas da resistência da espécie ao herbicida pyrazosulfuron-ethyl. Para isso, foram conduzidos experimentos em laboratório do BIOAGRO/UFV. O método utilizado baseou-se na metologia utilizada por CAREY et al. (1997) e adaptada por VARGAS et al. (1999), com algumas modificações. Foram avaliadas a concentração de substrato (piruvato) que fornece velocidade inicial igual à metade da velocidade máxima de reação (K M) e velocidade máxima de reação (Vmáx), bem como a atividade da enzima ALS na presença do inibidor (pyrazosulfuron-ethyl). Diante dos resultados, pode-se observar que a resistência de C. difformis a pyrazosulfuron-ethyl é decorrente da insensibilidade da enzima ALS ao herbicida, não acarretando, porém, prejuízo aos parâmetros cinéticos K M e Vmáx da enzima ALS.

Cyperus difformis L. is a weed that occurs in flooded rice, which has presented difficulty in controlling due to the resistance to ALS inhibiting herbicides. The objectives of this research were to investigate kinetic characteristics of ALS enzyme from C. difformis biotypes and to determine the biochemical bases of resistance from the species to pyrazosulfuron-ethyl herbicide. For that, experiments were conducted at the BIOAGRO/UFV laboratory. The method used was based on the methodology used by CAREY et al. (1997) and adapted by VARGAS et al. (1999), with some modifications. It was evaluated substratum concentration (pyruvate) that provides initial velocity equal to half the speed reaction (K M) and maximum velocity of reaction (Vmáx), as well the activity of the ALS enzyme in the presence of the inhibitor (pyrazosulfuron-ethyl). According to the results, it is possible to observe that C. difformis resistance to pyrazosulfuron-ethyl is due to the insensibility of the ALS enzyme to the herbicide, however without penalty to K M and Vmáx kinetic parameters of the ALS enzyme.

Duas α-galactosidases (P1 e P2) e uma invertase produzidas no sobrenadante da cultura do fungo Aspergillus terreus quando crescido por 168 h a 28ºC com farelo de trigo como fonte de carbono foram parcialmente purificadas por cromatografias de gel filtração e interação hidrofóbica. O pH e temperatura ótimos para as preparações P1, P2 e invertase foram entre 4,5-5,0, 5,5 e 4,0 e 60, 55 e 65ºC, respectivamente. O K M app para Ï�-nitrofenil-α-D-galactopiranosideo foi 1.32 mM e 0.72 mM para P1 e P2, respectivamente. O valor de K M app para invertase usando sacarose como substrato foi de 15,66 mM. As preparações enzimáticas P1 e P2 mantiveram suas atividades após 3 h de pré-incubação a 50 ºC e a invertase manteve cerca de 90% após 6 h a 55 ºC. P1 e P2 foram diferentemente sensíveis à inibição por Ag+, D-galactose e SDS. As preparações enzimáticas hidrolisaram os galactooligossacarídeos presentes em melaço de soja.

Two α -galactosidase (P1 and P2) and one invertase present in the culture of Aspergillus terreus grown on wheat straw for 168 h at 28ºC were partially purified by gel filtration and hydrophobic interaction chromatographies. Optimum pH and temperatures for P1, P2 and invertase preparations were 4.5-5.0, 5.5 and 4.0 and 60, 55 and 65ºC, respectively. The K M app for Ï� -nitrophenyl-α -D-galactopyranoside were 1.32 mM and 0.72 mM for P1 and P2, respectively, while the K M app value for invertase, using sacarose as a substrate was 15.66 mM. Enzyme preparations P1 and P2 maintained their activities after pre-incubation for 3 h at 50ºC and invertase maintained about 90% after 6 h at 55 ºC. P1 and P2 presented different inhibition sensitivities by Ag+, D-galactose, and SDS. All enzyme preparations hydrolyzed galacto-ologosaccharides present in soymolasses.

