Extracellular Lipase Production Research Articles

Tepki Yüzey Metodu ile Aeromonas caviae LipT51'den Ekstrasellüler, Termo-Alkali Lipaz Üretiminin İstatistiksel Optimizasyonu

Extracellular thermo-alkaline lipase production from Aeromonas caviae LipT51 was statistically optimized by response surface methodology (RSM). First, the one factor at a time approach was implemented to screen the sources of carbon (olive oil, tributyrin, sunflower oil, waste frying oil, glycerol, Tween 80, Tween 20, palm oil, and Triton X100) and nitrogen (peptone, yeast extract, tryptone, whey, urea, NaNO2, NH4NO3) for the highest lipase production. Then, optimum values for waste frying oil selected as carbon source, tryptone selected as nitrogen source and initial pH of the medium were determined by RSM using Box-Behnken design (BBD). The quadratic model of BBD for lipase production was statistically significant and reliable (p < 0.0001, R2 = 0.9881). The validated optimal conditions for maximum lipase production (1.6 U mL-1) were determined as 1.13% waste frying oil, 1.5% tryptone and pH 7.9. For the first time in this study, optimization of lipase production from an A. caviae strain was carried out and under optimized culture conditions using cheap waste material. The production efficiency of lipase enzyme, which is known to be valuable with its detergent activity, increased 2.7 times compared to non-optimized conditions.

Process parameter studies by central composite design of response surface methodology for lipase activity of newly obtained Actinomycete

Actinomycete strains from cottonseed soap-stock (a dark gelatinous waste from oil refinery) were obtained. Cultures were analyzed for the enzyme lipase, protease and cellulase production using plate assay(s). Best lipolytic isolates were further quantitatively analyzed for lipase production, using substrate p-nitrophenyl palmitate (p-NPP). Cultures were characterized by morphological and molecular studies. The potent lipase producing strain of actinomycete was optimized for pH, temperature, inoculum size, carbon and nitrogen sources. Further, the lipolytic strain of actinomycete was grown on synthetic media and growth factors were optimized by RSM (Response Surface Methodology) for maximum production of lipase. Total 49 cultures were obtained from enriched soap-stock samples; among them 7 isolates were strains of actinomycete. Potent actinomycete strains were identified as Nocardiopsis alba , Streptomyces leeuwenhoekii , Streptomyces caelestis , Streptomyces werraensis and Streptomyces sclerotialus . Based on quantitative lipase production study, Nocardiopsis alba was selected as the best lipase producer. Optimized results of process parameter studies for lipase production by Nocardiopsis alba were: 5mL inoculum size, temperature 40 ∘ C, shaking at 130 rpm, pH 8.0 and cottonseed oil as the best carbon source. Cottonseed oil, K 2 HPO 4 and cellulose were noted as main factors as optimized by Central Composite Design (CCD), where the culture showed highest (65.78 U/mL/min) lipase production. Findings of the research would offer potential environmental benefits along with socio-economic benefits if implemented suitably. • Response surface methodology has been reported to study Lipase production by Actinomycete • Analysis of variance (ANOVA) has been used in this study. • Highest extracellular lipase production was observed at 40°C at pH 8 of the growth medium. • Nocardiopsis alba can be used on higher scale under optimized process parameters.

Enhanced production of extracellular lipase by ethyl methane sulfonate from soil fungal strain in submerged and solid-state fermentation condition

Enhancement in the production of enzyme by utilizing different strains of microbe is one of the main prospects in biotechnology. In the present work, ethyl methane sulfonate (EMF) was selected as the chemical mutagen for inducing mutagenesis in fungi. It is a cheap method to induce random mutation as compared to other methods of recombinant technologies. Strain improvement was done by incubating the fungal spore suspension at variable concentrations of EMS i.e. 4% (v/v) and 10% (v/v) for the time period of 60, 90, and 120 min respectively. The set of control was treated with distilled water only. The fungal colonies were found to be maximum in control plate as compared to the EMF exposed plates. The number of fungal colonies was reduced as we raised the exposure time of EMF. Specific activity and the lipase activity of wild strain and hyperproducer were evaluated under the submerged (SmF) and solid-state fermentation (SSF). The wild strain denoted the 3.2 U/ml/min of enzymatic activity under SmF and 15.87 U/g/min of activity under SSF. In contrast, the best enzymatic activity was represented by S2St1 at 10% of EMS after the time period of 60 min i.e. 11.7 U/ml/min under SmF and 99.35 U/g/min under SSF after the time period of 72 hrs. Statistical analysis by using one-way ANOVA determined that the value of F calculated was lower than the F tabulated. So, there was a significant relation between the EMS percentage and time of exposure among the mutated strains. In conclusion, this soil fungal strain can be utilized to produce lipase enzyme for numerous industrial applications.

