Impact of selected factors on extraction yield and composition of fatty acids in microbial oil produced from Yarrowia lipolytica yeast cells

Drożdże Yarrowia lipolytica to modelowy gatunek mikroorganizmów olejogennych, zdolny do kumulacji lipidów wewnątrz komórki w ilości powyżej 20 % suchej masy. Oprócz prac nad zwiększaniem wydajności biosyntezy oleju mikrobiologicznego ważnym zagadnieniem jest także dobór warunków ekstrakcji tłuszczu z biomasy drożdży. Celem pracy była ocena wpływu wybranych parametrów (rodzaju rozpuszczalnika, sposobu przygotowania próbki oraz czasu trwania procesu) na wydajność ekstrakcji tłuszczu zawartego w komórkach dzikiego szczepu drożdży Y. lipolytica KKP 379, prowadzonej w aparacie Soxhleta oraz na skład kwasów tłuszczowych w uzyskanym oleju. Hodowle okresowe drożdży prowadzono w bioreaktorze laboratoryjnym w bogatym podłożu YPO oraz w podłożach mineralnych z limitowaną zawartością azotu, co umożliwiło otrzymanie biomasy komórkowej o zróżnicowanej zawartości oleju mikrobiologicznego. W badaniach zastosowano plany kwadratów łacińskich, co pozwoliło na odmaskowanie efektu wpływu sposobu przygotowania próbki na zmienną zależną, jaką była zawartość tłuszczu w biomasie drożdży. Najlepsze rezultaty osiągnięto po przygotowaniu biomasy metodą lityczną z wykorzystaniem komercyjnego preparatu Y-PER. Zastosowany czas ekstrakcji oraz rodzaj rozpuszczalnika (eter naftowy lub heksan) nie wpłynęły istotnie na wydajność procesu. Rodzaj rozpuszczalnika miał z kolei wpływ na zawartość trzech kwasów tłuszczowych spośród czternastu zidentyfikowanych w lipidach ekstrahowanych z biomasy drożdży, tj. kwasu palmitooleinowego, eikozapentaenowego i dokozatetraenowego. Większą zawartość wymienionych kwasów oznaczono w próbkach ekstrahowanych heksanem.Yarrowia lipolytica yeasts are a model species of oleaginous microorganisms capable of accumulating lipids inside the cell in an amount exceeding 20 % of dry matter. In addition to studies on how to increase the efficiency of oil biosynthesis, an important issue is, also, the selection of conditions for lipid extraction from yeast biomass. The objective of the research study was to assess the impact of some selected parameters (type of solvent, sample treatment method, and the duration of the process) on the yield of extraction, run in an Soxhlet extractor, of lipids contained in cells of wild strain of Y. lipolytica KKP 379 yeast, and on the composition of fatty acids in the microbial oil produced. The yeasts were cultured in a batch process, in a laboratory bioreactor in a rich YPO medium, as well as in nitrogen restricted mineral media; thus, it was possible to obtain a cell biomass with varying contents of microbial oil. In the research, Latin Square Designs were applied so that the effect of biomass treatment method could be unmasked on the dependent variable, i.e. the content of lipids in the yeast biomass. The best results were achieved when the biomass was prepared using a commercial preparation of Y-PER. The applied duration time of extraction and the used type of solvent (petroleum ether or hexane) did not significantly impact the process yield. However, the type of solvent impacted the content of three fatty acids of the fourteen identified in the lipids extracted from the yeast biomass, ie. the content of palmitoleic, eicosapentaenoic, and docosatetraenoic acids. Higher contents of the above mentioned compounds were determined in the hexane extracted samples

Utilization of waste from food and fuel industries by lipolytic yeast of Yarrowia lipolytica

Przemysł spożywczy i paliwowy wytwarzają znaczne ilości trudnych w utylizacji hydrofobowych odpadów, stąd istnieje potrzeba poszukiwania nowych metod ich zagospodarowania. Celem badań była ocena możliwości zastosowania wybranych hydrofobowych odpadów przemysłu spożywczego i paliwowego jako głównego źródła węgla oraz induktora syntezy enzymów lipolitycznych w hodowli szczepu drożdży Yarrowia lipolytica W29. Hodowle wstrząsane prowadzono przez 65 h w 28°C w zmodyfikowanym podłożu YPG, w którym glukoza została zastąpiona olejem po procesie wędzenia ryb, tłuszczem po procesie wędzenia wędlin wieprzowych, tłuszczem po pieczeniu kaczej tuszki, zjełczałym masłem klarowanym lub zużytym olejem silnikowym. Stwierdzono, że odpady te mogą być wykorzystywane jako źródło węgla w hodowli drożdży Y. lipolytica. Produkcję enzymów lipolitycznych zaobserwowano w podłożach zawierających tłuszczowe substraty, a aktywność enzymów korelowano ze składem kwasów tłuszczowych. Wykazano, że istnieje możliwość utylizacji zastosowanych substratów odpadowych w procesach mikrobiologicznych do syntezy enzymów o aktywności lipolitycznej.Waste disposal and by-product management in many branches of industry pose problems in the areas of environmental protection and sustainability. Hydrophobic waste substrates of food and fuel origin stands for one of the continuously gaining ground for waste management fields. The aim of the study was to evaluate the possibility to use food origin wastes and fuel industry waste as a carbon sources in the culture medium for Y. lipolytica W29 with their simultaneous valorization. Culture media contained 2% of waste substrates. In the study there were evaluated yeast biomass yield, number of yeast cells in 1 cm³ of medium and extracellular lipase activity after 65 h of yeast growth on a rotary shaker at 28°C. Five wastes were estimated: oily waste from duck roasting process, oily waste from sausages smoking process, rancid ghee, oily waste from fish smoking process and waste engine oil. Additionally fatty acid composition of lipid waste was analyzed using gas chromatography. It was shown the possibility of using these wastes in cultivation of yeast with their simultaneous valorization by obtaining valuable products, e.g. enzymes such as extracellular lipases as well as biomass intended for feed. Yeast biomass yielded from 19.77 g DM·dm⁻³ for oily waste from duck roasting process to 12.28 g DM·dm⁻³ for oily waste from fish smoking process. It has been found that waste substrates stimulate the synthesis of extracellular lipases with different efficiency. The highest activity was obtained in medium containing smoked fish oil (0.313 U·cm⁻³). Furthermore, in waste engine oil medium no lipase activity of Y. lipolytica yeast was observed, but cells did grow and formed a biofilm. The analysis of fatty acid compositions showed the highest oleic acid content in oily waste from duck roasting process (41.9%), slightly lower waste from sausages smoking process (37.8%) and two-fold lower in fish oil (17.3%). This is very important information, because some authors believe that lipolytic enzymes synthesized by the yeast Y. lipolytica show substrate specificity as compared to that oil. Furthermore, the waste oil from fish smoking process was characterized by the presence of polyunsaturated fatty acids containing more than 20 carbons in chain length (EPA and DHA). There was no correlation between lipolytic activity and oleic acid content in waste fat used as the carbon source in medium, but it can be concluded that Y. lipolytica yeast prefered unsaturated rather than saturated fatty acids in extracellular lipase production

