• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    A granular crosslinked copolymer suitable as a high speed liquid chromatographic packing and especially as a gel permeation chromatographic packing is disclosed. In a preferred embodiment of the invention the packing is a copolymer essentially consisting of (I) units of at least one vinyl alcohol, (II) units of at least one vinyl ester of a carboxylic acid and (III) units of at least one crosslinking monomer having an isocyanurate ring, the ratio of the units (I) to the units (II) in the copolymer being within the range satisfying the following equation: about about 0.8 wherein a and b are molar ratios of the units (I) and (II), respectively, in the total units (I), (II) and (III) of the copolymer, and a process for preparing the same by suspension polymerization is disclosed.
    公开号:SU1311631A3
    申请号:SU813303096
    申请日:1981-06-24
    公开日:1987-05-15
    发明作者:Янагихара Юзо；Ногути Кохдзи；Хонда Макото
    申请人:Асахи Касеи Когио Кабусики Кайся (Фирма)；
    IPC主号:
    专利说明:

    This invention relates to methods for chromatographic analysis, in particular to a liquid chromatography (LC) nozzle, and can be used in the field of biochemistry and medicine for high-speed and high-performance separation or analysis of substances dissolved in a solvent, in accordance with the mechanism of gel permeation chromatography (GPC). .
    The purpose of the invention is to ensure the suitability of the nozzle for high-speed separation and analysis of substances.
    The proposed gel is obtained by saponification of crosslinked copolymers of vinyl acetate (BA) or vinyl propionate (VP) with tri-allyl isocyanurate (TASH). Attitude
    but
    -g - Y-characterizes the degree of omya + D
    copolymer laziness (i.e., the number of pedroxyl groups) and gel hydrophilicity.
    The coefficients s and b are calculated from the density of hydroxyl groups (qOH) in the gel and the number of TASHCH units, qOH denotes the equivalent of BC units per weight unit of the gel and can be determined by reacting the gel with acetic anhydride in a pyridine solvent, while measuring the amount of acetic acid consumed anhydride in reaction with hydroxyl groups or a change in gel weight, and the concentration of hydroxyl groups is calculated from this measured value. For example, 1 mmol of acetic anhydride is consumed in the reaction with 1 g of dry gel, the qOH of this gel is 1 milliequivalent / g.
    The number of TABLE is determined by the nitrogen content obtained by elemental analysis of the gel. In other words, q can be obtained by qOH, and b - by the value obtained by subtracting the total number of BC and TAIC units from the total number of units in the copolymer, respectively.
    3s
    but
    It affects the degree of crosslinking of the copolymer. The coefficients q and b are determined by the specified method, and c by the values of the elementary analysis of the gel or by calculating the moles VA and TASHCH, which are supplied together (a + b) i, in the indicated formula.
    Attitude
    X character
    five
    0
    five
    When X is in the specified range, the gel with small particles has sufficient strength to withstand high pressure and high speed. Such a gel also possesses sufficient hydrophilicity and poorly adsorbs components in an aqueous solution, in particular proteins and amino acids, which increases the expediency of its use in high-speed GPC.
    The quantitative composition of the nozzle (100 parts by weight of copolymer and 100-198 parts by weight of iodine) characterizes the degree of swelling of the gel - Wg, which is respectively in the range of 1.0-1.98 g water / g of dry gel.
    . Wn is defined as follows.
    The gel, immersed in water and fully balanced, is centrifuged to remove water adhering to the surface of the gel, and its weight (W) is measured. The gel is dried and its dry weight is determined. Wg is calculated by the formula
    Wi - W2
    (wp.
    W,
    w.
    five
    0
    five
    0
    five
    The proposed gel is characterized by a maximum pore size in gel particles corresponding to the molecular weight of polyethylene glycol or dextran, equal to 1.9-10-1.0-10, at which the molecules of these polymers do not penetrate into the pores of the gel particles.
    Substances with a molecular weight below the critical value can be separated by GPC, and substances with a higher molecular weight than the critical value cannot penetrate into the pores of the gel, and pass directly through the gaps in the gel. Consequently, these substances give the same amount of elution, despite the molecular weight, and therefore cannot be separated by GPC.
    Mj, can be determined from the GPC calibration curve, which can be obtained by plotting the logarithm of molecular weights of individual samples, the molecular weight of which is added up on the ordinate, and the elution volume of each sample is on the abscissa relative to the gel column.
