Production of inulinase from Kluyveromyces marxianus YS-1 using root extract of Asparagus racemosus

Roots of Asparagus racemosus were used as a source of inulin for the production of inulinase from Kluyveromyces marxianus YS-1. Root extract prepared at 10 kg/cm² for 10 min showed the maximum production of inulinase. Various parameters of fermentation have been examined in order to improve overall enzyme yield. Inulinase yield of 40.5 IU/mL in an optimized medium containing inulin (3.5%), beef extract (0.5%), MnSO₄ (0.5 mM), CoCl₂ (0.05 mM), SDS (0.4 mM) and pH 6.5 at 30 °C under agitation (150 rpm) for 60 h has been obtained at shake flask level. After optimization the enzyme production has been increased 6.1 times at flask level. In a bioreactor (working volume 1 L) fermentation medium containing 4% inulin has shown maximum inulinase production (47.3 IU/mL) under optimized parameters of 200 rpm, 0.75 vvm aeration at 30 °C after 60 h.

Structural revision of shatavarins I and IV, the major components from the roots of Asparagus racemosus

The two major steroidal saponins from the roots of Asparagus racemosus were isolated by RP-HPLC and their structure determined by extensive NMR studies. Their structures did not match those reported previously for shatavarins. I and IV and were found to be 3-O-{[beta-D-glueopyranosy](1 -> 2)][alpha-L-rhamnopyranosyl(1 -> 4)]-beta-D-glucopyranosyl}-26-O-(P-D-glu(opyranosyl)-(25S)5 beta-furostan-3p,22 alpha,26-triol and 3-O-{[beta-D-glueopyranosyl(1 -> 2)][alpha-L-rhamnopyranosyl(1 -> 4)]-beta-D-glucopyrariosyl}-(25S)-5 beta-spirostan-3 beta-ol. (c) 2006 Elsevier Ltd. All rights reserved.

Asparinins, asparosides, curillins, curillosides and shavatarins: structural clarification with the isolation of shatavarin V, a new steroidal saponin from the root of Asparagus racemosus

A new steroidal saponin, shatavarin V, (3-O-{[alpha-L-rhamnopyranosy](1-2)][beta-D-glucopyranosyl(1 -> 4)]-beta-D-glucopyranosyl}-(25S)-5 beta-spirostan-3 beta-ol), was isolated from the roots of Asparagus racemosus by RP-HPLC, and its structure determined by 1D and 2D NMR studies. This data permits clarification of the structures reported for several known saponins: asparinins A and B; asparosides A and B; curillin H; curillosides G and H and shavatarins I and IV. (c) 2006 Elsevier Ltd. All rights reserved.

Production of high fructose syrup from asparagus inulin using immobilized exoinulinase from Kluyveromyces marxianus YS-1

Extracellular exoinulinase from Kluyveromyces marxianus YS-1, which hydrolyzes inulin into fructose, was immobilized on Duolite A568 after partial puriWcation by ethanol precipitation and gel exclusion chromatography on Sephadex G-100. Optimum temperature of immobilized enzyme was 55 °C, which was 5 °C higher than the free enzyme and optimal pH was 5.5. Immobilized biocatalyst retained more than 90% of its original activity after incubation at 60 °C for 3 h, whereas in free form its activity was reduced to 10% under same conditions, showing a signiWcant improvement in the thermal stability of the biocatalyst after immobilization. Apparent Km values for inulin, raYnose and sucrose were found to be 3.75, 28.5 and 30.7 mM, respectively. Activation energy (Ea) of the immobilized biocatalyst was found to be 46.8 kJ/mol. Metal ions like Co2+ and Mn2+ enhanced the activity, whereas Hg2+ and Ag2+ were found to be potent inhibitors even at lower concentrations of 1 mM. Immobilized biocatalyst was eVectively used in batch preparation of high fructose syrup from Asparagus racemosus raw inulin and pure inulin, which
yielded 39.2 and 40.2 g/L of fructose in 4 h; it was 85.5 and 92.6% of total reducing sugars produced, respectively.

