Acrylamide recrystallization

Acrylic acid (AA) and methacrylic acid (MAA) (purchased from Merck) are freed from inhibitor on a neutral aluminium oxid column and distilled. Acrylamide (AM) from Kebo, Stockholm, is recrystallized once from chloroform solution before use. Other monomers of analytical grade were purchased from Merck and used as received crotonic acid (CA), tiglic acid (TA), 3-methyl crotonic acid (3-MCA), and a-methyl cinnamic acid (oi-MCia) (Table 1). Benzophenone (analytical grade, Kebo) and acetone (spectroscope grade, Merck) were used as supplied. 

Materials. Monomers used in the preparation of the copolymers were as follows acrylamide as a 50% solution in water, stablized with cupric ion, supplied by American Cyanamid Company acrylic acid supplied by BASF and AMPS, 2-acrylamido-2-methylpropanesulfonic acid, (recrystallized grade) obtained from Lubrizol. The sodium salts of acrylic acid and AMPS were prepared by gradual neutralization of the monomers with sodium hydroxide solution, maintaining a temperature of 0 to 5°C, to give a final concentration of 50%. 

Toluene (300 mL) was added to (i-PrO)2TiCl2 (1.40 g. 5.9 mmol), the chiral diol (above equation, 3.12 g, 5.9 mmol) and 4 A molecular sieves (powder, 3.0 g) at 0 C and the mixture stirred for 30 min. The acrylamide 43 (R = Me 10.0 g, 71 mmol) in petroleum ether (200 mL) was added. Subsequently, a petroleum ether soln (100 mL) of ketene dimethyl thioacetal (44 10.0 g, 83 mmol) was added and the mixture stirred at 0 C for 1 h. The reaction was quenched by adding pi I 7 phosphate buffer and inorganic materials removed by filtration. The organic materials were extracted with EtOAc and the extracts washed with brine and dried (Na,S04). After evaporation of the solvent, the crude product was purified by eolumn chromatography yield 14.8 g (80%). Optically pure 45 was obtained by two recrystallizations from benzene/hexane (80% recovery) mp 90.5 92.0 C. 

Materials. Ion-exchanged and distilled water was used in all the polymerizations. Four kinds of acrylamides were used as comonomers (M2). Acrylamide (AA, Wako Chemicals Co.) and methacrylamide (MA, Tokyo Kasei Co.) were recrystallized from benzene. 

N-(Hydroxymethyl)acrylamide (HMA, Tokyo Kasei Co.) was recrystallized from ethyl acetate. N,N-Dimethylacrylamide (DMA, Tokyo Kasei Co.) and styrene (St, Kashima Kagaku Yakuhin Co.) were distilled at 54°C/3.5 mmHg and 40°C/14.5 mmHg, respectively. In some copolymerizations cross-linking reagents were added to reduce the formation of water-soluble polymer. N -Methylenebisacrylamide (MBA, Nakarai Chemicals Co.) and N-allylacrylamide (AAA, Polysciences, Inc.) were used as received. Divinylbenzene (DVB, Tokyo Kasei Co.) was treated with 10 % sodium hydroxide and dried. Two kinds of initiators were used Potassium persulfate (KPS, Taisei Kagaku Co.) was recrystallized from water and azobis(isopropyl-... 

Prepare solution B (acrylamide) by dissolving 22.2 g recrystallized acrylamide and 0.6 g N,N -methylene-bis(acrylamide) in sufficient distilled water to yield a final solution volume of 100 ml. This solution should be stored in a brown bottle at 4°C. Solutions more than a month old yield inferior gels and should therefore be replaced. Acrylamide is a strong, cumulative neurotoxin that is easily absorbed through the lungs or directly through the skin. Therefore, extreme care should be taken not to breathe unpolymerized acrylamide or permit it to come in contact with the skin. 

Recrystallized preparations of acrylamide are available commercially or may be prepared as follows. Dissolve acrylamide (70 g) in 1 liter chloroform at 50°C. Filter the solution hot and cool it to —20°C to bring about crystallization. Collect the crystalline acrylamide in a chilled Buchner funnel, wash with chilled (—20°C) chloroform and/or heptane, and dry. Additional discussion of the purification of reagents used for electrophoresis may be found elsewhere (35,36). 

