SU1153976A1 - Method of obtaining sorbent for separating oligonucleotides - Google Patents
Method of obtaining sorbent for separating oligonucleotides Download PDFInfo
- Publication number
- SU1153976A1 SU1153976A1 SU833670634A SU3670634A SU1153976A1 SU 1153976 A1 SU1153976 A1 SU 1153976A1 SU 833670634 A SU833670634 A SU 833670634A SU 3670634 A SU3670634 A SU 3670634A SU 1153976 A1 SU1153976 A1 SU 1153976A1
- Authority
- SU
- USSR - Soviet Union
- Prior art keywords
- sorbent
- resolution
- acetone
- treatment
- selectivity
- Prior art date
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/30—Processes for preparing, regenerating, or reactivating
- B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
- B01J20/3202—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the carrier, support or substrate used for impregnation or coating
- B01J20/3204—Inorganic carriers, supports or substrates
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
- B01J20/286—Phases chemically bonded to a substrate, e.g. to silica or to polymers
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/30—Processes for preparing, regenerating, or reactivating
- B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
- B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
- B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
- B01J20/3268—Macromolecular compounds
- B01J20/3272—Polymers obtained by reactions otherwise than involving only carbon to carbon unsaturated bonds
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J2220/00—Aspects relating to sorbent materials
- B01J2220/50—Aspects relating to the use of sorbent or filter aid materials
- B01J2220/54—Sorbents specially adapted for analytical or investigative chromatography
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- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Organic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Inorganic Chemistry (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
- Solid-Sorbent Or Filter-Aiding Compositions (AREA)
Abstract
СПОСОБ ПОЛУЧЕНИЯ СОРБЕНТА ДЛЯ РАЗДЕЛЕНИЯ ОЛИГОНУКЛЕОТИДОВ, включаю ций обработку кремнезема подиэтиленимином и раствором эпоксидной смолы или дибромэтана в диоксане, отличающийс тем, что, с целью повьшени селективности и разрешающей способности сорбента дополнительно осуществл ют обработку йодистым метилом в ацетонитриле. (ЛA method of producing a sorbent for separating oligonucleotides, including treating silica with ethylenimine and a solution of epoxy resin or dibromoethane in dioxane, characterized in that, in order to increase the selectivity and resolution of the sorbent, treatment with iodine methyl acetate in addition to acetone and resolution of the sorbent additionally carry out treatment with iodine methyl acetate in acetone and an increase in selectivity and resolution of the sorbent additionally carry out the treatment with iodine methyl acetone in acetone, and the resolution of the sorbent additionally carry out the treatment with iodine methyl acetate in acetone and a resolution of the sorbent additionally carry out treatment with iodine methyl amine in acetone and a resolution of the sorbent with a methyl iodine in acetone in order to increase the selectivity and resolution of the sorbent. (L
Description
елate
со со with so
О) Изобретение относитс к материал дл жидкостной хроматографин и може быть использовано в цинтезе сорбентов дл разделени и анализа олигонуклеотидов . Цель изобретени - повышение селективности и разрешаюо ей способнос ти сорбента. Сущность предложенного способа заключаетс в следующем. Высушенный в вакууме силикагель обрабатывают раствором полиэтиленим на в этиловом спирте, перемешивают, дегазируют,фильтруют, отмывают этиловым спиртом. Полученную композицию сущат и обрабатывают раствором эпоксидной смолы или дибромэтана в диоксане. После следующей промывки сорбента и сушки на него воздейству избытком йодистого метила в ацетони риле, дегазируют и кип т т реакцион ную смесь, промывают ацетонктрнпом окончательно высушивают. Пример 1. Синтез сорбента. 