Sementes de Caesalpinia peltophoroides (Leguminosae Caesalpinioideae) - sibipiruna - foram colocadas para embebição por 144 h, sendo retiradas amostras para análise de proteína, quantificações da atividade de alfagalactosidase e de açúcares presentes na micrópila. A germinação iniciou-se com 96 h de embebição, sem que fossem detectadas modificações na parede celular da micrópila. Nesta, observou-se maior proporção de arabinose, que mostrou tendência de aumento com o decorrer da embebição. A atividade específica da alfagalactosidase foi detectada em sementes secas, tanto no eixo embrionário quanto nos cotilédones, aumentando no primeiro a partir de 24 h de embebição. O aumento da atividade nos cotilédones foi mais lento, sendo mais acentuado a partir de 120 h de embebição. O teor de proteína decresceu continuamente no eixo embrionário a partir de 24 horas de embebição, enquanto se manteve estável nos cotilédones. A atividade da alfagalactosidase foi máxima nas temperaturas de 55 e 50 ºC para o eixo embrionário e para os cotilédones, respectivamente. O pH que mais estimulou a atividade da enzima foi na faixa de 5,5 a 6,0 para o eixo embrionário e na de 4,5 a 5,0 para os cotilédones. As alfagalactosidase do eixo embrionário e dos cotilédones foram inibidas por SDS, CuSO4, galactose e melibiose. Não houve efeito estimulante sobre a atividade da alfagalactosidase do eixo embrionário por nenhum dos efetores, enquanto o mercaptoetanol estimulou a atividade da enzima dos cotilédones. Os K M para o substrato ro-NPGal para a alfagalactosidase do eixo embrionário e dos cotilédones foram de 1,74 e 2,64 mM, respectivamente.

Caesalpinia peltophoroides (Leguminosae Caesalpinioideae) seeds were soaked in water for 144 hours. Samples were taken for protein analysis, quantification of alphagalactosidase activity and micropyle sugar composition. Germination began after 96 hours of imbibition, with no modifications in the micropyle cellular wall. The content of arabinose was higher in the micropyle region, with a tendency to increase during imbibition. Activity of alphagalactosidase was detected in dry seeds in the embryonic axis, as well as in the cotyledons, having increased 24 hours after imbibition. The increase of the activity in the cotyledons was observed only after 120 hours of imbibition. The protein content decreased continuously in the embryonic axis after 24 hours of imbibition, however it kept stable in cotyledons. The activity of the alphagalactosidase was raised by the temperature, being maximum at 55ºC for embryonic axis and at 50ºC for cotyledons. Enzyme activity was favored by pH ranging from 5.5 to 6.0 for the embryonic axis and 4.5 to 5.0 for cotyledons. The effectors SDS, CuSO4, galactose and melibiose inhibited enzyme activity of embryonic axis and cotyledons. Mercaptoethanol showed little stimulating effect on alphagalactosidase in cotyledons. The K M of the embryonic axis and the cotyledons were 1.74 and 2.64, respectively.

Este trabalho objetivou foi determinar a composição bioquímica de sementes de espécies florestais e caracterizar a enzima alfa-galactosidase de sementes germinadas de Platymiscium pubescens. Os maiores teores de lipídios foram determinados em sementes de Chorisia speciosa, Caesalpinia peltophoroides, Tabebuia serratifolia e Tabebuia velanedae, enquanto sementes de Enterolobium contortisiliquum, Schizolobium parahyba e Cassia grandis apresentaram os maiores teores protéicos. A alfa-galactosidase catalisa a hidrólise dos oligossacarídeos de rafinose, em sementes de leguminosas, durante a germinação. A maior atividade da alfa-galactosidase foi detectada em sementes de Platymiscium pubescens após 72 h de embebição. Duas formas de alfa-galactosidases, C1 e C2, foram purificadas de sementes germinadas de P. pubescens, usando-se fracionamento com sulfato de amônio e cromatografias de filtração em gel e de afinidade. Essas enzimas apresentaram atividade máxima em pH 5,5 e a 50-55 ºC. Os valores de Km ap das formas C1 e C2, para o substrato ro-nitrofenil-alfa-D-galactopiranosídeo, foram de 0,54 mM e 0,78 mM, e para a rafinose, de 4,64 mM e 5,09 mM, respectivamente. Essas enzimas exibiram estabilidade térmica moderada, mantendo 70% da atividade original após 3 h de incubação a 45 ºC. A atividade enzimática da C1 e C2 foi totalmente perdida na presença de CuSO4 e dodecil sulfato de sódio (SDS). Tais enzimas também hidrolisaram melibiose, rafinose e estaquiose, indicando potencial para aplicações biotecnológicas.