Development of a stable continuous flow immobilized enzyme reactor for the hydrolysis of inulin

A 23.5-fold purified exoinulinase with a specific activity of 413 IU/mg and covalently immobilized on Duolite A568 has been used for the development of a continuous flow immobilized enzyme reactor for the hydrolysis of inulin. In a packed bed reactor containing 72 IU of exoinulinase from Kluyveromyces marxianus YS-1, inulin solution (5%, pH 5.5) with a flow rate of 4 mL/h was completely hydrolyzed at 55 °C. The reactor was run continuously for 75 days and its experimental half-life was 72 days under the optimized operational conditions. The volumetric productivity and fructose yield of the reactor were 44.5 g reducing sugars/L/h and 53.3 g/L, respectively. The hydrolyzed product was a mixture of fructose (95.8%) and glucose (4.2%) having an average fructose/glucose ratio of 24. An attempt has also been made to substitute pure inulin with raw Asparagus racemosus inulin to determine the operational stability of the developed reactor. The system remained operational only for 11 days, where 85.9% hydrolysis of raw inulin was achieved.

Partial purification and characterization of exoinulinase from kluyveromyces marxianus YS-1 for preparation of high-fructose syrup

An extracellular exoinulinase( ^{2, 1}- ß- D fructan fructanohydrolase, EC 3.2.1.7), which catalyzes the hydrolysis of inulin into fructose and glucose, was purified 23.5-fold by ethanol precipitation, followed by Sephadex G-100 gel permeation from a cell-free extract of Kluyveromyces marxianus YS-1. The partially purified enzyme exhibited considerable activity between pH 5 to 6, with an optimum pH of 5.5, while it remained stable(100%) for 3 h at the optimum temperature of 50º c. Mn²⁺ and Ca²⁺ produced a 2A-fold and 1.2-fold enhancement in enzyme activity, whereas Hg²⁺ and Ag²⁺  completely inhibited the inulinase. A preparation of the partially purified enzyme effectively hydrolyzed inulin, sucrose, and raffinose, yet no activity was found with starch, lactose, and maltose. The enzyme preparation was then successfully used to hydrolyze pure inulin and raw inulin from Asparagus racemosus for the preparation of a high-fructose syrup. In a batch system, the exoinulinase hydrolyzed 84.8% of the pure inulin and 86.7% of the raw Asparagus racemosus inulin, where fructose represented 43.6mg/ml and 41.3mg/ml, respectively.

Molecole bioattive naturali e sintetiche per la terapia di malattie da aumentato riassorbimento osseo

Le terapie convenzionali per le malattie da aumentato riassorbimento osseo sono limitate dalla tossicità sistemica, bassa biodisponibilità farmacologica e scarsa aderenza alle terapie. In questo studio sono stati considerati approcci terapeutici innovativi basati su composti naturali e sintetici. I) Valutazione dell'attività biologica di composti naturali. Evidenze sperimentali hanno dimostrato l’attività antiproliferativa ed antiapoptotica di piante della Medicina ayurvedica. Queste proprietà sono sfruttabili nel trattamento di malattie da aumentato riassorbimento osseo, come l'osteoporosi. Per chiarire i possibili effetti terapeutici di questi composti, sono stati studiati i decotti di Rubia cordifolia, Hemidesmus indicus, Emblica officinalis, ed Asparagus racemosus. Hemidesmis indicus si è dimostrato il più efficace. II) Valutazione dell'attività biologica di composti sintetici. I bisfosfonati (BP) sono farmaci capaci di legarsi alle superfici minerali ossee e all’idrossiapatite, nei siti di rimodellamento osseo. Poiché i BP inibiscono la funzione degli osteoclasti, sono convenzionalmente impiegati nel trattamento di malattie da aumentato riassorbimento osseo, come l'osteoporosi. Tuttavia, gli elevati costi e gli effetti collaterali legati alla somministrazione determinano una scarsa aderenza al trattamento condizionandone l’efficacia. Scopo di questo studio è stato quello di valutare l'attività biologica di BP chimicamente innovativi, meno tossici e sintetizzati con strategie catalitiche semplificate ed ecocompatibili, in modo da ridurre i costi di produzione. È stato valutato l’effetto citotossico e antiosteoclastico dei composti e confrontato con quello dei BP comunemente impiegati in clinica (neridronato, pamidronato e alendronato). I risultati sono stati considerati raggiunti qualora fossero identificati BP di nuova sintesi non citotossici e capaci di conservare almeno il 90% della capacità dei substrati di base di inibire il riassorbimento osseo. Tutti i composti di nuova sintesi sono risultati meno tossici del BP convenzionale, anche a concentrazioni più elevate ed i più efficaci sono stati un BP coniugato con acido biliare, un BP aromatico contenente azoto ed un BP alifatico contenente zolfo.