    The graph in figure 1 shows the relationship between the elution volume
    3. 1311631
    (V) in milliliters and the molecular weight of the material to be separated. The sloping line and the line parallel to the ordinate are, in this graph, essentially straight lines. The part where the two lines intersect is called a curve. Mr. expressed as a quantity
    t1 rn
    on the ordinate at the point at which there are small pores uniformly distributed in the internal parts of the particles. Organic synthetic polymers having a crosslinked structure swell in a solvent having an affinity for these polymers, and shrink during drying. When using a soft gel, the pores filled with the solvent upon swelling are maintained
    The oblique straight line is relined only with a stitched mesh. No such stack
    cuts the continuation of the line parallel to the ordinate. M. represents one property that is inherent in a gel, and shows an exceptional molecular weight in which the gel can have a separating effect based on the difference in the size of the molecules. Substances having a greater molecular weight than the exclusive molecular weight are eluted essentially together without separation.
    Mp .. is determined by using
    as a standard of substances having a known molecular weight, polyethylene glycols or dextrans, and distilled water as; solvent. Since the water-soluble standard polymers supplied by industry have a molecular weight less than about 2,000,000, a full calibration curve cannot be obtained for a gel having a Md, exceeding 2,000,000. Therefore, Mg, such a gel is not precisely defined, but is evaluated
    15
    Vani in
    20
    25
    thirty
    maintained in the swollen state and, therefore, the gel shrinks most of the pores disappear. In this case, the specific surface area becomes essentially the size of the outer surface of the particles and generally has a very low value - less than 1. When using a solid gel with a solid structure after drying the gel, the pores can maintain essentially the same condition as at the moment of swelling, although they shrink slightly (i.e., the pores have a permanent character. Followed 5, the solid gel has a specific surface area that is much larger than a soft gel.
    Typically, the proposed gel has a specific surface area (S) of about 5-1000. If the gel has a specific surface area less than the lower limit of the range, this means that the gel has a uniform structure (soft gel), which
    at the intersection point at which the pores contain fine pores and, therefore, the repetition of the calibration curve is definitely not suitable for high-speed GPC.
    divided by molecular weights less than 2,000,000, crosses the continuation of a line parallel to the ordinate, which is determined under the same conditions on a gel having a smaller Mp, -.
    In order for the gel to have mechanical strength suitable for high-speed GPC, combined with a lack of adsorption capacity, the degree of crosslinking X should preferably be in the range of about 0.25-0.32 when M d is in the range of about 10 - 10, and the degree of crosslinking should preferably be about 0.27-0.35, when Mp, is in the range of about 10-10.
    The proposed gel is a completely porous solid gel that has a large specific surface area in a dry state. Completely porous structure means structure.
    having small pores evenly distributed in the inner parts of the particles. Organic synthetic polymers having a crosslinked structure swell in a solvent having an affinity for these polymers, and shrink during drying. When using a soft gel, the pores filled with the solvent upon swelling are maintained
    five
    0
    five
    0
    maintained in the swollen state and, therefore, the gel shrinks and most of the pores disappear. In this case, the specific surface area becomes essentially the size of the outer surface of the particles and generally has a very low value - less than 1. When using a solid gel with a solid structure after drying the gel, the pores can maintain essentially the same condition as at the moment of swelling, although they shrink slightly (i.e., the pores have a permanent character. Followed 5 The gel has a specific surface area that is much larger than a soft gel.
    Typically, the proposed gel, specific surface area (S) is about 5-1000. If the gel has a specific surface area less than the lower limit of the range, this means that the gel has the structure of a single; - native type (soft gel), which
    The specific surface area is determined by the BET method using nitrogen gas.
    Our gel typically has a weight average diameter of part (D) of about 4-198 microns. In particular, the most favorable characteristics for LC gel is when D, has a small value, for example, in the range of about 5-20 microns. When high separation capacity is required, the B gel should be in the range of about 5-12 microns. D is measured with a Coulter counter and is calculated using the formula.
    55
    DW
    .5 (; df) IKnj-dt)
    where d, - is the particle diameter; n- - particle frequency d, -.
    with diameter
    5131
    In liquid chromatography, the separation capacity can be increased by reducing the size of the packing particles. However, by passing the solvent through a column packed with a gel with smaller particles, the pressure loss increases significantly compared with a gel that has large particle sizes. Therefore, with a small mechanical strength, the gel is deformed or destroyed, causing abnormally large pressure losses, which makes it impossible for SJH to use a gel with small particle sizes. Since the mechanical strength of the proposed gel can be significantly improved by controlling various properties, including the number and the degree of crosslinking, the gel can hold high speeds and pressures despite the small particle size.