Palm oil wastes as feedstock for lipase production by Yarrowia lipolytica and biocatalyst application/reuse.

Palm oil production chain generates a greasy residue in the refining stage, the Palm Oil Deodorizer Distillate (PODD), mainly composed of free fatty acids. Palm oil is also used industrially to fry foods, generating a residual frying oil (RFO). In this paper, we aimed to produce lipase from palm agro-industrial wastes using an unconventional yeast. RFO_palm, from a known source, consisted of 0.11% MAG + FFA, 1.5% DAG, and 97.5 TAG, while RFO_commercial, from a commercial restaurant, contained 6.7% of DAG and 93.3% of TAG. All palm oil wastes were useful for extracellular lipase production, especially RFO_commercial that provided the highest activity (4.9 U/mL) and productivity (465 U/L.h) in 75h of processing time. In 48h of process, PODD presented 2.3 U/mL of lipase activity and 48.5 U/L.h of productivity. RFO_commercial also showed the highest values for lipase associated to cell debris (843 U/g). This naturally immobilized biocatalyst was tested on hydrolysis reactions to produce Lipolyzed Milk Fat and was quite efficient, with a hydrolysis yield of 13.1% and 3-cycle reuse. Therefore, oily palm residues seem a promising alternative to produce lipases by the non-pathogenic yeastY. lipolytica and show great potential for industrial applications.

Orchestration an extracellular lipase production from Aspergillus niger MYA 135: biomass morphology and fungal physiology

The impact of biomass morphology and culture conditions on fungal fermentation was widely reviewed in the literature. In this work, we presented three independent experiments in order to evaluate the influence of some of those input factors on a lipase production separately by using the Aspergillus niger MYA 135 and the two-stage fermentation technique. Regarding the culture modality, the biomass was pre-grown in a first reactor. Then, the washed mycelium was transferred to a second reactor to continue the study. Firstly, linear effects of fungal morphology and several physiological parameters on a lipase production were explored using the Plackett–Burman design. The dispersed fungal morphology was confirmed as a proper quality characteristic for producing an extracellular lipase activity. Concerning the impact of the carbon source on the biomass pre-growth, the sucrose (E = 9.923, p < 0.001) and the l-arabinose (E = 4.198, p = 0.009) presented positive and significant effects on the enzyme production. On the contrary, the supplementation of 0.05 g/L CaCl2 displayed a highly negative and significant effect on this process (E = − 7.390, p < 0.001). Secondly, the relationship between the enzyme production and the input variables N:C ratio, FeCl3 and olive oil was explored applying the central composite design. Among the model terms, the N:C ratio of the production medium had the most negative and significant influence on the enzyme synthesis. Thus, it was concluded that a low N:C ratio was preferable to increase its production. In addition, the bifunctional role of FeCl3 on this fungus was presented. Thirdly, a prove of concept assay was also discussed.

Statistical experimental design applied to extracellular lipase production by the marine Antarctic yeast Leucosporidium scottii CRM 728

Abstract The search for microbial enzymes from extreme environments may result in the discovery of biomolecules with different properties. In this sense, the aim of this study was to evaluate and optimize the conditions for lipase production by the marine-derived Antarctic yeast Leucosporidium scottii CRM 728 using an experimental design with different sources of carbon and nitrogen. The applied strategy was composed of three steps, including Plackett-Burman, Central Composite Design (CCD), and validation assay. Among nine variables applied, corn steep liquor, olive oil, and soybean oil showed a positive effect in the lipase production, while urea showed a negative effect. The production of lipase increased 9-fold in comparison to the basal activity when olive oil and corn steep liquor were used as nutrient sources. On the other hand, there was an increase of 4.8-fold when soybean oil and corn steep liquor were used as nutrient sources. The highest amounts of lipase were achieved in non-saline conditions and at 20.0 °C. In this study, the lipase production by the marine-derived psychrotolerant yeast L. scottii was for the first time reported, providing new knowledgement in the field of enzymatic production by extremophilic and marine microbial resource.