2 г силохрома С-80 с удельной поверх ностью 80 , предварительно высушенного в вакууме при 190°С в течение 3 ч, помещают в круглодонную кол обу на 100 мл и заливают 20 мл 10%ного раствора полизтиленимина (молекул рна масса 35 тыс.) в этиловом спирте. Хорошо перемешивают на встр хивателе 3 мин, дегазируют, оставл ют на 40 мин и повтор ют встр хивание. Затем силикагель на фильтрующей воронке промывают этиловым спиртом (6 X 30 мл) и сушат при 90С в вакууме в течение 2ч, перенос т в круглодонную колбу на tOO мл добавл ют 20 мл 10%-ного раствора . эпоксидной смолы ДЭГ-1 в диоксане, встр хивают 2 мин, дегазируют, остав л ют на 12 ч. После этого носитель промывают последовательно диоксаном (Эх 30 мл), спиртом (3 X 30 мл), ацетонитрилом (3 х 30 мл), спиртом (2 X 30 мл), сушат в вакууме при в течение 2 ч. Перенос т в круг лодонную колбу на 100 мл, добавл ют 50 мл 50%-ного раствора йодистого метила в ацетонитриле, перемешивают на встр хивателе 3 мин, дегазируют, кип т т 2 ч и оставл ют на 12 ч при комнатной температуре. Промывают сорбент ацетонитрилом (3 х 30 мл), этиловым спиртом (3 X 30 мл), сушат в течение 2 6 Содержание азота в полученном сорбенте по элементному анализу составл ет 1,4%, что соответствует емкости 1,0 мг-экв/1. Пример 2. Синтез сорбента. Синтез провод т аналогично примеру 1, но вместо раствора эпоксидной смолы берут 10%-ный раствор 1,2-дибромэтана в диоксане. После обработки йодистым метилом и сушки, сорбент заливают 50 мл смеси диметилбензиламина и ацетокитрила (1:1), встр хивают 2-3 мин, дегазируют и оставл ют на 10 ч. После этого носитель промывают на воронке с фильтрующим дном ацетонитрилом (3 х 30мл), этиловым спиртом (3 X 30 мл), водой (3 X 30 мл), сушат при ЮОС 2 ч. Содержание азота в полученном сорбенте по элементному анализу составл ет 1,4%, что соответствует 1,0 мг-экв/г. Полученный сорбент испытывали на жидкостном хроматографе высокого давлени модели 332 фирмы Альтекс (США). В качестве элюита использовали 0,1 М KHjPO, рН 6,5; 30% MeCN, скорость потока 0,5 мл/мин. Использовали сорбенты с размером частиц 10 мкм, колонки 3,2 х 250 мм. Селективность сорбента определ ют по известной формуле tx 3 to , коэффициент селективности; олигодезоксинуклеотид; d (CAAGCCTCCA); d (CTCTTACCCAA); d (AAAAACGCTTACAA); d (GAAAGGTGAATAA); врем удерживани декадезоксинуклеотида; врем удерживани х-го олигодезоксинуклеотида; врем выхода неудерживаемого на колонке вещества. Коэффициенты селективности предлагаемого , известного и фирменного сорбента Partisil SAX приведены в табл.1, Partisil SAX - сорбент фирмы Whatman (США), представл ющий собой сильный анионообменник, наиболее часто используемый дл выделени олигонуклеотидов . Коэффициент разрешающей способное ти сорбента вычисл ют по формуле R V. 2 , 1 .W W где R,j коэффициент разрешающей способности; Ц - врем удерживани у-го олигодезоксннуклеотида; t - врем удерживани декадезок синуклеотида;ными, w - ширина пика декадезоксинук- ни и леотида;дов. видно, сто сама высока степень разделени олигонуклеотидов достигаетс на предложенном сорбенте, что делает возможным полное разделение всех четырех олигонуклеотидов в отличие от известных сорбентов. Таким образом, предложенный способ получени сорбента позвол ет повысить селективность и раэт ешающую способность сорбента по сравнение с известчто повышает качество разделеанализа смесей олигонуклеотиO) The invention relates to a material for liquid chromatographic and can be used in the sorption of sorbents for the separation and analysis of oligonucleotides. The purpose of the invention is to increase the selectivity and resolution of the sorbent. The essence of the proposed method is as follows. Dried under vacuum, the silica gel is treated with a solution of polyethylene on ethanol, stirred, degassed, filtered, washed with ethyl alcohol. The resulting composition is present and treated with a solution of epoxy resin or dibromoethane in dioxane. After the next washing of the sorbent and drying it by the action of an excess of methyl iodide in acetone, it is degassed and the reaction mixture is boiled, washed with acetone and finally dried. Example 1. Synthesis of sorbent. 