Growth Response of Non-Conventional Yeasts on Sugar-Rich Media: Part 2: Citric Acid Production and Circular-Oriented Valorization of Glucose-Enriched Olive Mill Wastewaters Using Novel Yarrowia lipolytica Strains.

The global market for citric acid (CA) is one of the biggest and fastest expanding markets in the food industry. The CA production employing microbial bioprocessing with efficient GRAS strains and renewable waste streams is in line with the European Union binding targets for resource efficiency, sustainable consumption-production, and low-carbon technologies. In this work, the potential of three novel wild-type Yarrowia lipolytica strains (namely LMBF Y-46, LMBF Y-47 and ACA-YC 5033) regarding the production of CA and other valuable metabolites was tested on glucose-based media, and the most promising amongst the screened strains (viz. the strain ACA-YC 5033) was cultured on glucose-based media, in which part of the fermentation water had been replaced by olive-mill wastewaters (OMWs) in a novel approach of simultaneous OMW valorization and bioremediation. In the first part of this study, the mentioned strains were cultured under nitrogen-limited conditions with commercial (low-cost) glucose employed as a sole carbon source in shake-flask cultures at an initial concentration (S0) ≈ of 50 g/L. Variable quantities of secreted citric acid (CA) and intra-cellular compounds (viz. polysaccharides and lipids) were produced. All strains did not accumulate significantly high lipid quantities (i.e., maximum lipid in dry cell weight [DCW] values ≈30% w/w were noted) but produced variable CA quantities. The most promising strain, namely ACA-YC 5033, produced CA up to c. 24 g/L, with a yield of CA produced on glucose consumed (YCA/S) ≈ 0.45 g/g. This strain in stirred tank bioreactor experiments, at remarkably higher S0 concentrations (≈110 g/L) and the same initial nitrogen quantity added into the medium, produced notably higher CA quantities, up to 57 g/L (YCA/S ≈ 0.52 g/g). The potential of the same strain (ACA-YC 5033) to bioremediate OMWs and to produce value-added compounds, i.e., yeast cells, CA, and intra-cellular metabolites, was also assessed; under nitrogen-limited conditions in which OMWs had partially replaced tap water and significant glucose concentrations had been added (S0 ≈ 100 g/L, simultaneous molar ratio C/N ≈ 285 g/g, initial phenolic compounds [Phen0] adjusted to ≈1.0 g/L; these media were similar to the OMWs generated from the traditional press extraction systems) the notable CA quantity of 60.2 g/L with simultaneous YCA/S = 0.66 g/g, was obtained in shake flasks, together with satisfactory phenolic compounds removal (up to 19.5% w/w) and waste decolorization (up to 47.0%). Carbon-limited conditions with Phen0 ≈ 1.0 g/L favored the production of yeast DCW (up to 25.3 g/L), with equally simultaneous interesting phenolic compounds and color removal. The fatty acid profile showed that cellular lipids were highly unsaturated with oleic, linoleic and palmitoleic acids, accounting for more than 80% w/w. This study proposed an interesting approach that could efficiently address the biotreatment of toxic effluents and further convert them into circular-oriented bioproducts.

Growth Response of Non-Conventional Yeasts on Sugar-Rich Media: Part 1: High Production of Lipid by Lipomyces starkeyi and Citric Acid by Yarrowia lipolytica.