Kinetic and thermodynamic characterization of novel alkaline lipase from halotolerant Bacillus gibsonii.

A halotolerant bacterial strain isolated and identified as Bacillus gibsonii was used for extracellular lipase production. The bacterial strain was able to grow up to 1200mM salt concentration and showed maximum growth at 600mM NaCl concentration. The present study includes production of extracellular lipase enzyme and characterization of partially purified lipase with respect to its kinetic and thermodynamic behaviour. Maximum lipase activity was observed at 60°C under alkaline pH (9.0) condition. The kinetic parameters such as Vmax, Km and Kcat were calculated as 158.73U/mL, 0.539mM and 483.93min-1 at 60°C, respectively, suggested thermostable nature of the enzyme. The thermal inactivation energy [Ea(d)] was calculated as 66.98kJ/mol. The values of Gibb's free energy (86.31kJ/mol), enthalpy (64.26kJ/mol) and entropy (- 66.21 × 10-3kJ/mol/K) for the enzyme inactivation obtained at 60°C corroborated the assumption that 60°C was the optimum temperature. Further, the deactivation rate constant (kd) values calculated at 60°C and 80°C were found to be 0.0907 and 0.182min-1, respectively, which suggested that enzyme was more stable at 60°C and it was partly inactivated at 80°C.

Production, Purification and Characterization of Extracellular Lipase from a Mutated Strain of Penicillium Citrinum KU613360

In the present study lipase production, purification and characterization were carried out with a novel fungal strain of Penicillium citrinum KU613360 isolated from vegetable oil contaminated soil samples collected from oil mills located in and around Guntur District, Andhra Pradesh, India. The strain improvement was carried out by subjecting the strain to both UV and Ethidium Bromide treatments. The wild strain of P. citrinum KU613360 showed maximum lipase activity of 1.053±0.32IUmL-1 on optimized medium and while the mutated strain treated with combination of UV (300 sec) and Et Br (200 µgcm3), recorded the enzyme activity of 4.260±0.011IUmL-1, using the optimised medium at 6.5 pH and 40°C temperature. Thus, a 404% enhancement in the activity was achieved by using induced mutation on wild strain of P. citrinum KU613360. The molecular weight of the purified lipase from the mutated strain was found to be 35 kDa, when analysed on SDS PAGE. From our results it was concluded that the mutated strain has considerable capability and potentiality to be used in various industrial applications.

Profiling of antimicrobial metabolites of plant growth promoting Pseudomonas spp. isolated from different plant hosts.

The online version contains supplementary material available at 10.1007/s13205-020-02585-8.

Enhancing Lipase Biosynthesis by Aspergillus Melleus and its Biocatalytic Potential for Degradation of Polyester Vylon-200

Herein, response surface methodology (RSM) was employed to optimize the bioprocess parameters for improved production of extracellular lipase by Aspergillus melleus and evaluated its biocatalytic potential for degradation of polyester vylon-200. Our previous report showed that pH, incubation time, temperature, and additional nitrogen source had significant effects on lipase biosynthesis. The variance analysis revealed that the established RSM model based on a central composite design for lipase production was significant (p < 0.0001, R2 = 0.9925). Under the optimized bioprocess conditions of pH 5.68, incubation time 96 h, temperature 30 °C, and diammonium tartrate as a nitrogen source, maximum lipase titer of 1346.87 U/gds was achieved, 1.92-fold higher than lipase yield in basal medium. The optimally synthesized cell-free lipase extract was partially purified by ammonium sulfate fractionation and dialysis and used to degrade polyester vylon 200. The degradation profile revealed that the lipolytic enzyme demonstrated excellent hydrolytic potential resulting in a 76% weight of polyester vylon-200. Differential scanning calorimetry revealed a noticeable decrease in the glass transition temperature of PV-200 (from 72.6 oC to 63.9 oC). Scanning electron microscopy envisaged various micron-scale cracks and holes on the surface of film after enzymatic treatment. Likewise, significant weight loss of the PV-200 films was also corroborated by FTIR analysis. This study's findings illustrate lipase's potential as a green and ecofriendly biocatalyst for robust polyester degradation and depolymerization.