2 g of silochrome C-80 with a specific surface 80, previously dried in vacuum at 190 ° C for 3 hours, are placed in a 100 ml round bottom flask and poured 20 ml of a 10% solution of poly (stylenimine) (molecular weight 35 thousand) in ethyl alcohol. Mix well on a shaker for 3 minutes, degas, leave for 40 minutes, and shake again. Then the silica gel on a filtering funnel was washed with ethyl alcohol (6 X 30 ml) and dried at 90 ° C under vacuum for 2 hours, transferred to a too ml round bottom flask with 20 ml of a 10% solution added. DEG-1 epoxy resin in dioxane, shaken for 2 minutes, degassed, left to stand for 12 hours. After this, the carrier was washed successively with dioxane (Ex 30 ml), alcohol (3 X 30 ml), acetonitrile (3 x 30 ml), alcohol (2 X 30 ml), dried under vacuum for 2 hours. Transfer the flask to 100 ml in a circle, add 50 ml of 50% methyl iodide in acetonitrile, mix on a stirrer for 3 minutes, degas, boil for 2 hours and leave for 12 hours at room temperature. The sorbent is washed with acetonitrile (3 x 30 ml), ethyl alcohol (3 X 30 ml), dried for 2 6 The nitrogen content in the resulting sorbent according to elemental analysis is 1.4%, which corresponds to a capacity of 1.0 mEq / 1 . Example 2. Synthesis of sorbent. The synthesis is carried out analogously to example 1, but instead of a solution of epoxy resin, a 10% solution of 1,2-dibromoethane in dioxane is taken. After treatment with methyl iodide and drying, the sorbent is poured over 50 ml of a mixture of dimethylbenzylamine and acetoxytrile (1: 1), shaken for 2-3 minutes, degassed and left for 10 hours. After this, the carrier is washed on a filter bottom with acetonitrile (3 x 30 ml ), ethyl alcohol (3 X 30 ml), water (3 X 30 ml), dried at 10 ° C for 2 hours. The nitrogen content in the resulting sorbent according to elemental analysis is 1.4%, which corresponds to 1.0 mg eq / g . The resulting sorbent was tested on a high pressure liquid chromatograph model 332 from Altex (United States). 0.1 M KHjPO, pH 6.5; 30% MeCN, flow rate 0.5 ml / min. Sorbents with a particle size of 10 μm, columns 3.2 x 250 mm were used. The selectivity of the sorbent is determined by the known formula tx 3 to, the coefficient of selectivity; oligodeoxynucleotide; d (CAAGCCTCCA); d (CTCTTACCCAA); d (AAAAACGCTTACAA); d (GAAAGGTGAATAA); decadeoxynucleotide retention time; retention time of x-th oligodeoxynucleotide; the release time of the unsupported substance on the column. The selectivity coefficients of the proposed, well-known and proprietary sorbent Partisil SAX are given in Table 1, Partisil SAX is a sorbent from Whatman (United States), which is a strong anion exchanger most often used for the isolation of oligonucleotides. The resolution coefficient of the sorbent is calculated by the formula R V. 2, 1 .W W where R, j is the resolution coefficient; C is the retention time of the y-th oligodeoxane nucleotide; t is the retention time of the decucleotide synucleotide; n is the width of the decadetoxynuclei and leotide peak; it can be seen that the highest degree of separation of oligonucleotides is achieved on the proposed sorbent, which makes possible the complete separation of all four oligonucleotides, in contrast to the known sorbents. Thus, the proposed method for the preparation of a sorbent improves the selectivity and rat ability of a sorbent compared to lime, which improves the quality of the analysis section of oligonucleotide mixtures.
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Application Number | Priority Date | Filing Date | Title |
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SU833670634A SU1153976A1 (en) | 1983-12-08 | 1983-12-08 | Method of obtaining sorbent for separating oligonucleotides |
Applications Claiming Priority (1)
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SU833670634A SU1153976A1 (en) | 1983-12-08 | 1983-12-08 | Method of obtaining sorbent for separating oligonucleotides |
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SU1153976A1 true SU1153976A1 (en) | 1985-05-07 |
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SU833670634A SU1153976A1 (en) | 1983-12-08 | 1983-12-08 | Method of obtaining sorbent for separating oligonucleotides |
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SU (1) | SU1153976A1 (en) |
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1983
- 1983-12-08 SU SU833670634A patent/SU1153976A1/en active
Non-Patent Citations (1)
Title |
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G.B. Сох, C.R. Loscombe, M.J. Slucntt, К. Sugden, I.А. Upxield. The preparation, properties and some applications of bonded ionexchange packings based on microparticulate silica gel for HPLC. J. Chromofogr, 1976, v. 117, p. 269276. Офицеров В.И., Ямщиков В.Ф. Хроматографи олигонуклеотидов на пористых носител х, модифицированных полиэтиленимином. - Биоорганическа хими , 1983, т. 9, 9, с. 12481253. * |
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