Sugar-rich waste streams, generated in very high quantities worldwide, constitute an important source of environmental pollution. Their eco-friendly conversions into a plethora of added-value compounds through the use of microbial fermentations is currently a very "hot" scientific topic. The aim of this study, was to assess the potential of single cell oil (SCO), microbial mass and citric acid (CA) production by non-conventional yeast strains growing on expired ("waste") glucose. Six yeast strains (viz. Rhodosporidium toruloides DSM 4444, Rhodotorula glutinis NRRL YB-252, R. toruloides NRRL Y-27012, Yarrowia lipolytica LFMB Y-20, Y. lipolytica ACA-DC 50109 and Lipomyces starkeyi DSM 70296) were initially grown in shake flasks with expired glucose used as substrate under nitrogen limitation, in order to "boost" the cellular metabolism towards the synthesis of SCO and CA, and their growth response was quantitatively evaluated. Initial glucose concentration (Glc0) was adjusted at c. 50 g/L. Besides Y. lipolytica, all other yeast strains produced noticeable SCO quantities [lipid in dry cell weight (DCW) ranging from 25.3% w/w to 55.1% w/w]. Lipids of all yeasts contained significant quantities of oleic acid, being perfect candidates for the synthesis of 2nd generation biodiesel. The highest DCW production (=13.6 g/L) was obtained by L. starkeyi DSM 70296, while both Y. lipolytica strains did not accumulate noticeable lipid quantities, but produced non-negligible CA amounts. The most promising CA-producing strain, namely Y. lipolytica ACA-DC 50109 was further studied in stirred-tank bioreactor systems, while the very promising DCW- and SCO-producing L. starkeyi DSM 70296 was further studied in shake flasks. Both strains were grown on media presenting higher Glc0 concentrations and the same initial nitrogen quantity as previously. Indeed, L. starkeyi grown at Glc0 = 85 g/L, produced DCWmax = 34.0 g/L, that contained lipid =34.1% w/w (thus SCO was =11.6 g/L). The strain ACA-DC 50109 in stirred tank bioreactor with Glc0 ≈ 105 g/L produced CA up to 46 g/L (yield of CA produced on glucose consumed; YCA/Glc ≈ 0.45 g/g). Finally, in fed-batch bioreactor experiment, the significant CA quantity of 82.0 g/L (YCA/Glc = 0.50 g/g) was recorded. Concluding, "waste" glucose proved to be a suitable substrate for a number of non-conventional yeast strains. Y. lipolytica ACA-DC 50109 produced significant quantities of CA while L. starkeyi DSM 70296 was a very interesting DCW- and SCO-producing candidate. These strains can be used as potential cell factories amenable to convert glucose-based residues into the mentioned metabolic compounds, that present high importance for food, chemical and biofuel facilities.

Spent Mushroom Substrate Hydrolysis and Utilization as Potential Alternative Feedstock for Anaerobic Co-Digestion.

Valorization of lignocellulosic biomass, such as Spent Mushroom Substrate (SMS), as an alternative substrate for biogas production could meet the increasing demand for energy. In view of this, the present study aimed at the biotechnological valorization of SMS for biogas production. In the first part of the study, two SMS chemical pretreatment processes were investigated and subsequently combined with thermal treatment of the mentioned waste streams. The acidic chemical hydrolysate derived from the hydrothermal treatment, which yielded in the highest concentration of free sugars (≈36 g/100 g dry SMS, hydrolysis yield ≈75% w/w of holocellulose), was used as a potential feedstock for biomethane production in a laboratory bench-scale improvised digester, and 52 L biogas/kg of volatile solids (VS) containing 65% methane were produced in a 15-day trial of anaerobic digestion. As regards the alkaline hydrolysate, it was like a pulp due to the lignocellulosic matrix disruption, without releasing additional sugars, and the biogas production was delayed for several days. The biogas yield value was 37 L/kg VS, and the methane content was 62%. Based on these results, it can be concluded that SMS can be valorized as an alternative medium employed for anaerobic digestion when pretreated with both chemical and hydrothermal hydrolysis.

Biochemical Evaluation of Agaricus and Pleurotus Strains in Batch Cultures for Production Optimization of Valuable Metabolites.

The production of various biochemical compounds such as proteins, glucans and glucanases, from the mycelium of four strains of Basidiomycetes species, Agaricus bisporus, Agaricus subrufescens, Pleurotus eryngii and Pleurotus ostreatus, during batch culture in shaking flasks, was studied. Fungi were cultured for 26 days in defined media with glucose as carbon source and were primarily evaluated for their ability to consume glucose and produce mycelial mass and intracellular polysaccharides (IPS). Results showed that on the 26th day of cultivation, P. ostreatus produced the maximum biomass (16.75 g/L), whereas P. eryngii showed the maximum IPS concentration (3.82 g/L). All strains presented a similar pattern in total protein production, with A. bisporus having the highest percentage of total proteins (36%, w/w). The calculated correlation coefficients among ribonucleic acid (RNA) vs. biomass (0.97) and RNA vs. protein (0.97) indicated a very strong relation between RNA and biomass/protein synthesis. The studied strains exhibited an increase in total glucan and glucanase (ß-1,6) production during cultivation, with A. bisporus reaching the highest glucan percentage (8%, w/w) and glucanase activity (12.7 units/g biomass). Subsequently, processed analytical data were used in contour-graph analysis for data extrapolation to optimize future continuous culture.

