33 research outputs found
Caracterização estrutural de alguns polissacarídeos presentes no basidioma de Pleurotus pulmonarius e aplicações
No full textOrientador : Prof. Dr. Marcello IacominiCo-orientadora : Drª Eliane R. CarboneroDissertação (mestrado) - Universidade Federal do Paraná, Setor de Ciências Biológicas, Programa de Pós-Graduação em Bioquímica. Defesa: Curitiba, 2008Inclui bibliografiaO presente trabalho teve por objetivo elucidar a estrutura química de alguns
polissacarídeos presentes no basidioma de Pleurotus pulmonarius, bem como
verificar os efeitos antinociceptivos e antiinflamatórios de uma manogalactana
isolada e, analisar alguns parâmetros nutricionais. O corpo de frutificação do
basidiomiceto seco, moído e deslipidificado foi submetido a extrações aquosas
(a frio e a quente) e alcalina (KOH 1%). Os extratos aquosos foram
primeiramente submetidos à precipitação com etanol, enquanto o extrato
alcalino foi submetido a adição de ácido acético até pH 5,0. Os extratos
polissacarídicos obtidos foram submetidos aos processos de purificação por
congelamento e degelo, precipitação com solução de Fehling, ultrafiltração e
diálise por membranas de diferentes porosidades, de maneira sequencial. A
partir destes processos foram purificadas e caracterizadas diferentes
estruturas. A fração MG obtida pela extração aquosa a frio apresentou-se pura
e composta por manose, galactose e O-metil-galactose. Análises
espectroscópicas e de metilação sugerem que essa molécula apresenta cadeia
principal constituída de 3-O-Me-a-D-Galp e a-D-Galp ligadas (1 - 6), sendo que
ambas as unidades podem estar substituídas em O-2 por terminais não
redutores de b-D-Manp. Esse polissacarídeo apresentou um efeito analgésico,
porém não demonstrou atividade antiinflamatória, quando avaliado em teste de
contração muscular induzida por ácido acético em camundongos. A fração
PHW obtida pela extração aquosa a quente, após tratamento com NaOH
apresentou glucose como principal componente monossacarídico. Os
espectros de RMN-¹³C e HMQC dessa fração apresentaram sinais
característicos de uma b-glucana-(1 - 3), substituída em O-6 por b-Glcp,
semelhante à lentinana. O extrato alcalino obtido foi tratado com solução de
Fehling, originando uma fração sobrenadante (SF-GLC) e uma fração
precipitada (PF-GLC) neste reagente. Ambas apresentaram espectros de
RMN-¹³C semelhantes e característicos de b-glucanas. Entretanto, a análise de
metilação mostrou que SF-GLC é composta por alto teor de ligações (1- 3) e
(1 - 6), enquanto PF-GLC é composta principalmente por ligações (1 - 6). A
análise dos parâmetros nutricionais desse cogumelo mostrou que este é rico
em proteínas e aminoácidos essenciais apresentando baixos teores de lipídios.
Além disso, é uma boa fonte de minerais apresentando cálcio, magnésio,
cobre, zinco e manganês em sua composição.The present object work is an elucidation of chemical structures of some
polysaccharides extracted from the fruiting bodies of Pleurotus pulmonarius,
besides their biological applications and nutritional value. The dried and milled
basidiomycete was submitted to aqueous extraction (cold and hot), and alkaline
extraction (1% KOH). The aqueous extracts were firstly submitted to ethanol
precipitation, and the alkaline extract was neutralized with acetic acid. The
resulting polysaccharide extracts were submitted successively to a freezethawing
process, precipitation with Fehling solution, ultrafiltration and dialysis
with membranes of different Mr cut-offs. After these processes different
polysaccharides were characterized. Fraction MG, obtained from the cold water
extraction, was homogeneous and was composed of mannose, galactose and
O-methyl-galactose. Spectroscopic and methylation analysis suggest that this
molecule has a main chain composed of 3-O-Me-a-D-Galp e a-D-Galp (1 - 6)-
linked units, with both substituted at O-2 by b-D-Manp non-reducing end units.