The objective of this work was to determine seed biochemical composition of forest species and to characterize alpha-galactosidase enzyme of germinated seeds of Platymiscium pubescens. The highest lipid levels were found in seeds of Chorisia speciosa, Caesalpinia peltophoroides, Tabebuia serratifolia and Tabebuia velanedae, whereas seeds of Enterolobium contortisiliquum, Schizolobium parahyba and Cassia grandis showed the highest protein levels. alpha-galactosidase catalyzes the hydrolyzis of raffinose oligossacarides in legume seeds during germination. The highest activity of alpha-galactosidase was found in seeds of Platymiscium pubescens after 72 h of soaking in the water. Two forms of alpha-galactosidases, C1 and C2, were purified from germinated seeds of P. pubescens, using partition with ammonium sulfate, and gel filtration and affinity chromatographies. These enzymes presented maximum activity at pH 5.5, 50-55ºC. Km ap values in the C1 and C2 forms forrho-nitrophenyl-alpha-D-galactopyranoside substrate were 0.54 mM and 0.78 mM, and 4.64 mM and 5.09 mM for raffinose, respectively. These enzymes showed moderate thermal stability, maintaining 70% of the original activity after 3 h incubation at 45ºC. The C1 and C2 enzymatic activity was totally lost in the presence of CuSO4 and sodium dodecyl sulfate (SDS). These enzymes also hydrolyzed melibiose, raffinose and stachyose, indicating a potential for biotechnological applications.

O fungo termofílico Aspergillus fumigatus secreta as enzimas invertase (beta-frutofuranosidase) e alfa-galactosidase (alfa-D-galactosídeo galactohi-drolase) que estão envolvidas na hidrólise completa dos oligossacarídeos de rafinose. A enzima alfa-galactosidase foi produzida em meio de cultura do fungo Aspergillus fumigatus crescido por 36 h a 42 °C em meio mineral mínimo contendo os açúcares galactose, ou melibiose, ou rafinose como fontes de carbono. A enzima foi purificada por filtração em gel, seguida por duas cromatografias de troca iônica. A massa molecular da alfa-galactosidase determinada por SDS-PAGE foi de 54,7 kDa. A atividade máxima da enzima purificada, utilizando o substrato ro-nitrofenil-alfa-D-galactopiranosídeo (roNPGal) foi na faixa de pH entre 4,5 e 5,5 e a 55 °C. A enzima manteve aproximadamente 80% de sua atividade original mesmo após pré-incubação por 90 minutos a 50 °C. O valor de KM para o substrato roNPGal foi 0,3 mM. A enzima foi capaz de hidrolisar melibiose, mas sua atividade foi muito reduzida na presença do substrato rafinose.

Aspergillus fumigatus secreted invertase (beta-fructofuranosidase) and alpha-galactosidase enzymatic activities able to hydrolyzing raffinose oligosaccharides (RO). alpha-Galactosidase was induced by galactose, melibiose and raffinose, but galactose was the most efficient inducer. It was purified by gel filtration and two ion exchange chromatographies and showed Mw of 54.7 kDa. The purified enzyme showed maximal activity against p-nitrophenyl-alpha-D-galactopyranoside (pNPGal) at pH 4.5-5.5 and 55 °C, and retained about 80% of the original activity after incubation for 90 minutes at 50ºC. The KM for pNPGal was 0.3 mM. Melibiose was hydrolyzed by the enzyme but raffinose was very poor substrate.

Sementes de Platymiscium pubescens foram colocadas para embeber em água, sendo retiradas amostras para as caracterizações bioquímica e cinética da enzima alfa-galactosidase do eixo embrionário e dos cotilédones. A atividade específica no eixo embrionário aumenta de zero até o tempo de 96 horas de embebição, estabilizando em seguida. A atividade da enzima dos cotilédones mostrou pequeno incremento durante esse mesmo tempo. A alfa-galactosidase do eixo embrionário apresentou atividade máxima no intervalo de pH de 4,5 a 6,0. Por outro lado, para a enzima proveniente dos cotilédones, a maior atividade foi detectada na faixa de 4,0 a 6,0. A temperatura de 55ºC foi a que mais estimulou as atividades da alfa-galactosidase do eixo embrionário e dos cotilédones. As enzimas do eixo embrionário e dos cotilédones mostraram-se termotolerantes, não se alcançando a meia vida na temperatura de 40ºC, no período de 1.500 minutos. A atividade da alfa-galactosidase do eixo embrionário foi inibida por melibiose, CuSO4 e SDS, enquanto a dos cotilédones foi por todos os efetores, exceto com SDS, CuSO4 e galactose que tiveram efeito neutro sobre a atividade da alfa-galactosidase dos cotilédones. Os valores de K M para as alfa-galactosidases do embrião e para o cotilédone foram 3,37 e 0,26 mM, respectivamente.

