Οι ελαιογόνοι Ζυγομύκητες έχουν την ικανότητα να συσσωρεύουν σημαντικές ποσότητες λιπιδίων, πλουσίων σε πολυακόρεστα λιπαρά οξέα (PUFA) μεταβολίζοντας γλυκερόλη. Στο μυκήλιο όλων των στελεχών που μελετήθηκαν συσσωρεύτηκε μεγαλύτερη ποσότητα ουδέτερων λιπιδίων (NL) από ότι γλυκολιπιδίων, σφιγγολιπιδίων και φωσφολιπιδίων (P). Η βιοσύνθεση των P στους Mortierella ramanniana, Mucor sp. και Cunninghamella echinulata πραγματοποιήθηκε ενώ η διαδικασία συσσώρευσης των NL ήταν σε εξέλιξη. Η συγκέντρωση των PUFA μειώθηκε σε όλα τα λιπιδιακά κλάσματα του M. ramanniana κατά τη διάρκεια της αύξησης. Αντίθετα, στην περίπτωση του C. echinulata τόσο η συγκέντρωση του λινελαϊκού όσο και του γ- λινολενικού οξέος (GLA) αυξήθηκε με το χρόνο σε όλα τα κλάσματα, και κυρίως στο κλάσμα των P. Τα λιπίδια του Mucor sp. ήταν πλούσια σε ελαϊκό οξύ και GLA, ενώ τα Ρ του Thamnidium elegans παρόλο που ήταν πλούσια σε λινελαϊκό οξύ, αυτό δε μετατράπηκε σημαντικά σε GLA. Δεδομένου ότι η κύρια λειτουργία των PUFA στους Ζυγομύκητες σχετίζεται με τη συμμετοχή τους στις μυκηλιακές μεμβράνες θα μπορούσαμε να υποθέσουμε ότι η βιοσύνθεση αυτών των λιπαρών οξέων σχετίζεται άμεσα με τη μυκηλιακή αύξηση. Ωστόσο αυτό ισχύει μόνο για ορισμένες περιπτώσεις Ζυμομυκήτων όπως αυτή του M. ramanniana. Αντίθετα, υπό τις ίδιες πειραματικές συνθήκες, η βιοσύνθεση των PUFA στην περίπτωση του C. echinulata συνεχίζεται ακόμη και μετά την ολοκλήρωση της αύξησης, υποδεικνύοντας ότι σε αυτό το είδος η βιοσύνθεση δεν σχετίζεται άμεσα με την πρωτογενή αύξηση. / Oleaginous Zygomycetes have the ability to accumulate large amounts of lipids rich in polyunsaturated fatty acids (PUFA) that are of pharmaceutical and nutritional interest. In all investigated strains fungal mycelia contained higher amounts of neutral lipids (NL) than glycolipids plus sphingolipids (G+S) and phospholipids (P), while biosynthesis of P in Mortierella ramanniana, Mucor sp. and Cunninghamella echinulata occurred though NL accumulation process was in progress. PUFA concentration gradually decreased in all lipid fractions of M. ramanniana during growth. In contrast, in C. echinulata concentration of both linoleic acid and γ-linolenic acid (GLA) strongly increased with time in all lipids, especially in P. Lipids of Mucor sp. were enriched in both oleic acid and GLA, while P of Thamnidium elegans were enriched in linoleic acid, but this fatty acid was not efficiently converted to GLA. Taking for granted that the main function of PUFA in Zygomycetes is associated to their participation in mycelial membranes we could suppose that biosynthesis of these fatty acids is strongly associated to mycelial growth. However, this is accurate only for some Zygomycetes, such as M. ramanniana. On the contrary, under the experimental conditions used in this work, PUFA biosynthesis in C. echinulata persists after growth cessation, suggesting that in this species biosynthesis is not a strictly growth-associated process.