This polysaccharide presented an analgesic effect, independently of an
inflammatory response, when analyzed by a test of abdominal constriction
induced by acetic acid in mice. Fraction PHW obtained via hot water extraction,
after treatment with NaOH, had glucose as its principal monosaccharide
component. The ¹³C-NMR and HMQC spectra of this fraction contained signals
characteristic of a (1 - 3)-linked-b-glucan, substituted at O-6 by b-Glcp, similar
to lentinan. The alkaline extract was treated with Fehling solution giving rise to a
soluble (SF-GLC) and precipitated fraction (PF-GLC). Both fractions had similar
¹³C-NMR spectra and they were characteristic of b-glucans. Although
methylation data showed that SF-GLC was composed of a high proportion of
(1 - 3) and (1 - 6) linkages, and PF-GLC was composed mostly of (1 - 6)
linkages. Analysis of the nutritional value of this mushroom showed that it is rich
in protein and essential amino acids, besides containing lower levels of lipids. In
addition, it is a good source of minerals such as calcium, magnesium, copper,
zinc and manganese
Polysaccharides from <it>Agaricus bisporus </it>and <it>Agaricus brasiliensis </it>show similarities in their structures and their immunomodulatory effects on human monocytic THP-1 cells
No full textAbstract Background Mushroom polysaccharides have traditionally been used for the prevention and treatment of a multitude of disorders like infectious illnesses, cancers and various autoimmune diseases. Crude mushroom extracts have been tested without detailed chemical analyses of its polysaccharide content. For the present study we decided to chemically determine the carbohydrate composition of semi-purified extracts from 2 closely related and well known basidiomycete species, i.e. Agaricus bisporus and A. brasiliensis and to study their effects on the innate immune system, in particular on the in vitro induction of pro-inflammatory cytokines, using THP-1 cells. Methods Mushroom polysaccharide extracts were prepared by hot water extraction and precipitation with ethanol. Their composition was analyzed by GC-MS and NMR spectroscopy. PMA activated THP-1 cells were treated with the extracts under different conditions and the production of pro-inflammatory cytokines was evaluated by qPCR. Results Semi-purified polysaccharide extracts of A. bisporus and A. brasiliensis (= blazei) were found to contain (1→6),(1→4)-linked α-glucan, (1→6)-linked β-glucan, and mannogalactan. Their proportions were determined by integration of 1H-NMR signs, and were considerably different for the two species. A. brasiliensis showed a higher content of β-glucan, while A. bisporus presented mannogalactan as its main polysaccharide. The extracts induced a comparable increase of transcription of the pro-inflammatory cytokine genes IL-1β and TNF-α as well as of COX-2 in PMA differentiated THP-1 cells. Pro-inflammatory effects of bacterial LPS in this assay could be reduced significantly by the simultaneous addition of A. brasiliensis extract. Conclusions The polysaccharide preparations from the closely related species A. bisporus and A. brasiliensis show major differences in composition: A. bisporus shows high mannogalactan content whereas A. brasiliensis has mostly β-glucan. Semi-purified polysaccharide extracts from both Agaricus species stimulated the production of pro-inflammatory cytokines and enzymes, while the polysaccharide extract of A. brasiliensis reduced synthesis of these cytokines induced by LPS, suggesting programmable immunomodulation.</p
Strengths and weaknesses of the aniline-blue method used to test mushroom (1→3)-β-D-glucans obtained by microwave-assisted extractions
No full textThe parameters to extract polysaccharide-enriched fractions (PEF) from mushrooms using MAE (microwave-assisted extraction) were adjusted following a full factorial 3 2 experimental design. The highest yield and total carbohydrate values, using Lentinula edodes as model mushroom, were obtained at 180 °C and 30 min. Several mushroom species were submitted to MAE and their PEF yields ranged between 12.1–44.2%. (1→3)-β-Glucans determination using a conventional fluorimetric method changed depending on the standard utilized. NMR analyses of PEF indicated that the presence of other polysaccharides in the extracts or their specific folding, might impair the proper determination of (1→3) linkages by the fluorophore. Mushrooms from Cantharellales order