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1

Adsorbenty ze związanymi cieczami jonowymi i ich wykorzystanie w przygotowaniu próbek oligonukleotydów

Nuckowski Łukasz, Zalesińska Ewa, Studzińska Sylwia

Wiadomości Chemiczne

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2020

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[Z] 74, 7-8

545--565

EN

Oligonucleotides are short fragments of nucleic acids. They have a growing potential in medicine, especially as diagnostic and therapeutic agents. In most cases, these compounds are determined in the complex biological matrix. Thus, the sample preparation step is very important in their bioanalysis. Solid-phase extraction is a predominant technique in this field. However, presently used for this purpose adsorbents have disadvantages. They ensure low extraction effectiveness and procedures using them are labor-intensive or time-consuming. Ionic liquids, since their discovery, are objects of intensive interest of scientists. Their scientific attractiveness is connected with their unique properties. They are used in separation and sample preparation techniques, such as liquid-liquid extraction using water-immiscible ionic liquids. This approach was also used in the extraction of oligonucleotides. Adsorbents modified with ionic-liquids have growing potential in extraction techniques. Few types of materials are used, namely carbon, polymers, and silica. A common feature of these materials modified with ionic liquids is the ion exchange character. Nonetheless, carbon nanomaterials are coated or covalently modified with ionic liquids, and they are used mainly for nonpolar compounds. Polymer and silica-based adsorbents are used mainly for acidic compounds. Polymers are characterized by the highest stability of the presented materials. Due to their ion-exchange properties crosslinked poly(ionic liquids) were used also for extraction of unmodified and modified oligonucleotides. The optimized procedure applying the material with bonded zwitterion ionic liquid gives high recoveries. It is concurrent for presently used adsorbents, thus solves problems connected with their usage. Moreover, it can be used for biological samples without any pre-purification.

2

Klasyczne i nowe metody przygotowania próbek oligonukleotydów antysensownych przed analizą chromatograficzną

Nuckowski Łukasz, Kilanowska Anna, Studzińska Sylwia

Analityka : nauka i praktyka

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2020

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nr 1

4--11

PL

Chromatograficzna analiza oligonukleotydów antysensownych wymaga przygotowania próbek. Zwykle wiąże się to z czasochłonnymi procedurami lub też koniecznością zastosowania odczynników znacząco wpływających na czułość metod chromatograficznych.

3

Chromatograficzna analiza związków budujących kwasy nukleinowe

Studzińska S., Rola R., Łobodziński F., Krzemińska K.

Wiadomości Chemiczne

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2016

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[Z] 70, 9-10

633--656

EN

Understanding the characteristics, role and structure of nucleic acids allowed to answer questions about the disease processes. Today, nucleic acids and their constituents are tools, which are used by molecular biology in medicine and biotechnology. Antisense and gene therapy are intensively developing methods for possible treating or preventing disease. They use short fragments of DNA or RNA - oligonucleotides to silence the genes expression. They are not the only ones that allow analytical chemists to obtain information about the state of our body. Determination of modified nucleoside allows detection of cancer, while analysis of nucleotides allows the estimation of strengthening the immune system. There is a great need of sensitive, selective and precise methods of separation of nucleosides, nucleotides and oligonucleotides and their qualitative and quantitative analysis. Consequently liquid chromatography (LC) is the most commonly used for analysis of nucleic acid constituents. The most widely used modes of LC include Ion Exchange Chromatography (IEC) and Reversed Phase High Performance Liquid Chromatography (RP HPLC). Both techniques have their advantages and disadvantages in the analysis of nucleosides, nucleotides and oligonucleotides. In the case of IEC it is necessary to use high concentrations of the salt in the mobile phase or concentration gradients, which considerably limits the possibility of using MS detection. RP HPLC can be coupled with MS detection but only when volatile salts are mobile phase components. On the other hand there is a significant problem is the lack of sufficient selectivity for the most polar nucleosides and nucleotides. RP HPLC MS is still most often used in the determination of nucleosides and nucleotides, due to its high sensitivity and a comprehensive qualitative analysis. Another system used for the HPLC analysis of oligonucleotides is Ion Pair Reversed Phase High Performance Liquid Chromatography (IP RP HPLC). These compounds can not be analyzed by RP HPLC due to their high polarity. The advantage of IP RP HPLC is selectivity, achieved by a suitable choice of mobile phase composition and the possibility of using MS. A disadvantage of IP RP HPLC in the analysis of oligonucleotides is however lower sensitivity compared to RP HPLC. During the last few years Hydrophilic Interaction Liquid Chromatography (HILIC) was applied for the separation of mixtures of nucleosides, nucleotides, oligonucleotides extracted from a biological or food samples. The presented results demonstrate the usefulness of this method, however, the resolving power is limited due to the asymmetric peak shape. On the other hand proper selection of the mobile and stationary phase can lead to a high selectivity in the analysis of the most polar nucleosides, nucleotides and oligonucleotides, which can not be separated by RP HPLC.

