Isolation and purification of all-trans diadinoxanthin and all-trans diatoxanthin from diatom Phaeodactylum tricornutum

Two diatom-specific carotenoids are engaged in the diadinoxanthin cycle, an important mechanism which protects these organisms against photoinhibition caused by absorption of excessive light energy. A high-performance and economical procedure of isolation and purification of diadinoxanthin and diatoxanthin from the marine diatom Phaeodactylum tricornutum using a four-step procedure has been developed. It is based on the use of commonly available materials and does not require advanced technology. Extraction of pigments, saponification, separation by partition and then open column chromatography, which comprise the complete experimental procedure, can be performed within 2 days. This method allows HPLC grade diadinoxanthin and diatoxanthin of a purity of 99 % or more to be obtained, and the efficiency was estimated to be 63 % for diadinoxanthin and 73 % for diatoxanthin. Carefully selected diatom culture conditions as well as analytical ones ensure highly reproducible performance. A protocol can be used to isolate and purify the diadinoxanthin cycle pigments both on analytical and preparative scale. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10811-016-0961-x) contains supplementary material, which is available to authorized users

Expression of three diadinoxanthin de-epoxidase genes of Phaeodacylum tricornutum in Escherichia coli Origami b and BL 21 strain

In the diadinoxanthin cycle the epoxy group is removed from diadinoxanthin and diatoxanthin is created. This conversion takes place e.g. in diatoms with the involvement of the enzyme diadinoxanthin de-epoxidase. In one of the diatom species, Phaeodactylum tricornutum (CCAP 1055/1 strain with genome sequenced) three de-epoxidase genes (PtVDE, PtVDL1, PtVDL2) have been identified, but only one of them (PtVDE) corresponds to violaxanthin de-epoxidase, an enzyme which is commonly found in higher plants. In these studies, the expression of two de-epoxidase genes of another Phaeodactylum tricornutum strain (UTEX 646), which is commonly used in diatom studies, were obtained in Origami b and BL21 E. coli strains. The molecular masses of the mature proteins are about 49 kDa and 60 kDa, respectively, for VDE and VDL2. Both enzymes are active with violaxanthin as a substrate

Zeaxanthin epoxidation : an in vitro approach

Zeaxanthin epoxidase (ZE) is an enzyme operating in the violaxanthin cycle, which is involved in photoprotective mechanisms. In this work model systems to study zeaxanthin (Zx) epoxidation were developed. Two assay systems are presented in which epoxidation of Zx was observed. In these assays two mutants of Arabidopsis thaliana which have active only one of the two xanthophyll cycle enzymes were used. The npq1 mutant possesses an active ZE and is thus able to convert Zx to violaxanthin (Vx) but the violaxanthin de-epoxidase (VDE) is inactive, so that Vx cannot be converted to Zx. The other mutant, npq2, possesses an active VDE and can convert exogenous Vx to Zx under strong light conditions but reverse reaction is not possible. The first assay containing thylakoids from npq1 and npq2 mutants of A. thaliana gave positive results and high efficiency of epoxidation reaction was observed. The amount of Zx was reduced by 25%. To optimize high efficiency of epoxidation reaction additional factors facilitating both fusion of the two types of thylakoids and incorporation of Zx to their membranes were also studied. The second kind of assay contained npq1 mutant thylakoids of A. thaliana supplemented with exogenous Zx and monogalactosyldiacylglycerol (MGDG). Experiments with different proportions of Zx and MGDG showed that their optimal ratio is 1:60. In such system, due to epoxidation, the amount of Zx was reduced by 38% of its initial level. The in vitro systems of Zx epoxidation described in this paper enable analysis some properties of the ZE without necessity of its isolation

Temperature effect on growth, and selected parameters of Phaeodactylum tricornutum in batch cultures