Materiala. Nonionic surfactants Brij 52 (B52) and Brij 30 (B30) were obtained from the Sigma Chemical Company and used as received. These surfactants are ethoxylated alcohols with the nominal structures Cie a and C12E4, respectively, where E represents the number of ethylene oxide units. Acrylamide was obtained from the Aldrich Chemical Company (Gold Label 99-1-%) and recrystallized twice from chloroform. Azo bis(isobutyrnitrile) (AIBN), obtained from the Alfa Products Division of Morton Thiokol, was recrystallized from methanol. Water was doubly deionized. Propane obtained from Union Carbide Linde Division (CP Grade) and ethane from Air Products (CP Grade) were used without further purification. 

Solid monomers acrylamide and methacrylamide were recrystallized twice from methanol. Bulk monomers were polymerized in the manner described above, but in the solid state (no melting). Aqueous solutions of these monomers were handled as the liquid monomers. 

Materials. Scoured cotton cellulose fibers of the Acala variety supllied by Kanebo Co Ltd. were piirified by extracting with hot benzene-ethanol mixture (l l vol. ratio) for 2kh. Then the cotton fibers were washed with methanol and distilled water and air-dried. Acrylamide (AM) was purified by recrystallization from benzene for several times. Bis-(beta-chloroethyl) vinylphosphonate (Fy) was purified by passing the monomer through a column filled up with activated alumina to remove inhibitors of polymerization. Other chemicals used were reagent grade, and were used without further purification. 

Rehydratable gels were obtained from Micro-Map, Inc., Boca Rotan, FL, at both 3 per cent T, 3.5 per cent C and 5 per cent, T 3.5 per cent C. (T refers to the per cent of monomer present, while C is defined as the per cent of cross-linker relative to T. The per cent of the gel is the total of these two values) For comparative studies conventional gels were prepared at the same acrylamide concentration from the same bottle of two times recrystallized acrylamide (Serva). Acrylamide was recrystallized also from chloroform as previously described [2]. These gels were 250 or 375 1 thick and were bonded to silanized glass plates, or covalently bonded to GelBond Pag 7 mil mylar sheets (Marine Colloids, Rockland ME ) using previously published methods 118-191 and were randomized between the two laboratories. Single lots of ampholytes from various manufacturers were used for the comparative studies in each laboratory to eliminate lot variation. 

The polyacrylamide gels were prepared by a standard redox reaction with ammonium persulfate and tetramethylethylenediamine (TEMED). Recrystallized acrylamide monomer (5 g), N,N -methylenebis(acrylamide) (0.133 g), ammonium persulfate (40 mg), and TEMED (400 xL) were dissolved in water to make a total volume of 100 mL. After thorough mixing, the preparation was poured through a small aperture into cylindrical or spherical glass molds. Upon gelation, the molds were broken and the gel samples were transferred into a cell containing an excess of water. This time was taken to be zero if = 0) for the kinetics experiments. 

Acrylamide (Cyanamid C.V.) was recrystallized once from chloroform (Caledon, reagent grade), washed with benzene (BDH, reagent grade), dried in vacuo, and stored over silica gel in a desiccator. For the kinetic studies, DMAEM was purified... 

Reagents. Acrylamide (Aldrich Chemical Company) was recrystallized from acetone twice before use. Electrophoresis grade sodium dodecyl sulfate (SDS) (Polysciences, Inc.) was used without further purification. Distilled water was purified additionally by treatment with a water purification system (Milli-Q, Millipore) to give water with a resistivity of greater than 18 Mft-cm. 

NaAMB Series. 3-Acrylamido 3-methylbutanoic acid (AMBA) was synthesized via a Ritter reaction involving acrylonitrile and 3,3-dimethylacrylic acid in the presence of water and an excess of sulfuric acid. The synthesis basically followed the procedure set forth by Hoke and Robins. The crude product was twice recrystallized from a mixture of methyl ethyl ketone and petroleum ether prior to use (m.p. 89-91 C). The product was analyzed by elemental analysis and FTIR. Acrylamide was twice recrystallized from acetone prior to use. 

Acrylamide was purified by recrystallization from distilled methanol. 