Screening of microfungi for lipolytic activity and optimization of process parameters in lipase production by solid substrate fermentation using selected microfungi (Penicillium aurantiogriseum)

In the present study, eighty-four strains of microfungi were screened for lipolytic activity, and Penicillium aurantiogriseum showing the highest enzyme activity was selected. Then extracellular lipase production from this species was carried out using the solid state fermentation (SSF) method, in which various agricultural wastes were used as substrates. Among the various agricultural wastes used in SSF, sunflower pulp was determined as the best solid substrate. Optimum fermentation conditions were found as follows: 6-day incubation time at 25 oC and moisturizing liquid pH: 5.5 (distilled water), initial moisture level 75% (w/v), initial inoculum concentration 1 ml (1x106 spores/ml), and 1% sesame oil as a carbon source. As a result of the optimum conditions, the volume activity was recorded as 0.4 U/ml, and the specific enzyme activity was obtained as 1.13 U/mg.

Three different C: N ratios for Pacific white shrimp, Penaeus vannamei under practical conditions: Evaluation of growth performance, immune and metabolic pathways

A 28-day experiment was carried out to evaluate different C/N ratios (10:1, 15:1 and 20:1) on growth performance, water quality, survival rate and gene expression related to metabolic pathways of Penaeus vannamei in a biofloc system. To maintain the C/N ratio of the feed input, different carbon sources (molasses, rice bran and wheat flour) in combination were applied after fermentation for a day with biofloc consortium. Results showed that shrimps reared in C/N 10 (630 mg) and C/N 15 (646 mg) biofloc systems showed significantly (p = .0229 and .0263) higher growth when compared to C/N 20 (528 mg) and control (374 mg). With regard to the production of extracellular enzymes, protease, lipase and xylanase were found predominantly in colonized bacteria isolated from biofloc treatments, whereas amylase was commonly found in all the treatments. The proPO activity was considerably increased in C/N 10 and 15 treatments than C/N 20 and control. The ranges of relative expression levels of potential genes entailed in metabolite pathway like hexokinase (HK- 6.57–8.17), pyruvate kinase (PK-5.79–7.96), crustacean hyperglycaemic hormone (CHH- 6.99–7.66) and fatty acid synthase (FAS- 6.57–8.86), triacylglycerol lipase (TGL-6.99–8.66) and immune gene (Toll receptor- 4.57–4.86) fold up-regulated expression were observed in the biofloc treatments than that of the control. The overall findings indicate that the rearing of P. vannamei shrimp in C/N 10 and 15 biofloc systems could improve the growth performance, water quality and development of immune and metabolic responses.

Multilayered Nano-Entrapment of Lipase through Organic-Inorganic Hybrid Formation and the Application in Cost-Effective Biodiesel Production.

Significant components of cost-effective medium for Magnusiomyces capitatus A4C extracellular lipase (ECL) production were optimized via a five-level factorial design. A simplistic, economical, and green approach was adopted for biomimetic mineralization to prepare multilayered nano-entrapped ECL, which were then applied as biocatalysts for the production of fatty acid methyl ester (FAME). The optimal ECL (0.8mg protein/mL) and CuSO4∙5H2O (1.2mM) showed the highest capacity for enzyme loading. The ECL-CuSO4-hybrid showed an 89.7% conversion of triacylglycerides into FAME via transesterification and a 98.7% conversion of oleic acid into FAME via esterification at 72h. The ECL-CuSO4-hybrid gave 65% and 78.7% FAME production after 5 successive reuses via transesterification and esterification reactions, respectively. Therefore, these ECL-inorganic hybrid biocatalysts have high economical potential to be used for the production of biodiesel as the future petrodiesel replacement.

Characterization of Thermophilic Microorganisms in the Geothermal Water Flow of El Chichón Volcano Crater Lake

This study reports for the first time the isolation, identification and characterization of lipase-producing thermophilic strain from the geothermal water of the El Chichón volcano crater lake. Two strains were identified by 16S rRNA sequencing as Geobacillus jurassicus CHI2 and Geobacillus stearothermophilus CHI1. Results showed that G. jurassicus CHI2 is Gram-positive, able to ferment maltose, fructose and sucrose and to hydrolyze starch and casein; while G. stearothermophilus CHI1 showed to be Gram-variable, able to ferment maltose and fructose and to hydrolyze starch. Colonies of both strains presented irregular shape, umbilicated elevation of gummy texture and cells presented flagellar movement to survive in fluids with high temperature and mass gradients due to complex phenomena of heat and mass transfer present in the geothermal fluids. Lipase production for G. stearothermophilus CHI1 was also evaluated. It was found that this strain possesses a growth associated with extracellular lipase production with a high activity of 143 U/mL at 8.3 h of incubation time, superior to the activities reported for other microorganisms of genus Geobacillus; for this reason, it can be said that the thermal flow of the El Chichón volcano crater lake can be a useful source of lipase-producing thermophilic bacteria.