Made available in DSpace on 2014-06-12T15:04:02Z (GMT). No. of bitstreams: 2 arquivo4414_1.pdf: 421863 bytes, checksum: 1dc983d91fb57f58244502a9a24c4406 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2003 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O perfil de crescimento e o consumo de fosfato e glicose, bem como o conteúdo de fósforo, a distribuição, estrutura e localização de polifosfato foram avaliados em Cunninghamella elegans cultivada em meios contendo diferentes concentrações de fosfato. Os resultados permitiram evidenciar a influência dessas concentrações sobre a produção de biomassa e o metabolismo do polifosfato no organismo estudado. A maior concentração de fosfato no meio de cultivo proporcionou maior rendimento da biomassa ao longo do crescimento. Observou-se uma relação entre consumo de fosfato e glicose com o crescimento e a quantidade de polifosfato total nos micélios cultivados em diferentes concentrações de fosfato. Diferentes frações do polifosfato celular de C. elegans foram identificadas e quantificadas através de extrações seqüenciais. A citoquímica ultrastrutural foi utilizada, com sucesso, para identificar a localização e a distribuição de polifosfato em C. elegans. Os resultados revelaram diferenças no padrão de marcação citoquímica nas diferentes fases do crescimento e meios de cultivo. Uma marcação uniforme do polifosfato foi observada sobre a superfície celular, em especial, na parede celular e na membrana citoplasmática. Produtos de reação, resultantes da marcação citoquímica, foram também visualizados em estruturas trabeculares, vacuolares e vesiculares, sob a forma de corpos eletrondensos e grânulos dispersos no citoplasma. Os resultados demonstraram o potencial de C. elegans na acumulação de polifosfato, sugerindo uma possível aplicação em processos biotecnológicos

The present thesis presents new renewable, antimicrobial and biodegradable superabsorbent polymers (SAPs), produced from the cell wall of zygomycetes fungi. The cell wall was characterized and chitosan, being one of the most important ingredients, was extracted, purified, and converted to SAP for use in disposable personal care products designed for absorption of different body fluids. The cell wall of zygomycetes fungi was characterized by subsequent hydrolysis with sulfuric and nitrous acids and analyses of the products. The main ingredients of the cell wall were found to be polyphosphates (4-20%) and copolymers of glucosamine and N-acetyl glucosamine, i.e. chitin and chitosan (45-85%). The proportion of each of these components was significantly affected by the fungal strain and also the cultivation conditions. Moreover, dual functions of dilute sulfuric acid in relation to chitosan, i.e. dissolution at high temperatures and precipitation at lowered temperatures, were discovered and thus used as a basis for development of a new method for extraction and purification of the fungal chitosan. Treatment of the cell wall with dilute sulfuric acid at room temperature resulted in considerable dissolution of the cell wall polyphosphates, while chitosan and chitin remained intact in the cell wall residue. Further treatment of this cell wall residue, with fresh acid at 120°C, resulted in dissolution of chitosan and its separation from the remaining chitin/chitosan of the cell wall skeleton which was not soluble in hot acid. Finally, the purified fungal chitosan (0.34 g/g cell wall) was recovered by