Non-alcoholic fatty liver disease and hepatocellular carcinoma: Clinical challenges of an intriguing link.

Non-alcoholic fatty liver disease (NAFLD) has emerged as the most common liver disorder worldwide mainly attributed to the epidemic spread of obesity and type 2 diabetes mellitus. Although it is considered a benign disease, NAFLD can progress to non-alcoholic steatohepatitis, liver cirrhosis and hepatocellular carcinoma (HCC). Most data regarding the epidemiology of NAFLD-related HCC are derived from cohort and population studies and show that its incidence is increasing as well as it is likely to emerge as the leading indication for liver transplantation, especially in the Western World. Although cirrhosis constitutes the main risk factor for HCC development, in patients with NAFLD, HCC can arise in the absence of cirrhosis, indicating specific carcinogenic molecular pathways. Since NAFLD as an underlying liver disease for HCC is often underdiagnosed due to lack of sufficient surveillance in this population, NAFLD-HCC patients are at advanced HCC stage at the time of diagnosis making the management of those patients clinically challenging and affecting their prognostic outcomes. In this current review, we summarize the latest literature on the epidemiology, other than liver cirrhosis-pathogenesis, risk factors and prognosis of NAFLD-HCC patients. Finally, we emphasize the prevention of the development of NAFLD-associated HCC and we provide some insight into the open questions and issues regarding the appropriate surveillance policies for those patients.

Association of cardiovascular factors in diabetic patients with non-alcoholic fatty liver disease.

PURPOSE: To evaluate the association between severity of hepatic steatosis/fibrosis with clinical, laboratory, and echocardiographic characteristics, including visceral obesity and type 2 diabetes mellitus (T2DM)-related micro- and macrovascular complications in diabetic patients with non-alcoholic fatty liver disease (NAFLD). METHODS: We studied 60 consecutive NAFLD outpatients with T2DM, recording several demographic and clinical characteristics, trunk and visceral fat, cardiac ultrasound, and micro- and macrovascular complications of diabetes mellitus including microalbuminuria, diabetic peripheral neuropathy, peripheral vascular disease, and cardiac autonomic function. Severity of steatosis and fibrosis was evaluated with abdominal ultrasound and liver stiffness measurements, respectively. RESULTS: Twenty-three (41%) of the patients had grade 1 steatosis and mean liver stiffness was 7.5 ± 3 kPa. After applying Bonferroni correction for multiple comparisons, ferritin concentration was the only factor significantly different between patients with mild (grade 1) compared to those with moderate/severe (grade 2/3) steatosis and showed good discriminative ability for the presence of moderate/severe steatosis (AUC: 0.74, sensitivity 88%, specificity 48%, PPV 74%, and NPV 72%). In addition, waist circumference was the only factor associated with the presence of significant fibrosis (≥ F2) with very good discriminative ability (AUC: 0.77, sensitivity 89%, specificity 45%, PPV 75%, and NPV 70%). CONCLUSION: Specific clinical and laboratory characteristics, which may be determined via widely accessible and noninvasive techniques, were associated with severity of diabetics NAFLD, taking into account echocardiographic characteristics, visceral obesity, and T2DM-related systemic complications.

Undergraduate Medical Education During the Pandemic: How do Academic Staff Members in Greece and Cyprus Feel About it?