The control of climbing asparagus (Asparagus africanus Lam.) in remnant Brigalow scrub in south-east Queensland

A replicated trial was conducted at Tallegalla in south-east Queensland to assess the effectiveness of a range of control methods for climbing asparagus Asparagus africanus Lam. A total of 18 treatments using mechanical, cut stump, basal bark, foliar spray and splatter gun techniques were trialled with a range of herbicides and application rates. Removing the plant and placing it above the ground surface was most effective in killing climbing asparagus. Basal bark spraying of 24 g triclopyr ester (40 mL Garlon® 600) or 10 g fluroxypyr ester (50 mL Starane® 200) L-1 diesel and the cut stump application of neat diesel or 225 g glyphosate (500 mL Glyphosate CT®) L-1 water offered the best chemical control of climbing asparagus.

The chemical control of the environmental weed basket asparagus (Asparagus aethiopicus L. cv. Sprengeri) in Queensland

A replicated trial to determine effective chemical control methods for the invasive species, basket asparagus (Asparagus aethiopicus L. cv. Sprengeri) was conducted at Currumbin Hill, Queensland, from June 1999 to August 2000. Four herbicides (metsulfuron-methyl, dicamba, glyphosate and diesel) were applied at different times of the year (winter, spring, summer and autumn). Neat diesel applied to adult crowns effectively killed basket asparagus. However, germination of basket asparagus and other weeds was not prevented. An overall spray of 0.06 g metsulfuron-methyl (0.1 g Brush-Off®) + 1 mL BS 1000® L-1 water gave slower but more selective long-term control of basket asparagus when compared to diesel, especially when applied in winter and spring. High rates of foliar applied dicamba were most effective in spring and glyphosate splatter gunned on base of stems in autumn. The combination of increased selectivity, ease of application and likelihood of reduced environmental impacts on native plants, other than coast she-oak (Casuarina equisetifolia L. var. incana Benth.), of metsulfuron-methyl makes it more suitable for controlling large infestations of basket asparagus.

Comparative seed and dispersal ecology of three exotic subtropical Asparagus species

The genus Asparagus includes at least six invasive species in Australia. Asparagus aethiopicus and A. africanus are invasive in subtropical Australia, and a third species, A. virgatus is naturalized and demonstrates localized spread in south east Queensland. To better understand how the attributes of these species contribute to their invasiveness, we compared fruit and seed traits, germination, seedling emergence, seed survival, and time-to-maturity. We further investigated dispersal ecology of A. africanus, examining the diet of a local frugivore, the figbird (Sphecotheres viridis) and the effect of gut passage on seedling emergence. Overall, A. aethiopicus was superior in germination and emergence, with the highest mean germination (98.8%) and emergence (94.5%) under optimal conditions and higher emergence (mean of 73.3%) across all treatments. In contrast, A. africanus had the lowest germination under optimal conditions (71.7%) and low mean seedling emergence (49.5%), but had fruits with the highest relative yield (ratio of dry pulp to fruit fresh weight) that were favored by a local frugivore. Figbirds consumed large numbers of A. africanus fruits (~30% of all non-Ficus fruits), and seedling germination was not significantly affected by gut passage compared to unprocessed fruits. Asparagus virgatus germinated poorly under cool, light conditions (1.4%) despite a high optimum mean (95.0%) and had low mean performance across emergence treatments (36.3%). The species also had fruits with a low pulp return for frugivores. For all species, seed survival declined rapidly in the first 12 mo and fell to < 3.2% viability at 36 mo. On the basis of the traits considered, A. virgatus is unlikely to have the invasive potential of its congeners. Uniformly short seed survival times suggest that weed managers do not have to contend with a substantial persistent soil-stored seed bank, but frugivore-mediated dispersal beyond existing infestations will present a considerable management challenge.