contained (1→3)-β-glucans but they were not detected with the fluorimetric method. Therefore, although the method (after adjustments) was sensitive enough to detect their presence in many mushroom extracts, it cannot be used for all species and it is also not recommended for quantitative determinationsThis research was supported by national R+D program from the Spanish Ministry of Science and Innovation (Project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain ( S2013/ABI-2728 ). The authors would also like to thank the Brazilian funding agencies CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) , CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior) and the Fundação Araucária
Testing the effect of combining innovative extraction technologies on the biological activities of obtained β-glucan-enriched fractions from Lentinula edodes
No full textInnovative technologies as ultrasound-assisted extraction (UAE) (550 W, 60% amplitude, 50 °C) or subcritical water extraction (SWE) (200 °C, 11.7 MPa) were more effective than hot water extractions to obtain β-glucan-enriched fractions from shiitake mushrooms. UAE required longer extraction time (60 min) than SWE (15 min). Combination of UAE + SWE or pre-treatment of the raw material with supercritical CO2 (SFE) (40 °C, 35 MPa, 3 h) before both extractions yielded extracts containing larger β-glucan concentrations. Fluorimetric/colorimetric determinations indicated that obtained fractions contained (1 → 3)- and (1 → 3),(1 → 6)-β-glucans. NMR confirmed their presence as well as (1 → 3)-α-glucans and heteropolymers including mannose and galactose. SWE (15 min), SFE + SWE or UAE + SWE extracts showed larger glucose levels and lower mannose and galactose residues than the other extractions suggesting certain extraction specificity towards β-glucans. They also included more chitin-derivatives than UAE. The extracts obtained after combination of technologies partially retained their immunomodulatory properties but they showed high hypocholesterolemic activities according to in vitro studies.This research was supported by national R+ D program from the Spanish Ministry of Science and Innovation (project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain (S2013/ABI-2728).Peer reviewe
Production of a β-d-glucan-rich extract from Shiitake mushrooms (Lentinula edodes) by an extraction/microfiltration/reverse osmosis (nanofiltration) process
No full textA pilot-scale process combining extraction of Shiitake mushroom (Lentinula edodes) powder in water (98 °C, 1 h), cross-flow microfiltration and reverse osmosis (nanofiltration) was performed to obtain β-glucan-rich extracts. Suspensions (45–80 L) obtained after 3 extractions were clarified by microfiltration reducing their turbidity to <1 NTU. Membrane flux was completely recovered after filtration. One of the clarified extracts was concentrated (to 6–7 L) by reverse osmosis (Nanomax95) and the other two by nanofiltration (Nanomax50 and ALNF99-2517). Different physicochemical parameters (permeate flux, total soluble substances, total suspended particles and electrical conductivity) were monitored during filtration and the bioactive compounds present in the obtained fractions (β-glucans, total carbohydrates, chitins, eritadenine, lenthionine, ergosterol) were analyzed. The more adequate membrane for Shiitake extract concentration was Nanomax50 because it showed higher filtration flux and higher values of bioactive compounds in the obtained extract than the extracts obtained with the other two membranes.This research was supported by national R + D program from the Spanish Ministry of Science and Innovation (project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain (S2013/ABI-2728).Peer reviewe
Caracterización bioquímica y funcional de extractos ricos en fibra dietética obtenidos de Lentinula edodes
No full textResumen del trabajo presentado a las III Jornadas Científicas CIAL Fórum, celebradas del 22 al 23 de noviembre de 2018 en el Instituto de Investigación en Ciencias de la Alimentación (CIAL).Las setas comestibles han formado parte de la dieta desde hace siglos y sus propiedades nutricionales y beneficiosas para la salud han estimulado un crecimiento exponencial de su consumo en las últimas décadas. En particular, la seta shiitake (Lentinula edodes) es una de las más valoradas, no solo por sus características culinarias sino también por su contenido en compuestos bioactivos.