This study used a questionnaire to examine how academic staff members in Greece and Cyprus feel about the changes in undergraduate medical education during the COVID-19 pandemic. In general, e-learning was not considered an adequate way of teaching and was less enjoyable. Participants aged 31-40 years experienced more (P = .001), while staff in higher academic ranks less difficulty (P < .001) in adjustment. There was a small increase in workload, which was higher among respondents researching into COVID-19 (P = .001). During the pandemic, daily screen use >6 h was increased from 28.8% to 57.5%. The majority (74.2%) stated that scientific and educational training opportunities were not affected by the pandemic. In conclusion, the pandemic has induced important changes in undergraduate medical education. This new condition was considered adequate but not pleasant, with younger faculty members being more burdened.

Impact of olive mill wastewaters on the physiological behavior of a wild-type new Ganoderma resinaceum isolate.

A new wild-type Ganoderma resinaceum isolate was cultivated on glucose-enriched liquid cultures with olive mill wastewaters (OMWs) in initial phenolic compounds concentrations 0.0 (control), 0.5, 0.8, and 1.5 g/l. The effect of the fungus on the reduction of phenolics and color was assessed, whereas biomass production, glucose consumption, intra-cellular (IPS) and extra-cellular (EPS) polysaccharides biosynthesis, antioxidant activity of the biomass, and laccase synthesis were monitored. Results showed that significant phenolic reduction (94.5%) and decolorization (76.5%) occurred, 14.6 g/l of biomass was produced, glucose was almost totally consumed, EPS were produced in sufficient amounts (0.79 g/l), whereas the presence of OMWs enhanced the synthesis of IPS (maximum absolute values 4.0-5.2 g/l corresponding to 35-42% w/w). Kinetic analysis demonstrated that EPS and IPS values fluctuated with time, regardless of the available amount of glucose in the media, showing a maximum in the 17th day of culture. Laccase was highly synthesized in the middle of the fermentation, reaching the maximum value of 14 U/ml. Little growth was however observed at 1.5 g/l phenolics. Strong correlation between total phenolic content and free radical scavenging activity has been noticed in the methanolic extracts of the mycelium. Results strongly suggest the potentiality of G. resinaceum utilization in the OMW waste treatment.

Valorization of low-cost, carbon-rich substrates by edible ascomycetes and basidiomycetes grown on liquid cultures.

Three ascomycetes (Morchella vulgaris AMRL 36, M. elata AMRL 63, Tuber aestivum AMRL 364) and four basidiomycetes strains (Lentinula edodes AMRL 124 and 126, Agaricus bisporus AMRL 208 and 209) were screened for their ability to grow on liquid static flask cultures of glucose, glycerol, molasses and waste flour-rich hydrolysates with C/N ratio of 20 and produce biomass, exopolysaccharides and lipids. The profile of lipid fatty acids was also assessed. Selected strains were furthermore cultivated in C/N = 50. Results showed that substrate consumption, biomass formation and secondary metabolites production were strain, substrate and C/N ratio dependent. The maximum biomass (X), lipid (L) and exopolysaccharides (EPS) values noted were Xmax = 25.2 g/L (C/N = 20; molasses) and Lmax = 6.51 g/L (C/N = 50; rice cereal hydrolysates) by T. aestivum strain AMRL 364 and EPSmax = 2.41 g/L by M. elata strain AMRL 63 (C/N = 50; molasses), respectively. When C/N ratio of 50 was applied, biomass, lipid production and substrate consumption seem to be negatively affected in most of the trials. The adaptation and capability of the mushroom strains to be cultivated on substrates based on agro-industrial waste streams and infant food of expired shelf date offers the opportunity to set a circular oriented bioprocess.

Production of added-value microbial metabolites during growth of yeast strains on media composed of biodiesel-derived crude glycerol and glycerol/xylose blends.