Platymiscium pubescens seeds were placed to soak in water and sampled for biochemistry and kinetic characterizations of embryonic axis and the alfa-galactosidase cotyledon enzyme. The specific activity in the embryonic axis increased from zero to 96 hours of imbibition, stabilizing soon after. The activity of the cotyledon enzyme showed a small increase in the same period. The alfa-galactosidase of the embryonic axis presented its maximum activity in the interval of pH 4.5 to 6.0. On the other hand, for the cotyledon enzyme, the highest activity was detected in the interval of 4.0 to 6.0. The temperature of 55ºC was best to stimulate embryonic axis and cotyledon alfa-galactosidase acticvity. Enzymes of the embryonic axis and the cotyledons were heat tolerant, not reaching the half life at 40ºC, in 1.500 minutes. The embryonic axis alfa-galactosidase activity was inhibited by melibiose, CuSO4 and SDS, while that of cotyledons was for all the effectors, except for SDS, CuSO4 and galactose which had neutral effect. The values of K M for the embryo and for the cotyledon alfa-galactosidases were 3.37 and 0.26 mM, respectively.

Este trabalho teve como objetivo estudar as alterações fisiológicas causadas por métodos de quebra da dormência em sementes de Tachigalia multijuga (Benth) provenientes de três matrizes. Compararam-se os efeitos do ácido sulfúrico, da água fervente e do desponte na porcentagem de embebição, na porcentagem e velocidade de germinação, na atividade de alfagalactosidase e betamananase, na síntese de proteína e na alteração da membrana que recobre o embrião. Não houve germinação em sementes tratadas com água quente. Todos os tratamentos resultaram em porcentagem de germinação superior (P<0,05) à da testemunha, com exceção das sementes da matriz Cachoeira, em que o tratamento com ácido sulfúrico por 10 minutos foi semelhante. Entretanto, a velocidade de germinação da testemunha foi diferente (P<0,05) da de todos os tratamentos somente em sementes da matriz Araponga 2. A porcentagem de umidade das sementes tratadas com água quente por 60 segundos foi semelhante àquelas da testemunha e diferente (P<0,05) das tratadas com água quente por 30 minutos e com ácido por 20 minutos. As atividades das enzimas e teores de proteínas durante a germinação foram diferentes (P<0,05) entre os tratamentos com água e ácido.

This work aimed to study the effects of methods to overcome the dormancy of Tachigalia multijuga (Benth) seeds. It was compared the effects of the sulfuric acid, of the boiling water and of the it blunts in the imbibition percentage, in the percentage and speed of germination, in the activity of alpha galactosidase and beta mananase, in the protein synthesis and in the alteration in the membrane that recovers the embryo. There was no germination in seeds treated with hot water. All the treatments resulted in germination percentage superior (P<0.05) to the control, except for the seeds of Cachoeira, where the treatment with sulfuric acid for 10 minutes was the same. On the other hand, the speed of germination of the control was only different (P<0.05) from all the treatments in seeds of Araponga 2. The water percentage of the seeds treated with hot water by 60 seconds went the same to those of the control and different (P<0.05) from the treated with hot water by 30 minutes and by acid for 20 minutes. The activities of the enzymes and proteins content during the germination were different (P<0.05) among the treatments with water and acid. It is discussed the alterations of the membrane sugars content that recovers the embryo seeds.

The involvement of lipoxygenase isozymes in several physiological processes of plants has been described but their role is not well understood and more biochemical studies are needed to elucidate the role of the "Lipoxygenase Pathway" in plant physiology. Thus, the biochemical and kinetic characterization of a lipoxygenases "pool" from soybean leaves was carried out. Two genotypes were used: IAC-100 (a normal variety having lipoxygenases in the seeds) and IAC-100 TN (genetically modified genotype, which is devoid of lipoxygenases in the seeds). The plants were submitted to the application of fatty acids (lipoxygenase substrates) on leaves. The results of the biochemical and kinetic studies of lipoxygenase isozymes from leaves of the two genotypes analysed showed that genetic removal of lipoxygenase from seeds did not affect the response of the plant to the treatment, since both genotypes showed similar results.