La obtención de la fracción rica en fibra dietética se realizó sometiendo a cuerpos fructíferos de shiitake pulverizados a elevadas temperaturas (98ºC) en medio acuoso para eliminar compuestos hidrosolubles y concentrar aquellos insolubles, tales como polímeros de difícil degradación durante la digestión humana, que suelen ser fermentados por el microbioma del colon, es decir, actúan como fibra dietética. El extracto obtenido era rico en quitinas y ciertos tipos de β-glucanos y también contenía ergosterol, un lípido que reduce la absorción de colesterol y puede dar lugar a vitamina D2 mediante la irradiación con luz ultravioleta. La quitina presente puede ser sometida a protocolos de desacetilación para dar lugar a quitosano, polímero bien estudiado por sus propiedades hipolipidémicas, antimicrobianas y antitumorales. En este trabajo, además de su caracterización bioquímica, se llevó a cabo la evaluación de su capacidad antioxidante e hipocolesterolémica y sus propiedades reológicas. Además, se evaluaron diferentes condiciones para elaborar un protocolo de irradiación con luz ultravioleta para la conversión de ergosterol en vitamina D2 y de desacetilación química para la conversión de quitina en quitosano.Los autores agradecen la financiación otorgada por el Ministerio de Ciencia, Innovación y Universidades a través del proyecto AGL2014-56211-R y a la Comunidad de Madrid (S2013/ABI-2728).Peer reviewe
High Molecular Weight Glucan of the Culinary Medicinal Mushroom Agaricus bisporus is an α-Glucan that Forms Complexes with Low Molecular Weight Galactan
No full textAn a-glucan was isolated from the culinary medicinal mushroom A. bisporus by hot water extraction, ethanol precipitation and DEAE-cellulose chromatography. The resulting material showed a single HMW peak excluded from a Sephadex G50 column that could completely be degraded by α-amylase treatment. After heating in 1% SDS a small additional peak of low MW eluted from the G50 column. The monosaccharide composition of the main peak was evaluated by HPLC, and was found to consist of a majority of glucose (97.6%), and a minor proportion of galactose (2.4%). Methylation analysis and degradation by a-amylase indicated the presence of an a-glucan with a main chain consisting of (1®4)-linked units, substituted at O-6 by α-D-glucopyranose single-units in the relation 1:8. Mono- (13C-, 1H-NMR) and bidimensional [1H (obs.),13C-HSQC] spectroscopy analysis confirmed the a-configuration of the Glcp residues by low frequency resonances of C-1 at d 100.6, 100.2, and 98.8 ppm and H-1 high field ones at d 5.06, 5.11, and 4.74 ppm. The DEPT-13C-NMR allowed assigning the non-substituted and O-substituted –CH2 signals at d 60.3/60.8 and 66.2 ppm, respectively. Other assignments were attributed to C-2, C-3, C-4, C-5 and C-6 of the non-reducing ends at d 71.8; 72.8; 70.0; 71.3 and 60.3/60.8 ppm, respectively. The minor proportion of galactose that was demonstrated was probably derived from a complex between the a-glucan and a low molecular weight galactan
Isolation and comparison of α- and β-D-glucans from shiitake mushrooms (Lentinula edodes) with different biological activities
No full textA polysaccharide-enriched extract obtained from Lentinula edodes was submitted to several purification steps to separate three different D-glucans with β-(1→6), β-(1→3),(1→6) and α-(1→3) linkages, being characterized through GC–MS, FT-IR, NMR, SEC and colorimetric/fluorimetric determinations. Moreover, in vitro hypocholesterolemic, antitumoral, anti-inflammatory and antioxidant activities were also tested. Isolated glucans exerted HMGCR inhibitory activity, but only β-(1→6) and β-(1→3),(1→6) fractions showed DPPH scavenging capacity. Glucans were also able to lower IL-1β and IL-6 secretion by LPS-activated THP-1/M cells and showed cytotoxic effect on a breast cancer cell line that was not observed on normal breast cells. These in vitro results pointed important directions for further in vivo studies, showing different effects of each chemical structure of the isolated glucans from shiitake mushrooms.This research was supported by national R + D program from the Spanish Ministry of Science and Innovation (Project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain (S2013/ABI-2728). Part of this study was also financed by the Brazilian funding agencies Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) – Finance Code 001 and the Fundação Araucária.Peer reviewe