Producción de primicia en invernadero de híbridos de espárrago verde (Asparagus officinalis var. Altilis) en su cuarto año productivo

Resumen: La producción de espárragos en Argentina está caracterizada por una elevada estacionalidad concentrada en el periodo octubre-diciembre, por lo que el empleo de invernaderos brinda la posibilidad de ampliar el calendario de oferta, anticipando la entrada en producción. Con el objetivo de evaluar el rendimiento de primicia de diferentes híbridos de espárrago verde, se realizó un ensayo en invernadero con ocho híbridos, en UCA Buenos Aires, iniciado el 15/11/2006, mediante plantines de 100 días a 1m*0,3m. Se evaluaron los siguientes genotipos: Italo, Zeno, Eros, Ercole, H-668, Marte, y Giove, de origen italiano, y UC-157 de origen americano, como testigo por ser el tradicionalmente cultivado en Argentina. Se evaluaron 22 cosechas, con una frecuencia de día por medio, del 17/08/2011-25/10/2011. Se estudiaron diferencias en kg totales y comerciales (PFT y PFC), Nº turiones totales y comerciales/ha (NTT y NTC) y distribución de calibres (DC): Jumbo (J), Extra-Large (XL), Large (L), Medium (M), Small (S) y Asparagina (A). Se efectuó un análisis multifactor ANOVA LSD test (P>0.05). En promedio se obtuvieron: PFT: 17053; PFC: 7904 kg.ha-1; NTT: 670566 y NTC: 520938 turiones.ha-1. Se destacaron: en PFT: Italo: 29458a, Zeno: 23056b, Giove: 23034b y H-668: 18568bc; en PFC: Italo: 14850a, Giove: 9856b, Zeno: 9130bc y H-668: 8228bcd; en NTT: Italo: 842512a, H-668: 754512a, Giove: 715000b y Eros: 707498b; en NTC: Italo: 844998a, H-668: 667502b y Eros: 542498bc y en DC: en J: Italoa, Gioveb, Zenobc y UC-157bcd; en XL: Italoa, Zenoab y Gioveb; en L: Giovea, Italoa, Zenoab, H-668abc y UC-157abc; en M: Italoa, H-668ab y Erosab; en S: H-668a, Erosa e Italoab y en A: H-668a y Marteab. Por lo expuesto resulta alentadora la productividad de Italo, Giove, Zeno y H-668 para producción de espárrago verde de primicia en invernadero.

Evaluación del crecimiento y producción de turiones de espárrago verde (Asparagus officinalis L.) producidos bajo dos sistemas de inicio, arañas y plantines, en Azul, Buenos Aires

En dos plantaciones de espárrago verde (Asparagus officinalis L.) en el centro de la Pcia. de Buenos Aires, del cultivar UC 157 F2, iniciadas con arañas y plantines, durante su primer año de cosecha, se determinó la tasa de elongación diaria y total hasta el momento de la ramificación de los turiones, el número de días hasta alcanzar el largo exportable (22.5 cm) y los días hasta el momento de la ramificación con el objetivo de estudiar su comportamiento en la zona centro de la provincia de Buenos Aires. Las mediciones se efectuaron durante seis semanas, desde el 26 de septiembre hasta el 4 de noviembre de 2005, desde la emergencia del turión hasta el día de su ramificación (rama de 5 mm sobre su bráctea). La elongación máxima hasta el día de ramificación en promedio y para ambos sistemas de inicio, varió 2 desde 35.3 cm al principio de la cosecha hasta 22.8 cm al final de la misma. Las tasas de elongación relativas variaron en promedio para ambos sistemas de inicio, entre 0.11 cm cm-1 d-1 y 0.38 cm cm-1 d-1 dependiendo de la temperatura del aire. Al mismo tiempo las parcelas constituyentes del ensayo fueron sometidas a una cosecha comercial durante las seis semanas de evaluación. Los espárragos cosechados constituyeron el rendimiento total (kg ha-1) para cada sistema de inicio entre los cuales no se encontraron diferencias estadísticamente significativas. Este estudio permitiría explicar las diferencias de rendimiento comercial para un mismo cultivar en diferentes zonas de producción. Se concluye que para una zona dada, la elongación y sobre todo la ramificación, son variables prioritarias para la determinación del potencial productivo de un cultivar, debiendo tener en cuenta las condiciones de temperatura en las cual se pretende producir este cultivo y el mercado de destino de la producción debido a las mercadas preferencias entre sus consumidores.