Molecular, physiological, and biochemical characterization of extracellular lipase production by Aspergillus niger using submerged fermentation.

BackgroundExtracellular production of fungal lipases especially the lipases obtained from the Aspergilli has gained immense interest in recent years due to its diverse biotechnological applications. In this study, we focused on determining the fermentation parameters required for the optimal lipase production.MethodsA total of 256 fungal isolates were obtained from oil seeds. From each genus, one isolate was selected to evaluate lipase production using phenol red and tributyrin plate assays. Lipase activity was estimated using the spectrophotometric pNPP hydrolysis assay. The highest lipase producer isolates were identified using 18S ribosomal RNA gene sequencing. The genetic variability was determined by random amplified polymorphic DNA (RAPD) analysis and the dendrogram was constructed using the unweighted pair group method with arithmetic averages method. The isolates were examined in a submerged fermentation culture (Smf) to measure the effect of temperature, pH, incubation time, carbon source, nitrogen source, inoculum volume, and lipid source on lipase production.ResultsEleven isolates belonging to the genus Aspergillus were analyzed for lipase production where they were found to be the highest lipase producers among various fungal genera. All the tested isolates were identified as A. niger using 18s rRNA sequencing. Genetic diversity was evaluated among all of the studied A. niger isolates using RAPD primers. The RAPD primers were used to amplify 285 loci, of which five were polymorphic (1.75%) and seven were monomorphic (2.45%). Thus, a high level of genetic diversity was observed among all isolates. The tributyrin test and the lipase activity assay identified five strains of A. niger as high lipase producers, and their optimal enzyme activities were 709.74, 532.54, 735.64, 794.62, and 787.69 U/ml. The optimal conditions for lipase production were as follows: 40 °C, pH 7.5, 1% fructose as the carbon source, 1% yeast extract as the nitrogen source, 2% palm oil, 2.5 × 107 spores/ml suspension, and 3 days of incubation.ConclusionsThe current study provides a comprehensive characterization of the optimal conditions, which are essential to enhance lipase production in five A. niger isolates.

Optimization of extracellular lipase production by halotolerant Bacillus sp. VITL8 using factorial design and applicability of enzyme in pretreatment of food industry effluents

The effect of macro and micronutrients of media components on lipase production by Bacillus sp. VITL8 was investigated using classical as well as statistical methods. Initially, the carbon source, nitrogen source, inducer and metal ions that affect lipase production were selected using the classical approach. Subsequently, selected nutrients along with other key factors (such as pH, agitation rate, gum acacia and tween 80) were investigated using Placket Burman design. Finally, three significant factors, viz., olive oil, peptone and tween 80 were studied using a 22 full factorial central composite design. Under optimized condition [6% (v/v) of olive oil, 0.7% peptone, 0.9% tween 80 and 25 h of incubation], the enzyme production was found to be 2.2 times higher with an overall enzyme production of 325.0 ± 1.4 U mL−1. Laboratory scale experiment proved that the enzyme could be utilized for pretreatment of food industry effluent rich in fat and oil (dairy, bakery and poultry). The enzyme was capable of hydrolyzing more than 50% of the initial fat present in all these effluents and enabled the reduction in the levels of biological oxygen demand (BOD) and chemical oxygen demand (COD) of the effluent samples. The study thus reveals the utility of the lipase produced by the halotolerant bacterium Bacillus sp. VITL8 in the pretreatment of industrial effluents contaminated with oil and fat.HighlightsLipase production was enhanced by 2.2-fold using statistical methodsOne of the few reports on lipase production by a halotolerant bacterium, especially by Bacillus sp.Production of 325.0 ± 1.4 U mL−1 lipase within 25 h by a halotolerant bacteriumPretreatment of food industry effluents using Bacillus sp. VITL8 lipaseImprovement in effluent quality within 8 h of treatment