A total of 11 yeast strains of Yarrowia lipolytica, Metschnikowia sp., Rhodotorula sp. and Rhodosporidium toruloides were grown under nitrogen-limited conditions with crude glycerol employed as substrate in shake flasks, presenting interesting dry cell weight (DCW) production. Three of these strains belonging to Metschnikowia sp. accumulated significant quantities of endopolysaccharides (i.e. the strain V.V.-D4 produced 11.0 g/L of endopolysaccharides, with polysaccharides in DCW ≈ 63% w/w). A total of six Y. lipolytica strains produced either citric acid or mannitol. Most of the screened yeasts presented somehow elevated lipid and polysaccharides in DCW values at the early steps of growth despite nitrogen appearance in the fermentation medium. Lipid in DCW values decreased as growth proceeded. R. toruloides DSM 4444 cultivated on media presenting higher glycerol concentrations presented interesting lipid-accumulating capacities (maximum lipid = 12.5 g/L, maximum lipid in DCW = 43.0-46.0% w/w, conversion yield on glycerol = 0.16 g/g). Replacement of crude glycerol by xylose resulted in somehow decreased lipid accumulation. In xylose/glycerol mixtures, xylose was more rapidly assimilated from glycerol. R. toruloides total lipids were mainly composed of triacylglycerols. Total cellular fatty acid composition on xylose presented some differences compared with that on glycerol. Cellular lipids contained mainly oleic and palmitic acid.

Spent mushroom substrate for a second cultivation cycle of Pleurotus mushrooms and dephenolization of agro-industrial wastewaters.

Spent mushroom substrate (SMS) of Pleurotus ostreatus was supplemented with wheat bran and soybean flour and used as substrate for a new cultivation cycle of the oyster mushrooms Pleurotus ostreatus and Pleurotus pulmonarius. The bioconversion efficiency of mushrooms produced over substrate (BE%) used and the chemical composition of sporophores were evaluated. The concentration of mycelial mass, crude exopolysaccharide content and laccase enzyme activity were also determined at the supplemented SMS before inoculation, at 50% and 100% of colonization stages in the new cultivation and in the final re-utilized SMS. The laccase enzyme was extracted to examine SMS potential for the dephenolization of olive mill and winery wastewaters. Results showed that both Pleurotus species exhibited BE over 185%, demonstrating this bioprocess could represent a promising strategy to convert SMS into nutritional food. Data also indicate the strong positive impact that SMS could have in the solid wastes' management and agribusiness enhancement.

Upgrading Grape Pomace through Pleurotus spp. Cultivation for the Production of Enzymes and Fruiting Bodies.

Grape pomace, a by-product derived from winery industries, was used as fermentation media for the production of added-value products through the cultivation of two Pleurotus species. Solid-state (SSF), semiliquid (SLF), and submerged (SmF) fermentations were carried out using grape pomace as substrate. The effect of the different fermentations on the consumption of phenolic compounds, the production of mycelial mass and enzymes was evaluated using P. ostreatus and P. pulmonarius. The production of fungal biomass and enzymes was influenced by the fermentation mode. The maximum biomass values of ~0.5 g/g were obtained for both P. pulmonarius and P. ostreatus in SmF. Laccase production was induced in SSF and a maximum activity of 26.247 U/g was determined for P. ostreatus, whereas the highest endoglucanase activity (0.93 U/g) was obtained in the SmF of the same fungi. Analysis of phenolic compounds showed that both strains were able to degrade up to 79% of total phenolic content, regardless the culture conditions. Grape pomace was also evaluated as substrate for mushroom production. P. pulmonarius recorded the highest yield and biological efficiency of 14.4% and 31.4%, respectively. This study showed that mushroom cultivation could upgrade winery by-products towards the production of valuable food products.

Evaluation of Biomass and Chitin Production of Morchella Mushrooms Grown on Starch-Based Substrates.

Morchella sp. is one of the most expensive mushrooms with a high nutritional profile. In this study, the polysaccharide content of Morchella species was investigated. Specifically, mycelium growth rate, biomass production, sclerotia formation, and glucosamine and total polysaccharides content of six Morchella species grown on a starch-based media were evaluated. Submerged fermentations in potato dextrose broth resulted in a glucosamine content of around 3.0%. In solid-state fermentations (SSF), using potato dextrose agar, a high linear growth rate (20.6 mm/day) was determined. Increased glucosamine and total polysaccharides content were observed after the formation of sclerotia. Biomass and glucosamine content were correlated, and the equations were used for the indirect estimation of biomass in SSF with agro-industrial starch-based materials. Wheat grains (WG), potato peels (PP), and a mixture of 1:1 of them (WG-PP) were evaluated as substrates. Results showed that the highest growth rate of 9.05 mm/day was determined on WG and the maximum biomass yield (407 mg/g) on WG-PP. The total polysaccharide content reached up to 18.4% of dried biomass in WG-PP. The results of the present study proved encouraging for the efficient bioconversion of potato and other starch-based agro-industrial waste streams to morel biomass and sclerotia eliciting nutritional and bioactive value.