    The pore size of the proposed gel can be adjusted over a wide range, so the gel can be used not only to separate or analyze water-soluble polymers of saccharides or proteins, but also to analyze components with a molecular weight of several hundred thousand and millions that are present in the blood. and urine, which are closely interlinked with disease
    1 2 3 4 5 b 7 8 9 10
    five
    0
    five
    0
    ni renal or liver or symptoms, such as carcinomas. Since the proposed gel has the same high characteristics as the gel for high-speed GPC, these analyzes can be performed in a short period of time with obtaining more information.
    The pre-gel is used as a column nozzle. The column used is usually a cylinder made of stainless steel, but depending on the purpose any column can be chosen.
    Examples 1-19. Gels of a crosslinked copolymer of BC with BA and TAIC (A) or BC with VP and TAIC (B) are placed as a packing in a stainless steel column with a diameter of 7.5 mm and a length of 50 cm.
    The characteristic values of the gels used (X, Y, Dy, W, S., and Mp.) Are determined by the described procedures. The determination of Мр - is carried out on an aqueous solution of dextrans and polyethylene glycols using the detector Septek Reflector 111, model S E-11, pump and injector model 635A.
    The indicators of the used gels are tabulated.,
    Z-Yu
    1.9-10
    3
    8-10
    2.JO
    5-10
    5-10
    2-10
    1 -10
    1-10
    2-W
    Mp determined by polyethylene glycol, dextran.
    V-globulin, bovine serum alyumin, ovalumin and myoglobulin were analyzed on a packed column according to Example 1 using sodium chloride and 0.1 M phosphate, sodium, and an ultraviolet absorption spectrum detector as a quality solution. The proteins eluted in order, with a high molecular weight protein yield of essentially 100%. All samples were measured at a flow rate of 1 mp / min.
    In addition, the solution of a sample of sublimated human short is analyzed, the resulting chromatogram is shown in Fig. 2, where peak A mainly shows alumbo peak B — creatinine and peak C — uric acid. Some components were eluted in larger quantities compared to the free volume of the column due to the weak absorption capacity of the gel, but a number of components were detected and isolated after elution of the -j globulin and aluminumine.
    On a column with a nozzle according to example 2 was performed analysis of the sample
    solvent water containing 0.3 M chloro
    Table continuation
    in other examples - by

    0
    five
    0
    five
    sublimated human syvno- i rotkn. The resulting chromatogram is shown in FIG. 3, where peak D shows mainly alubumin, peak E — craaginine, and peak F — uric acid. The number of peaks is smaller compared to figure 2 due to the low sensitivity of the measurements. The basic pattern of the elution curve is the same as in FIG. Therefore, using the gel of this example, a number of components can be detected and separated.
    On a packed column according to Example 5, standard samples of aluminum bovine serum, ovalumin and myoglobulin were analyzed. The resulting chromatogram is shown in Fig. 4, where curve G shows the elution curve of aluminum, curve I shows the elution curve of ovalumin, and curve I shows the elution curve of myoglobulin.
    The results show that the gel of this example is very effective for the separation of proteins. All samples were measured at a flow rate of 1 ml / min. Each analysis was carried out for 20 minutes.
    9131
    On a column with nozzles according to Examples 3-4, 6-14, proteins were also analyzed, including j-globulin, egg albumin, ovalumin, and myoglobulin. The analysis was carried out at a flow rate of 0.5–2 MPa (min for 10–20 min. The chromatograms obtained show the elution of proteins in the order of increase — molecular weights, the degree of protein extraction 90–100%.
    On a column with gels according to comparative examples 15-19, the analysis of α-globulin and bovine short-ovine albumin, ovalumin and myoglobulin was performed. However, due to the insufficient gel strength and large pressure losses in the column and almost complete absorption of proteins on the gel, elution peaks to the chromatogram were not detected.
    As follows from the examples, the proposed nozzle provides high-speed separation and analysis of substances by gel permeation liquid chromatography.