Production and Optimization of Extracellular Lipase Expressed by Aspergillus flavus Isolated from Petroleum-Contaminated Soil

Production and Optimization of Extracellular Lipase Expressed by Aspergillus flavus Isolated from Petroleum-Contaminated Soil

Arthrographis curvata and Rhodosporidium babjevae as New Potential Fungal Lipase Producers for Biotechnological Applications

Abstract Fungi have always attracted a lot of attention as they are able to produce a vast repertoire of enzymes that find a broad spectrum of uses in biotechnological and industrial fields. Undoubtedly, one of the most promising biocatalysts is the lipase, which has been widely used for the biotransformation of a number of commercial products due to its high stability, high catalytic efficiency, versatility and selectivity, making it one of the most attractive and best-studied enzymes. In this study we report the isolation and molecular identification of new lipase-producing fungi from different environmental samples from Morocco. The production and activity of extracellular lipases, at different parameters, was evaluated using the Rhodamine B agar, submerged fermentation and biochemical methods. Two fungal strains Arthrographis curvata and Rhodosporidium babjevae, were isolated and found to produce large amounts of lipases. The optimal activity of the extracellular lipase was detected at 40°C and pH 9.0 for A. curvata and at 40 °C and pH 8.0 for R. babjevae. This study add new information at the growing list of fungal species producing lipases with improved physicochemical proprieties which could constitute a new line of research for further studies and to be exploited for industrial or bioremediation purposes.

Enhancement of lipase production by ethyl methane sulfonate mutagenesis of soil fungal isolate

Strain improvement through random mutagenesis is an extremely developed practice and it plays an important role in the economic growth of microbial agitation processes. The present study comprises of genetic improvement of fungus isolated from petrol pump soil by ethyl methane sulfonate (EMS) mutagenesis for increased production of extracellular lipase. Random mutagenesis was performed by incubating the spore suspension of fungus with EMS at a concentration of 5% (v/v) and 8% (v/v) for 30, 60 and 90 min, respectively. Control set was prepared by incubating the spore suspension with sterile distilled water. Control plate showed maximum number of fungal colonies whereas number of colonies was decreased as we increased exposure time of EMS from 30 to 90 min. The lipase activity of six mutagenic strains and wild strain was determined under submerged fermentation and solid state fermentation. Treated culture named as EMS5%-60min (obtained after 60 min exposure with 5% EMS) exhibited maximum activity (32.09 ± 1.84 IU/ml/min) in SmF as compared to wild strain (8.77 ± 3.52 IU/ml/min) and another treated strain named as EMS8%-90min (obtained after 90 min exposure with 8% EMS) exhibited maximum activity (7.99 ± 0.12 IU/g/min) in SSF as compared to wild strain (1.77 ± 0.71 IU/g/min). The activity of mutagenic strain i.e. EMS5%-60min was increased to 365.90% as compared to 100% activity of wild strain in SmF whereas activity of another mutagenic strain i.e. EMS8%-90min was increased to 451.41% as compared to 100% activity of wild strain in SSF.

Newly Isolated Lipolytic and Oleaginous Fungal Strain, Production, Optimization and Biochemical Characterization of the Extracellular (phospho)lipase

The present study investigates the biochemical characterization of an extracellular (phospho)lipase from a wood fungus Peziza sp. (medium optimization, inducer concentration and substrate specificity measurements). The strain was identified on the basis of ITS1/ITS4 primers. A 604 bp fragment was amplified by PCR and the obtained nucleotide sequence, showed 99% identity with the ITS region of isolates named as Peziza sp. Interestingly, Peziza sp. has both lipase and phospholipase activities with the same level which require both the presence of Ca2+ and bile salts. Our result shows that the lipase hydrolyzes preferably the olive oil at 45 °C, pH 8. Whereas, the phospholipase activity was detected on pure PC at 45 °C, pH 9. Lipid extraction from dry biomass using chloroform/methanol (2/1) and quantitative measurement using electron microscope showed that intracellular triglycerides content was significantly high and reaches 20.88%. Gas chromatography analysis shows a majority of C18:1 (76, 98%) and C18:2 (9, 33%). Whereas, saturated fatty acids ranging C16–C20 represent only 11.5% of total lipids composition. Lipid accumulation test in Pezizales sp under fluorescence microscope