precipitation at lowered temperatures and pH 8-10. The purity and the yield of fungal chitosan in the new method were significantly higher than that were obtained in the traditional acetic acid extraction method. As a reference to pure chitosan, SAP from shellfish chitosan, was produced by conversion of this biopolymer into water soluble carboxymethyl chitosan (CMCS), gelation of CMCS with glutaraldehyde in aqueous solutions (1-2%), and drying the resultant gel. Effects of carboxymethylation, gelation and drying conditions on the water binding capacity (WBC) of the final products, were investigated. Finally, choosing the best condition, a biological superabsorbent was produced from zygomycetes chitosan. The CMCS-based SAPs were able to absorb up to 200 g water/g SAP. The WBC of the best SAP in urine and saline solutions was 40 and 32 g/g respectively, which is comparable to the WBC of commercially acceptable SAPs under identical conditions (34-57 and 30-37 g/g respectively). / <p>Disputationen sker fredagen den 1 oktober kl. 10.00 i KA-salen, Kemigården 4, Chalmers, Göteborg</p>

Organisms belonging to the Kingdom Fungi are known to occupy a wide variety of ecological niches and are found globally in virtually all environments. Two members of the smallest of the fungal phylum, the Zygomycota, have also been found to harbor intercellular bacteria initially described as being from or closely related to organisms from the Genus Burkholderia. In this study two microaerophilic members of the species Verticilium from the phyla Ascomycota were characterized. Both appear to carry two bacterial endosymbionts. This is the first evidence of bacterial endosymbionts found within a member of the Ascomycota. Through the use of fluorescent stains, isolation of the intercellular bacteria, DNA analysis and fluorescent in situ hybridization (FISH) it appears that the newly isolated Verticilium sp. fungi contain not one but two bacterial endosymbionts from the family Proteobacteria. One putative symbiont is from the genus Bradyrhizobium, a member of the α-Proteobacteria, and one from the genus Burkholderia, a member of the β-Proteobacteria. This is the first evidence of a fungus containing not one, but two distinct endosymbionts from two separate bacterial families. Additionally the fungi were found to grow from spore across a large pH gradient (pH 1.2 to pH 13.5) and in conditions lacking given nutrient. They were tolerant of concentrations of Fe(II) up to 50mM and grew better with low oxygen levels (1.6%) than without.

Zygomycetes is a class of fungi living worldwide as saprobes, as part of mycorrhizae, and as parasites. Humans have used some zygomycetes for centuries in the production of traditional foods, e.g. Indonesian tempe. In the present thesis, the experimental focus was on two zygomycetes strains, Mucor indicus CCUG 22424 and Rhizopus sp. IT. One of the distinguishing features of M. indicus is its dimorphism. The different cell forms were influenced by the culturing conditions. After inoculation, when the initial spore concentration was high (6-8×106 spores/ml), yeast-like growth dominated under anaerobic conditions. With a smaller inoculum, yielding 1-2×105 spores/ml, and access to oxygen, filamentous forms dominated. Only negligible differences in ethanol yield (390-420 mg/g hexoses), productivity (3-5 g/l/h), and inhibitor tolerance were observed. Differential expressions of probably four genes were observed