    权利要求:
    Claims (1)
    [1]
    Invention Formula
    Nozzle for aqueous chromatography based on an aqueous gel of a crosslinked hydroxyl-containing copolymer, characterized in that, in order to ensure the suitability of the nozzle for high-speed separation
    five
    0
    five
    ten
    and analyzing substances, it contains, as a cross-linked hydroxyl-containing copolymer, a copolymer of vinyl alcohol with vinyl acetate or vinyl propionate and trilyl isodiodianurate at a molar ratio of units a / a + b 0.4-0.8 and 3c / a + b + 3c 0 , 25-0.4, where a, b and c are the molar ratios of the vinyl alcohol, vinyl acetate and vinyl propionate and trilyl isocyanurate units to the total number of units in the copolymer, with an average diameter of the gel particles of 4-198 µm and the maximum pore diameter in them, molecular weight polyethylene glycol or de Kstrana, equal to 1.9-10 - 1.0-10, in which the molecules of these polymers do not penetrate into the pores of the gel particles, in the following ratio of components, May, hours: To sew a copolymer Voda 100-198 LLP
    Priority by feature .:
    25.06.80 — using a cross-linked copolymer for adjusting the aqueous gel with a particle size, pore diameter, and water content determined by the formula;
    26, 122, a crosslinked copolymer with a molar ratio of comonomer units defined by the formula for the aqueous gel nozzle;
    Mnt
    1dM
    gjuff.l
    V
    Yu
    but
    30 min
    L.
    g
     7
    t- 8
    Editor M.Petrova
    Compiled by B. Filimonov
    Tehred, L.Oleinik Proofreader A.Ilin
    Order, 1907/58 Circulation 777 Subscription
    VNISh State Committee of the USSR
    for inventions and discoveries 113035, Moscow, Zh-35, Raushsk nab., 4/5
    Production and printing company, Uzhgorod, Projecto st., 4
    72
    10 // fia
    73 L
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    同族专利:
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    EP0043074A3|1982-01-13|
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    EP0043074B1|1984-09-26|
    EP0043074A2|1982-01-06|
    US4368275A|1983-01-11|
    DD159908A5|1983-04-13|
    CA1157996A|1983-11-29|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    LT3061B|1992-07-16|1994-10-25|Fermentas Biotech Inst|Process for the preparation of polymeric gels for sorbents manufacturing|NL6702806A|1966-03-31|1967-10-02|
    JPS5519243B2|1975-11-27|1980-05-24|
    JPS5731925B2|1976-03-09|1982-07-07|
    US4314032A|1978-10-26|1982-02-02|Kureha Kagaku Kogyo Kabushiki Kaisha|Crosslinked polyvinyl alcohol gel|JPH0140953B2|1981-02-12|1989-09-01|Asahi Chemical Ind|
    JPH0144725B2|1981-05-18|1989-09-29|Asahi Chemical Ind|
    JPS5861463A|1981-10-07|1983-04-12|Kureha Chem Ind Co Ltd|Carrier for liquid chromatography and separating and refining method for fat soluble material using said carrier|
    CA1204720A|1982-09-30|1986-05-20|Hajimu Kitahara|Packing materials for chromatographic use and amethod for analysis of an enantiomer mixture usingthe same|
    US4543363A|1983-06-15|1985-09-24|Asahi Kasei Kogyo Kabushiki Kaisha|Ion exchanger having hydroxyl groups bonded directly to backbone skeleton|
    US4904519A|1986-05-12|1990-02-27|Minnesota Mining And Manufacturing Company|Ink-receptive sheet|
    EP0264997B1|1986-10-24|1992-04-08|Asahi Kasei Kogyo Kabushiki Kaisha|A hydrophobic crosslinked copolymer and a method for producing the same|
    US6573327B1|1997-04-01|2003-06-03|Nippon Gohsei Kagaku Kogyo Kabushiki Kaisha|Highly stable aqueous solution of partially saponified vinyl ester resin|
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    US6881761B2|2000-05-29|2005-04-19|Showa Denko K.K.|Porous polymer particle, anion exchanger, producing method thereof, column for ion chromatography, and method for measuring anions|
    JP4717253B2|2000-12-19|2011-07-06|昭和電工株式会社|Porous polymer particles, alkali-resistant anion exchanger, production method thereof, ion chromatography column, and anion measurement method|
    JP4268347B2|2001-05-30|2009-05-27|富士フイルム株式会社|Control serum for dry analytical elements|
    EP1275436A1|2001-07-09|2003-01-15|Showa Denko Kabushiki Kaisha|Packing material and cartridge for solid phase extraction|
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    WO2009020649A1|2007-08-08|2009-02-12|Jordi Flp|Suspension homopolymerization of an isocyanurate|
    GB201405624D0|2014-03-28|2014-05-14|Synthomer Uk Ltd|Method of making a branched polymer, a branched polymer and uses of such a polymer|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    JP55085243A|JPH0115822B2|1980-06-25|1980-06-25|
    JP55183703A|JPH0219902B2|1980-12-26|1980-12-26|
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