Evaluation of microwave-assisted and pressurized liquid extractions to obtain β-D-glucans from mushrooms
No full textMicrowave-assisted extraction (MAE) and pressurized liquid extraction (PLE) were compared as advanced technologies to obtain polysaccharides (particularly biologically active β-glucans) from Pleurotus ostreatus and Ganoderma lucidum fruiting bodies. Extraction effectiveness was compared by a full-factorial experimental design (response surface methodology, RSM), using water as extraction solvent. Total carbohydrate content of the obtained extracts and polysaccharide yields were the variable responses investigated, while temperature and extraction time were the experimental factors. Temperature showed stronger influence in the polysaccharide extraction than time. The latter factor slightly affected MAE but not PLE extractions. Optimal conditions within the studied range were determined for each extraction method and species based on the desirability functions. Regarding the polysaccharide composition, the main differences between the species were more quantitative rather than qualitative, since NMR analyses indicated that all extracts contained mainly β- and α-glucans and heteropolysaccharides. Both extraction systems were effective for polysaccharide extraction from mushrooms.The authors would like to thank Juncao Brazil (Taboão da Serra-SP, Brazil), for the donation of Ganoderma lucidum, to CTICH (Centro Tecnológico de Investigación del Champiñón de La Rioja, Autol, Spain) for the Pleurotus ostreatus strain, and to Anton-paar GmbH for access to Monowave EXTRA instrument. F.R.S. thanks to the Brazilian funding agencies CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico), CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior) and the Fundação Araucária. B.G.L. thanks Spanish MINECO (Ministerio de Economía y Competitividad) for her Juan de la Cierva postdoctoral research contract (ref. JCI-2012-12972). The research was also supported by national R + D program from the Spanish Ministry of Science and Innovation (project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain (S2013/ABI-2728).Peer Reviewe
Vitamin D-enriched extracts obtained from shiitake mushrooms (Lentinula edodes) by supercritical fluid extraction and UV-irradiation
No full textThe combination of supercritical fluid extraction followed by UV-irradiation resulted in an interesting strategy to obtain ergosterol- and Vitamin D2-enriched extracts from Lentinula edodes to design novel functional food. Extractions carried out following a specific experimental design pointed out that extraction yields and ergosterol concentrations were more influenced by pressure than by the extraction temperature although, high temperatures (55–75 °C) might induce transformation of ergosta-7,22-dienol and fungisterol into ergosterol. After extraction, the ergosterol-enriched extracts should be dissolved in methanol or ethanol and irradiated (25 °C, 4 cm) to partially transform ergosterol into vitamin D2. Irradiation at 365 nm was less effective than at 254 nm. The fastest transformation was obtained using a UV lamp covering the complete UV spectrum for a maximum of 1 h. However, this lamp also induced vitamin D4 formation although in lower amounts than vitamin D2 or lumisterol2 while with irradiation at 254 nm most of ergosterol was transformed into vitamin D2.This research was supported by national R + D program from the Spanish Ministry of Science and Innovation (project AGL2014-56211-R) and the regional program from the Community of Madrid, Spain (S2013/ABI-2728). F.R.S. thanks to the Brazilian funding agency CNPq (Conselho Nacional de Desenvolvimento Cientifico e Tecnologico).Peer reviewe