between the yeast-like and filamentous growth forms. Lignocelluloses are a suitable substrate for cultivating zygomycetes, as they occur in abundance, particularly since zygomycetes, unlike Saccharomyces cerevisiae, can utilise pentoses. Lignocelluloses require pretreatment to achieve efficient hydrolysis of the cellulose. N-methylmorpholine-N-oxide (NMMO) was tested for pretreatment of spruce and birch. Reducing wood chip size and/or prolonged pretreatment, promoted hydrolysis yield. Best yields were achieved from &lt;2 mm chips and 5 h pretreatment. The hydrolysate was used for fermentation with M. indicus, resulting in 195 and 175 mg ethanol/g wood, and 103 and 86 mg fungal biomass/g wood, from spruce and birch respectively. Orange peel is another potential substrate. However, the hydrolysate contained 0.6 % (v/v) D-limonene, ten times higher than the concentration inhibiting S. cerevisiae. M. indicus was more resistant and successfully fermented the hydrolysate, producing 400 mg ethanol/g hexoses and 75 mg fungal biomass/g sugars. Both M. indicus and Rhizopus sp. grew in 1.0 % and 2.0 % D-limonene, although the latter was unable to grow in the hydrolysate. A third substrate was also used, spent sulphite liquor (SSL), which is a by-product from sulphite paper pulp mills. The SSL was diluted to 50 % and used for airlift cultivations of Rhizopus sp. In 1.0 vvm aeration, up to 340 mg biomass/g sugars was produced. Prolonged cultivations generally decreased the protein (from 500 to 300 mg/g) and lipid (from 70 to 20 mg/g) contents. In contrast, the cell wall fraction, measured as alkali-insoluble material (AIM), increased (160-280 mg/g), as did the glucosamine (GlcN) content (220-320 mg GlcN/g AIM). The produced fungal biomass could serve as animal feed, e.g. for fish. / <p>Akademisk avhandling som för avläggande av teknologie doktorsexamen vid Chalmers tekniska högskola försvaras vid offentlig disputation den 9 februari 2012, klockan 10.00 i KS101, Kemigården 4, Göteborg.</p>

Λόγω της αύξησης των τιμών του αργού πετρελαίου, τα τελευταία χρόνια η παραγωγή βιοντίζελ (biodiesel) έχει αποκτήσει μεγάλο ενδιαφέρον. Η χρήση των φυτικών ελαίων ως πρώτη ύλη στη βιομηχανία παραγωγής βιοντίζελ έχει πολλά μειονεκτήματα. Επομένως, το ενδιαφέρον των ερευνητών έχει στραφεί στη χρήση των SCO (ελαίων μονοκύτταρων οργανισμών), ως μία εναλλακτική πρώτη ύλη. Στην παρούσα διπλωματική εργασία, η βιομηχανική γλυκερόλη μετατράπηκε σε SCO από διάφορα στελέχη ελαιογόνων Ζυγομυκήτων, οι οποίοι καλλιεργήθηκαν υπό μη-ασηπτικές συνθήκες, χρησιμοποιώντας εκλεκτικά, περιοριστικά ως προς την πηγή αζώτου θρεπτικά μέσα καλλιέργειας, τα οποία ανέστειλαν τη βακτηριακή αύξηση. Όταν οι μύκητες Thamnidium elegans, Mortierella ramanniana, Mortierella isabellina, Zygorhynchus moelleri, Mucor sp. και Cunninghamella echinulatα καλλιεργήθηκαν σε pH6 υπό μη-ασηπτικές συνθήκες, ήταν ικανοί να παράγουν 1,2- 4,1 g/l βιομάζα και να συνθέτουν 17,5- 49,3% wt/wt λιπίδια επί της ξηρής τους βιομάζας. Τα επίπεδα σύνθεσης της βιομάζας ήταν ~50% χαμηλότερα από εκείνα που επιτεύχθηκαν όταν οι Ζυγομύκητες καλλιεργήθηκαν υπό ασηπτικές συνθήκες. Διάφορες αντι-βακτηριακές ουσίες (αιθέρια έλαια, αντιβιοτικά) χρησιμοποιήθηκαν ώστε να επιτευχθεί υψηλότερη συσσώρευση βιομάζας και λιπιδίων από τους Ζυγομύκητες. Σε pH5, ο μύκητας M. ramanniana παρήγαγε 4,4 g/l βιομάζα που περιείχε 22% wt/wt λιπίδια, ενώ ο μύκητας T. elegans παρήγαγε 3,4 g/l βιομάζα που περιείχε 45% wt/wt