The effect of optimal and stress temperatures on the growth kinetics of the Phaeodactylum tricornutum CCAP/1055/1 strain (a model diatom with a known genome sequence) in batch cultures was examined. The analysis of the obtained results showed two phases of culture growth. There were significant positive correlations between OD increase of chlorophyll a chlorophyll c and protein concentration at different temperatures. The Fv/Fm parameter achieved a maximum level on the 6th or 7th day and then decreased to the values registered on the first day of observation. Genetic material undergoes gradual degradation 10 days after inoculation. The size of the cells was invariable

Moje doświadczenia edukacji osób z niepełnosprawnością. Autoetnografia

Celem badań jest zmiana w kompetencjach badawczych uczestniczek, nabycie wiedzy metodologicznej, umiejętność jej zastosowania w praktyce, jak również przyjrzenie się własnym doświadczeniom związanym z edukacją osób z niepełnosprawnościami. Autorki opisały doświadczenia, które były dla nich przełomowe, szokujące albo zmieniały ich sposób patrzenia na osoby z niepełnosprawnością, wykorzystując metodę autoetnografii, która dla większości była nową formą wypowiedzi pisemnej. Wykorzystując wątki autobiograficzne, autorki ujawniają refleksje nad doświadczeniem edukacyjnym. Praca nad artykułem pozwoliła autorkom na zmianę w podejściu do badań naukowych, na poznanie metody autoetnograficznej i zmianę w postrzeganiu opisywanej rzeczywistości społecznej. Każda autorka artykułu jest jednocześnie badaczką i badaną.The aim of the research is a change in research competency of participants, getting methodology knowledge, ability of using it in practise and reflection on their own experiences in education of people with disabilities. Using autobiography elements authors reveal their reflections on education experiences. Working on the article allowed authors to change their approach to scientific research, getting to know autoethnographic method and a change in perception of the described social reality. All article’s authors are researchers, as well as the ones who were examined

Comparison of Magnetic Resonance Enterography Findings and Clinical Index of the Disease Activity Score in Crohn\u27s Disease

Introduction Crohn\u27s disease (CD) is a chronic inflammatory disease of the gastrointestinal tract. It usually affects younger people, with the peak of illness between the age of 15 and 25. MR enterography (MRE) is a proven and reliable modality in the evaluation of the lesion extent within this group of patients

A retrospective observational study on patients intoxicated by drugs and other xenobiotics

Objectives Many research studies conducted in various toxicology centers point to drugs as the most common cause of intoxication. Long-term observations make it possible to clarify the nature of these poisonings. The aim of this study was to examine the trends and reasons of intoxication in patients hospitalized over a 10-year observation period (2005–2015), as well as to compare the number of patients poisoned with nonsteroidal anti-inflammatory drugs (NSAIDs), mainly over-the-counter (OTC) drugs. Material and Methods A retrospective observational study examined the medical documentation of patients hospitalized in the Department of Toxicology and Internal Diseases of the T. Marciniak Lower Silesian Specialist Hospital in Wrocław in 2005–2015, including the analysis of the causes of intoxication as well as total poisoning-related death statistics. Quarterly and annual analyses of the numerical data, and comparisons of the frequency of poisonings were included. The patient population from the area of Lower Silesia, Poland, was examined. Results The number of hospitalized patients has increased, with attempted suicide being the leading cause of death. Male intoxication and mortality have been found to predominate. Drugs are the most common cause of poisoning, and among these the most common are sedatives and psychotropic drugs. Intoxication due to NSAIDs, especially OTC drugs, increased significantly in the observation period. In 2005 no fatal cases were reported as a result of NSAID intoxication, while in 2015 mortality significantly increased to 43%. Conclusions The lack of a common trend in poisonings is observed but the number of hospitalized patients has increased, especially among young people, which is consistent with global trends. Drugs are the most common cause of mortality, and a significant increase in NSAID (mainly OTC) poisonings in particular indicates the growing prevalence of an uncontrolled use of these drugs. Int J Occup Med Environ Health. 2019;32(4):489–50