Valorization of spent oyster mushroom substrate and laccase recovery through successive solid state cultivation of Pleurotus, Ganoderma, and Lentinula strains.

Spent mushroom substrate (SMS) of Pleurotus ostreatus was supplemented with wheat bran and soybean flour in various proportions to obtain C/N ratios of 10, 20, and 30, and their effect was evaluated in successive cultivation of Pleurotus ostreatus, Pleurotus pulmonarius, Ganoderma adspersum, Ganoderma resinaceum, and Lentinula edodes strains with respect to mycelium growth rate, biomass concentration, recovery of the enzyme laccase and crude exopolysaccharides, and also with additional fruiting body production. All fungi showed the highest growth rate on unamended SMS (C/N 30), with G. resinaceum being the fastest colonizer (Kr = 9.84 mm day-1), while biomass concentration maximized at C/N 10. Moreover, supplementation affected positively laccase activity, with P. pulmonarius furnishing the highest value (44,363.22 U g-1) at C/N 20. On the contrary, L. edodes growth, fruiting, and laccase secretion were not favored by SMS supplementation. Fruiting body formation was promoted at C/N 30 for Ganoderma and at C/N 20 for Pleurotus species. Exopolysaccharide production of further studied Pleurotus strains was favored at a C/N 20 ratio, at the initial stage of SMS colonization. The obtained results support the potential effective utilization of supplemented SMS for laccase production from Ganoderma spp. and for new fruiting body production of Pleurotus spp.

Adaptation of Volvariella volvacea metabolism in high carbon to nitrogen ratio media.

Increasing glucose concentration (up to 46%) in the culture medium of Volvariella volvacea resulted in a noticeable biomass increase (63.82%) and intracellular polysaccharide (IPS) production (32.84% in absolute and 51.34% in relative values). In high glucose concentrations, decreased biomass yields on glucose consumed were recorded. A non-negligible lipid was synthesized (up to 32.22%, w/w) in the first growth steps. Lipids and IPS were decreased as fermentation proceeded. Fatty acid composition of lipids was not remarkably affected by the C/N ratio imposed, whereas the concentration of neutral lipids increased in the high C/N ratio media. Phospholipid was the richest in saturated fatty acids lipid fraction. The major phospholipid classes, i.e. phosphatidyl-choline and phosphatidyl-inositol, were differentially affected by the high glucose concentrations, as the former increased its proportion and the latter decreased. Linoleic was the predominate fatty acid. C/N ratio and fermentation time affected IPS composition, as glucose increased in a high C/N ratio, while fructose and mannitol decreased.

Patterns of major metabolites biosynthesis by different mushroom fungi grown on glucose-based submerged cultures.

The biosynthetic potential of four basidiomycetes (Agrocybe aegerita, Flammulina velutipes, Ganoderma applanatum and Pleurotus pulmonarius) and one ascomycete (Morchella esculenta) was examined in regard to biomass, intracellular (endopolysaccharides and lipids) and extracellular (exopolysaccharides) compounds' production in liquid media with glucose as substrate, in static and agitated cultures. Exopolysaccharides' production presented significant negative correlation with biomass, endopolysaccharides and lipids, while biomass was positively related to the production of endopolysaccharides and lipids. Maximum values of biomass, endo- and exo-polysaccharides obtained were quite impressive: P. pulmonarius produced 22.5 g/L of biomass, A. aegerita 60.4 % (w/w) of endopolysaccharides and F. velutipes 1.2 g/L of exopolysaccharides. Polysaccharides and lipids synthesized at the early growth stages were subjected to degradation as the fermentation proceeded. Mycelial lipids of all strains were highly unsaturated, dominated by linoleic acid, whereas glucose was the main building block of endopolysaccharides. The ability of the examined mushroom fungi to synthesize in high quantities biomass and polysaccharides, products with biotechnological and medicinal interest, renders these fungi as potential candidates in sugar-based bio-refineries.