λιπίδια επί της ξηρής τους βιομάζας. Όταν προστέθηκε στο μέσο καλλιέργειας αιθέριο έλαιο θυμαριού, 4,8 g/l βιομάζα συντέθηκαν από το μύκητα Τ. elegans που περιείχαν 42,6% wt/wt λιπίδια. Επιπλέον, όταν προστέθηκαν στο μέσο καλλιέργειας αντιβιοτικά και αιθέριο έλαιο, ο μύκητας T. elegans ήταν ικανός να παράγει 7,9 g/l βιομάζα που περιείχε 31,5 % wt/wt λιπίδια. Τα αποτελέσματα έδειξαν ότι αν και ο πληθυσμός των βακτηρίων ανέστειλε την αύξηση των μυκήτων, η συσσώρευση των λιπιδίων δεν επηρεάστηκε από την παρουσία των βακτηρίων στο μέσο καλλιέργειας, συγκρινόμενα με αποτελέσματα πειραμάτων που πραγματοποιήθηκαν υπό ασηπτικές συνθήκες (control). Ως εκ τούτου, αναπτύχθηκε μια διεργασία δύο σταδίων στις κωνικές φιάλες και στο βιοαντιδραστήρα, κατά την οποία η αύξηση του μύκητα πραγματοποιήθηκε υπό ασηπτικές συνθήκες (1ο στάδιο) και ακολούθησε η συσσώρευση των λιπιδίων που πραγματοποιήθηκε υπό μη-ασηπτικές συνθήκες (2ο στάδιο). Στο 2ο στάδιο, προστέθηκε στο θρεπτικό μέσο καλλιέργειας αιθέριο έλαιο θυμαριού ως αντι- βακτηριακή ουσία. Από αυτή τη διεργασία, υψηλές ποσότητες λιπιδίων συσσωρεύτηκαν μέσα στο μυκήλιο του μύκητα, με απόδοση που έφτασε περίπου 13% wt/wt μικροβιακού ελαίου ανά καταναλωθείσα γλυκερόλη. / Biodiesel production has gained much interest during last years as an alternative fuel source, which arises from the escalating prices of petroleum fuels. The use of plant oils as feedstock for the biodiesel manufacture has many drawbacks, thus the interest has turned to single cell oil (SCO) as an alternative. In this thesis, raw glycerol was converted into SCO by oleaginous Zygomycetes, cultivated under non-aseptic conditions, using selective nitrogen limited media that inhibit the bacterial growth. Thamnidium elegans, Mortierella ramanniana, Mortierella isabellina, Zygorhynchus moelleri, Mucor sp. and Cunninghamella echinulata, cultivated at pH6 under non-aseptic conditions, were able to produce 1.2- 4.1 g/l of biomass and synthesized 17.5- 49.3 % wt/wt of lipids in their biomass. These accumulation levels in biomass were ~50% lower compared to those achieved when Zygomycetes grown under aseptic conditions. Various anti-bacterial compounds, including essential oils and antibiotics were used in order to achieve a higher biomass and lipid accumulation. At pH5, M. ramanniana produced 4.4 g/l biomass containing 22% wt/wt lipids in their biomass, while T. elegans produced 3.4 g/l biomass containing 45% wt/wt of lipids. When thyme essential oil was added into the growth medium, T. elegans produced 4.8 g/l biomass containing 42.6% wt/wt lipids. Furthermore, with the addition of antibiotics together with essential oil into the medium, the production of SCO was further improved with T. elegans being able to produce 7.9 g/l of biomass containing 31.5% wt/wt lipids. The obtained data showed that although bacterial populations inhibited the fungal growth, lipid accumulation remained unaffected by the presence of bacteria in the growth medium compared to control experiments (conducted under aseptic conditions). Therefore, a two-stage process was developed in both flasks and bioreactor, in which growth was performed under aseptic conditions (1st stage) followed by lipid accumulation performed under non-aseptic conditions (2nd stage) in the presence of thyme essential oil as an antibacterial agent. Large amounts of lipids were accumulated inside the mycelia, yielding around 13% wt/wt of oil per glycerol consumed.