4

Zastosowania „click chemistry” w modyfikacjach nukleozydów i oligonukleotydów

Gładysz M., Milecki J.

Wiadomości Chemiczne

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2014

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[Z] 68, 7-8

617--643

EN

Since the year 2001 new ideology of clean and simple synthesis in organic chemistry has been established. The outstanding scientists Meldal and Sharpless presented their concepts of Click Chemistry. Among the reactions chosen for this concept the reaction of Copper(I) Catalyzed Alkyne-Azide Cycloaddition (CuAAC) became the most popular one. It is the basis of syntheses employed for building blocks synthesis in medicinal chemistry and material science. Libraries of potentially pharmacologically active anticancer and antivirus compounds possessing neutral triazol linkage could be easily obtained. Remarkable efficiency of CuAAC reaction influenced on DNA- and RNAbased synthesis of novel oligonucleotides derivatives. Many of nucleic acid molecular modifications found applications in enzymatic transformation, nucleic acid hybridization, molecular tagging and gene silencing. The CuAAC reaction allows for introducing modifications into practically every region of nucleoside/nucleotide/ oligonucleotide. This includes versatile modifications of the base moiety both aiming at the base pairing ability or specific labeling of the nucleoside unit. Different conjugates (bio-, fluorescent-, affinity- or spin labels) are being attached to the base part of the nucleic acid taking advantage of the presence of azide or alkyne substituents, which can be installed without great difficulty. Labeling at the sugar part of the nucleoside can be realized at the position 2’, 3’ or 5’, the latter two giving rise to the end-labeled oligonucleotides and the 2’ position serving as the attachment point for labeling inside the oligonucleotide chain. These kind of nucleic acid modifications are very promising. Versatility of CuAAC reactions is demonstrated by numerous examples of introducing modifications into practically every reactive site of the nucleotide/oligonucleotide molecule. The review systematically presents application of the “click” technique for modification of nitrogenous base, sugar or pseudosugar moiety or phosphorus center. Possibility of creating new kind of chain linkage, devoid of negative charge and nuclease resistant is also shown. This allows to design a new class of nucleic acid analogues, similar in its DNA-mimicking properties to PNA’s.

5

Biochromatografia oligonukleotydów

Pietrzak L., Studzińska S., Buszewski B.

Analityka : nauka i praktyka

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2014

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nr 1

20--25

PL

Oligonukleotydy to krótkie, jednoniciowe fragmenty kwasów nukleinowych. Ich podstawowym budulcem są nukleotydy składające się z trzech elementów: reszty fosforanowej, zasady azotowej oraz pentozy. Najważniejszą fizyczną właściwością oligukleotydów jest ich hydrofobowo-hydrofilowy charakter.

6

Od syntezy oligonukleotydów do Mycoplasma laboratorium

Laskowski M.

LAB Laboratoria, Aparatura, Badania

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2011

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R. 16, nr 3

22-25

7

Dynamics of methanogenic archaeal communities based on the rrna analysis and their relation to methanogenic activity

Walczak M., Lalke-Porczyk E., Idzikowski W.

Environment Protection Engineering

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2011

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Vol. 37, nr 3

85-92

EN

Changes in phylogenetic groups of methanogenic microorganisms during methane fermentation have been studied. Phylogenetic groups of methanogens were quantified and visualized by hybridization of oligonucleotide probes complementary to rRNA of the major phylogenetic groups. At the beginning of fermentation, Eubacteria are the main group and the count of Archaea constitutes only an insignificant percentage of the population of microorganisms. During the process of fermentation, a very significant increase in methanogenic microorganisms was recorded after 70 days of the process in progress. It is reflected in the quantity and the composition of released biogas.