Mushroom polysaccharides and lipids synthesized in liquid agitated and static cultures. Part II: study of Volvariella volvacea.

Volvariella volvacea strains were studied in relation with their ability to produce biomass, lipids and polysaccharides. Firstly, screening of four strains (AMLR 188, 190, 191 and 192) was performed in agar cultures, where the mycelial growth rate of the strains was measured, and in static liquid cultures, where the production of biomass, the biosynthesis of total cellular lipids and the consumption of glucose were monitored. For all strains, biomass production was significant (13-15 g l(-1)) and total lipid in dry weight (%, w/w) ranged from 3 to 12 %. Afterwards, a detailed kinetic analysis of mycelial biomass, extra- and intra- cellular polysaccharides (EPS, IPS, respectively) as well as lipid production by a V. volvacea selected strain was conducted in submerged static and agitated cultures. Maximum values of 15 g l(-1) biomass, ~1.0 g l(-1) EPS and 5.5 g l(-1) IPS were recorded. Agitation did not have severe impact on biomass, EPS and IPS production, but it increased total lipid in dry weight quantities. EPS, IPS and lipid in dry weight values decreased with time. Glucose was the major cellular carbohydrate detected. Total fatty acid analysis of cellular lipids was performed for all V. volvacea strains and linoleic acid (Δ9,12)C18:2 was predominant. Neutral lipids constituted the major fraction of cellular lipids, but their quantity decreased as fermentation proceeded. Phospholipids were the most saturated lipid fraction.

Mushroom polysaccharides and lipids synthesized in liquid agitated and static cultures. Part I: screening various mushroom species.

The effect of four synthetic media containing glucose (initial concentration 30 g l(-1)) on mycelial growth, exopolysaccharides (EPS) and cellular lipids production was examined in 11 mushroom species after 12 and 16 days of culture in static- and shake-flasks. Fatty acid analysis of cellular lipids produced was also performed. Agitation had a positive effect on biomass production, glucose consumption and lipid biosynthesis. Media that favoured the production of biomass were not suitable for EPS biosynthesis and vice versa. Biomass values varied from ~1.0 g l(-1) (Lentinula edodes) to ~19 g l(-1) (Pleurotus ostreatus), while the highest EPS quantity achieved ranged between 1.6 and 1.8 g l(-1) (for Ganoderma lucidum and L. edodes, respectively). Quantities of total cellular lipids varied between 2.5 and 18.5 % w/w, in dry mycelial mass for the fungi tested. Lipid in dry weight values were influenced by the medium composition. Cellular lipids presented noticeable quantities of poly-unsaturated fatty acids like linoleic acid. Compared to most of the mushrooms tested, lipids of Volvariella volvacea were more saturated. The ability of several mushroom species of our study to produce in notable quantities the above-mentioned added-value compounds renders these fungi worthy for further investigations.

Residue evaluation of famoxadone and trifloxystrobin in cultivated mushrooms.

Dissipation of the fungicides famoxadone and trifloxystrobin in basidiocarps of Agaricus bisporus was studied in mushroom growing rooms. The mushroom samples taken at all three consecutive production flushes following single or split applications of the fungicides were extracted with solvents and the residues were determined by using a gas chromatograph equipped with an electron capture detector (GC-ECD). Recoveries from the fortified control samples ranged from 87 to 105%. Following drench applications at 0.1-1 g/m2 of culture bed area, the highest famoxadone residue determined in basidiocarps was 0.1447 mg/kg. Analysis of trifloxystrobin revealed a quantitative relationship between the application rate (0.8-1.8 g/m2) and the residue levels of both the parent compound and its acid metabolite. The maximal combined residues of trifloxystrobin and its metabolite were 0.1313 mg/kg. Short- and long-term dietary risk assessment for both fungicides was carried out using consumption data from World Health Organization and the UK Pesticide Safety Directorate's Ten Consumer Model. The potential acute and chronic residue intakes via mushroom consumption were below toxicologically significant indicators.