As doenças cardiovasculares (DCVs) são um grupo de desordens do coração e dos vasos sanguíneos capazes de causar ataques cardíacos e acidentes vasculares cerebrais, eventos geralmente agudos oriundos principalmente de bloqueios que impedem o sangue de fluir para o coração ou o cérebro. Proteases fibrinolíticas são agentes que degradam a fibrina, principal componente de trombo sanguíneo, capazes de evitar e tratar DCVs. Fungos filamentosos têm se mostrado uma boa alternativa para a produção de enzimas fibrinolíticas, dentre esses os fungos da divisão dos zigomicetos. A cepa UCP 1262 do zigomiceto Mucor subtilissimus foi estudada com o objetivo geral de produzir protease com atividade fibrinolítica com base em trabalhos publicados em fermentação em estado sólido (FES) e cultivo submerso com a mesma cepa. Foram realizados cultivos submersos desta cepa em frascos agitados e em 2 diferentes biorreatores, com determinação da cinética de crescimento, consumo de glicose e produção de enzima. O meio utilizado foi o MS-2, com farelo de trigo como fonte de nitrogênio e glicose como fonte preferencial de carbono. A FES com a mesma fonte de nitrogênio e o cultivo submerso em diferentes condições de taxa de inóculo, pH e agitação deste estudo foram comparados, assim como duas metodologias diferentes para dosagem de concentração de biomassa, gravimetria e ergosterol. As maiores concentrações de atividade fibrinolítica e produtividade volumétrica obtidas foram de, respectivamente, 12,0 U/mL e 0,22 U/(mL.h), 92% e 89% menores, respectivamente, do que o valor reportado em FES. O valor de ?Xmax foi calculado com base no perfil de biomassa, glicose e produção de CO2 e não apresentou correlação com a produção da enzima. A maior simplicidade operacional e os dados de rendimento obtidos indicam que a FES é uma alternativa melhor para a produção dessa enzima com esta cepa. O zigomiceto Mucor subtilissimus demonstrou ser muito difícil de ser cultivado em sistemas submersos com o meio MS-2, em especial pela sua morfologia e aderência ao biorreator. / Cardiovascular diseases (CVDs) are a group of heart and blood vessels disorders capable of causing heart attacks and strokes, usually acute events caused mainly by blockages that prevent blood from flowing to the heart and brain. Fibrinolytic proteases are agents that degrade fibrin, the major component of blood thrombus. Filamentous fungi were shown to be a good alternative for the production of fibrinolytic enzymes, among them fungi of the zygomycetes division. The UCP 1262 strain of the zygomycete Mucor subtilissimus was studied with the general objective of producing protease with fibrinolytic activity based on published works on solid state fermentation (SSF) and submerged culture with the same strain. Submerged cultures of this strain were carried out in shaken flasks and in 2 different bioreactors, with determination of growth kinetics, glucose consumption and enzyme production. The medium used was MS-2, with wheat bran as the nitrogen source and glucose as the preferred carbon source. The SSF with the same nitrogen source and the submerged culture under different inoculum ratios, pH and agitation conditions of this study were compared, as well as two different methodologies for biomass, gravimetry and ergosterol dosage. The highest concentration of fibrinolytic activity and volumetric productivity obtained were, respectively, 12.0 U/mL and 0.22 U/(mL.h), 92% and 89% lower, respectively, than the value reported in SSF. The value of ?Xmax was calculated based on the biomass concentration, glucose and CO2 production profile and showed no correlation with the enzyme production. The greater operational simplicity and yield data obtained indicate that SSF is a better alternative for the production of this enzyme with this strain. The zygomycete Mucor subtilissimus showed to be very difficult to cultivate in submerged culture with the MS-2 medium, especially for its morphology and adherence to the bioreactor.