High rate of mosaicism in tuberous sclerosis complex.

Six families with mosaicism are identified in a series of 62 unrelated families with a mutation in one of the two tuberous sclerosis complex (TSC) genes, TSC1 or TSC2. In five families, somatic mosaicism was present in a mildly affected parent of an index patient. In one family with clinically unaffected parents, gonadal mosaicism was detected after TSC was found in three children. The detection of mosaicism has consequences for genetic counseling of the families involved, as changed risks apply to individuals with mosaicism, both siblings and parents. Clinical investigation of parents of patients with seemingly sporadic mutations is essential to determine their residual chance of gonadal and/or somatic mosaicism, unless a mosaic pattern is detected in the index patient, proving a de novo event. In our data set, the exclusion of signs of TSC in the parents of a patient with TSC reduced the chance of one of the parents to be a (mosaic) mutation carrier from 10% to 2%. In the five families with somatic mosaicism, the parent was given the diagnosis after the diagnosis was made in the child

Comparative Economics of a 12-Gene Assay for Predicting Risk of Recurrence in Stage II Colon Cancer

© 2014, The Author(s). Background: Prior economic analysis that compared the 12-gene assay to published patterns of care predicted the assay would improve outcomes while lowering medical costs for stage II, T3, mismatch-repair-proficient (MMR-P) colon cancer patients. This study assessed the validity of those findings with real-world adjuvant chemotherapy (aCT) recommendations from the US third-party payer perspective.Methods: Costs and quality-adjusted life-years (QALYs) were estimated for stage II, T3, MMR-P colon cancer patients using guideline-compliant, state-transition probability estimation methods in a Markov model. A study of 141 patients from 17 sites in the Mayo Clinic Cancer Research Consortium provided aCT recommendations before and after knowledge of the 12-gene assay results. Progression and adverse events data with aCT regimens were based on published literature. Drug and administration costs for aCT were obtained from 2014 Medicare Fee Schedule. Sensitivity analyses evaluated the drivers and robustness of the primary outcomes.Results: After receiving the 12-gene assay results, physician recommendations in favor of aCT decreased 22 %; fluoropyrimidine monotherapy and FOLFOX recommendations each declined 11 %. Average per-patient drugs, administration, and adverse events costs decreased $US2,339,$US733, and $US3,211, respectively. Average total direct medical costs decreased$US991. Average patient well-being improved by 0.114 QALYs. Savings are expected to persist even if the cost of oxaliplatin drops by \u3e75 % due to generic substitution.Conclusions: This study provides evidence that real-world changes in aCT recommendations due to the 12-gene assay are likely to reduce direct medical costs and improve well-being for stage II, T3, MMR-P colon cancer patients

Evaluation of shark cartilage in patients with advanced cancer: A north central cancer treatment group trial

BACKGROUND. Shark cartilage has been a popular complementary or alternative medicine intervention. The basis for this popularity is the claim that sharks rarely get cancer because of the high proportion of cartilage in the shark\u27s body. However, early studies were equivocal. Therefore, a clinical trial was conducted to look at the impact of shark cartilage in patients with advanced cancer. The primary goal of this trial was to determine whether a shark cartilage product improved overall survival for patients with advanced cancer who were getting standard care. Secondary research goals were to evaluate toxicities, tolerability, and quality of life associated with this shark cartilage product. METHODS. The study was a two-arm, randomized, placebo-controlled, double-blind, clinical trial. Patients with incurable breast or colorectal carcinoma had to have good performance status and organ function. Patients could be receiving chemotherapy. Patients were all to receive standard care and then to be randomly selected to receive either a shark cartilage product or an identical-appearing and smelling placebo 3 to 4 times each day. RESULTS. Data on a total of 83 evaluable patients were analyzed. There was no difference in overall survival between patients receiving standard care plus a shark cartilage product versus standard care plus placebo. Likewise, there was no suggestion of improvement in quality of life for patients receiving the shark cartilage, compared with those receiving placebo. CONCLUSION. This trial was unable to demonstrate any suggestion of efficacy for this shark cartilage product in patients with advanced cancer. © 2005 American Cancer Society