Many industrially important fungi can grow in the form of small spherical pellets.Such pellets reduce the viscosity and enhances mass transfer rates in culture broths.The pelleted morphology also influences the fungus’s metabolism, directing it tospecific metabolites. The pellets are easily harvested from the broth and recycled.These properties makes pelleted morphology very attractive.The pelleted morphology of four Zygomycetes strains was studies. Several differentnutrient media used by other researchers for achieving pelleted growth was tested.The influence of eight factors on pelletization of Rhizopus sp. in a completely definedmedium was determined using a fractional central composite design and logistic regression.Pelleted growth of all four Zygomycetes was achieved, with very good results for twoRhizomucor sp. strains. A simple medium containing calcium carbonate was foundto induce pelletization with very high reproducibility.Immobilization of yeast cells was attempted in pellets of Rhizomucor. It was foundthat a flocculating yeast can be immobilized inside pellets of fungal mycelium, forming”biocapsules”. This is accomplished by first using a medium that induces pelletizationof the filamentous fungus and does not allow for growth of the yeast. Thepellets are then inoculated into a second medium that induces growth and flocculationof the yeast and inhibits further growth of the filamentous fungus.Non-flocculating yeasts could not be immobilized, suggesting that the flocculin proteinsin the cell wall of flocculating strains are important for proper immobilization.The flocculation and immobilization arises due to expression of several differentFLO-genes and the importance of these genes for successful immobilization isdiscussed.The results demonstrate that the morphology of Zygomycetes can be controlled andthat this may be useful in industrial fermentation. The new immobilization techniquereveals the great importance of flocculation and cell surface hydrophobicity. Usingyeast strains that express certain FLO-genes may be beneficial in fermentation oflignocellulosic hydrolysates.Microscopy techniques were developed that allows for high quality microphotographyof pellets and thin cross-sections of pellets and biocapsules. / Program: MSc in Resource Recovery - Industrial Biotechnology

The present work is synthesizing novel protein-based superabsorbent hydrogels from albumin protein (AP) and isolated zygomycetes protein (IZP) via modification with an acylating reagent and a bifunctional crosslinker, also investigation on their swelling behaviors under some conditions. The hydrophilic acylating reagent was introduced into albumin and zygomycetes proteins and hydrophylicity of albumin and isolated zygomycetes protein were increased by ethylenediaminetetraacetic dianhydride (EDTAD). Provided protein hydrogels through this method include modification of lysyl residues in the unfolded proteins via adding of one or more hydrophilic carboxyl groups. The reaction was developed with a dialdehyde crosslinking reagent (glutaraldehyde) in order to stabilize the modified protein configuration. The maximum capacity of EDTAD-AP hydrogel swelling was observed 296 g water per g of dry gel, and for EDTAD-IZP hydrogel was 87 g water per g of dry gel.In this study, effect of selected physical and chemical parameters such as protein structure, extent of modification, protein concentration, various pH, ionic strength, gel particle size, temperature as important factors affecting the water uptake behavior of superabsorbent hydrogel were investigated. In addition, the effect of some organic solvents particularly absolute ethanol for increasing the swelling properties was studied. / Uppsatsnivå: D

A criopreservaÃÃo à a metodologia de escolha em muitos bancos de microrganismos pois conduz a uma situaÃÃo de dormÃncia metabÃlica e em conseqÃÃncia as culturas mantÃm-se estÃveis por longos perÃodos. Ao reduzir a temperatura pode todavia ocorrer a formaÃÃo de cristais de gelo que tendem a promover lesÃo celular A lesÃo celular por sua vez pode ser evitada pelo processo de vitrificaÃÃo que ocorre, combinando uma soluÃÃo concentrada com o congelamento rÃpido A literatura relata o uso de carboidratos como agentes vitrificadores e crioprotetores TambÃm informa que a tÃcnica de imobilizaÃÃo de cÃlulas em alginato de sÃdio permite o fracionamento da cultura e favorece a preservaÃÃo de microrganismos O objetivo deste trabalho consiste em avaliar a viabilidade dos fungos dos gÃneros Malassezia spp e do grupo dos Zigomicetos imobilizados em alginato de sÃdio utilizando glicose e lactose como crioprotetores Doze cepas de espÃcies de Malassezia spp (9 M furfur 2 M globosa 1 M simpodialis) e doze cepas de Zigomicetos (7 Mucor s e 5 Rhizopus sp) pertencentes à micoteca do Centro Especializado em Micologia MÃdica da Universidade Federal do Cearà O gÃnero Malassezia abrange leveduras lipodependentes associadas a vÃrias enfermidades dermatolÃgicas compreendem na atualidade 13 espÃcies Zigomicetos, por sua vez sÃo fungos saprÃfitas e ubÃquos cujas hifas por apresentarem elevado tamanho e reduzido conteÃdo protÃico estÃo mais vulnerÃveis a danos mecÃnicos Dessa forma constituem grupos fÃngicos de preservaÃÃo difÃcil em estoque portanto uma metodologia que mantenham a viabilidade de Malassezia spp e Zigomicetos tambÃm poderia ser empregada com eficiÃncia para a estocagem de outros fungos As estruturas fÃngicas foram removidas e transferidas para duas soluÃÃes de estoque, formadas por 3% de caldo Sabouraud 15% de alginato de sÃdio e diferenciadas nas concentraÃÃes de glicose e lactose de 9% ou 23% Em seguida, foram adicionadas esferas plÃsticas, medindo 07cm de diÃmetro com orifÃcio central e 0,5cm de comprimento ApÃs a gelificaÃÃo iÃnica com cloreto de cÃlcio, cada cepa foi estocada à temperatura de -20 ÂC e -80 ÂC distribuÃdas em cinco tubos criogÃnicos de 15mL contendo cinco esferas cada uma sendo avaliados em cinco perÃodos ApÃs nove meses foi verificado que nÃo houve recuperaÃÃo de nenhuma cepa de Malassezia spp Com relaÃÃo Ãs espÃcies de Zigomicetos observou-se que apÃs nove meses de estoque quatro cepas mostraram viÃveis a -80 ÂC empregando glicose a 9%, seis cepas foram preservadas a -80 ÂC, utilizando glicose a 23% finalmente seis e sete exemplares foram recuperados a -80 ÂC utilizando lactose a 9% e 23% respectivamente. Dessa forma, o uso de carboidratos como crioprotetores em conjunto com a tÃcnica de imobilizaÃÃo de cÃlulas em alginato de sÃdio constitui alternativa à manutenÃÃo de algumas espÃcies fÃngicas / Cryopreservation is the method of choice in many banks of microorganisms because it leads to a situation of metabolic dormancy and consequently cultures are stable for long periods. By reducing the temperature may however be formed of ice crystals which tend to cause cell damage to cell damage in turn can be avoided by vitrification process which takes place by combining a solution with the quick freezing the literature reports the use of carbohydrates vitrificadores cryoprotectants agents and also advises that the technique of immobilization of cells in sodium alginate allows the fractionation of culture and promotes the preservation of microorganisms The objective of this study is to assess the viability of the fungi of the genus Malassezia spp and the Zygomycetes group of assets in sodium alginate and lactose using glucose as cryoprotectants Twelve strains of species of Malassezia spp (9 M furfur 1 M 2 M globosa simpodialis) and twelve strains of zygomycetes (seven Mucor Rhizopus sp is 5) belonging to the mycology collection of the Specialized Center for Medical Mycology Federal University of Cearà The genus Malassezia comprises yeasts lipodependent associated with various dermatologic diseases currently comprise 13 Zygomycetes species, in turn, are ubiquitous and saprophytic fungi whose hyphae because they have high protein content and reduced size are more vulnerable to mechanical damage this way are groups of fungi in stock so hard to preserve a methodology to maintain the viability of Malassezia spp and Zygomycetes could also be used effectively for the storage of other fungi fungal structures were removed and transferred to two stock solutions, formed by 3% broth Sabouraud 15% sodium alginate and different concentrations of glucose and 9% lactose and 23% were then added plastic spheres, measuring 07cm in diameter and 0.5 cm center hole length After ionic gelation with calcium chloride Each strain was stored at -20  C and -80  C cryogenic distributed in five tubes containing 15 mL each of five levels being evaluated in five periods after nine months it was found that there was no recovery of any strain of Malassezia spp Regarding Zygomycetes species showed that after nine months of inventory four strains were viable at -80  C using glucose 9%, six strains were preserved at -80  C, using 23% glucose last six seven specimens were recovered at -80  C using a 9% lactose and 23% respectively. Thus, the use of cryoprotectants such as carbohydrates in conjunction with the technique of immobilization of cells in sodium alginate is an alternative to maintaining some fungal species