Highly purified 2-hydroxyethyl methacrylate (HEMA), supplied by Pa. R6hm Haas, Darmstadt, FRG, and recrystallized acrylamide (AAm), supplied by Fa. Stockhausen, Krefeld, FRG, were used as graft monomers. In the case of grafting with AAm, a 10 molar solution of AAm in methanol was used. 

Acrylamide and methacrylamide were used after recrystallization from chloroform and ethyl acetate, respectively. 

Polyetherurethanes "WHS" (Biosearch Inc., Raritan, NJ) and "WALOPUR" (Wolff, Walsrode, FRG) were extracted in an Et0H/H20 mixture prior to use. Liquid monomers 2-hydroxyethylmethacrylate (HEMA), 2-epoxypropyl-methacrylate (GMA), 2-dihydroxypropylmethacrylate (GOMA) and acrylic acid (AAc) were distilled under reduced pressure. Acrylamide (AAm) was recrystallized from methanolic solution. Gases for glow discharge treatment... 

Acrylamide (ultra pure electrophoresis grade form Polysciences, Inc.) was used without further purification. Laurylacrylate (6) (Monomer-Polymer Labs) was purified by washing with 1 M NaOH followed by vacuum distillation. The 1,1-dihydroperfluorobutyl acrylate (2) and 1,1-dihydroperfluorooctyl acrylate (3) were purchased from Monomer-Polymer Laboratories and used as received. The fluorine containing surfactants and monomers, 2-(AT-ethylperfluorooctane-sulfoamido)ethylacrylate (4) and -methacrylate (5) technical grade were donated by the 3M Company. The monomers were recrystallized from MeOH. All of the solvents and salts were of reagent grade or ACS grade quality. The AMPS monomer (sodium salt Aldrich) was used as received. 

Materials. Palladium-graphite (Pd-Gr) was prepared by the reaction of potassium intercalated graphite and palladium chloride (II) as described in the literature.(i6) The content of palladium in Pd-Gr was determined to be 17 wt% by ICP plasma emission spectroscopy. iV . V -(3,4 -Oxydiphcnylcne)bis(acrylamide) JL was prepared by the condensation of 3,4 -oxydianiline and acrylic acid cMoride. The yield was 56 % and the structure was confirmed as described in the literature.(2) Bis(4-iodophenyl) ether 2. was prepared by the reaction of diphenyl ether, iodine, and bis[ istrifluoroacetoxy)iodobenzene in carbon tetrachloride at room temperature. The precipitate was filtered, washed with methanol, and purified by recrystallization from n-hexane. TTie yield was 59 % and the structure was confirmed as described in the literature. [Yoneyama, 1989 8] Ethyl acrylate, iodobenzene, trialkylamines, 1-decene, nitrobenzene, and all solvents used for the reaction were purified by distillation. Other materials were used as received. 

Materials. NIPAAm (Aldrich) was recrystallized from n-hexane, and dimethylacrylamide (DMAAm Aldrich) and diethylacrylamide (DEAAm Polysciences, Inc.) were distilled to remove inhibitor. Methylacrylamide (MAAm ABCR), ethylacrylamide (EAAm ABCR), acrylamide (AAm Life Technologies), ammonium persulfrite (APS, Aldrich), N,N -methylenebisacrylamide (BIS Aldrich),... 

Acrylic acid (AAc), from Eastman Chemical Co., Rochester, New York, was distilled before use. Acrylamide (AAm), from J. T. Baker Chemical Co., Toronto, Ontario, was recrystallized from chloroform before use. N,N -methylene-bisacrylamide (Bis), from Eastman Chemical Co., was used as received. 

A series of copolymers of acrylic acid (AAc) and acrylamide (AAm), cross-linked with Bis, were either prepared or, as in the case of CO(AAc/AAm)V, purchased. Those synthesized in the laboratory were produced from freshly distilled AAc and AAm recrystallized from chloroform. The solution copolymerization and crosslinking, when carried to approximately 5% conversion, yielded an insoluble gel that was subsequently broken in a blender and forced through a 20-mesh screen, washed in 0.5N HCl, washed in distilled water, freeze-dried, and stored in a desiccator under vacuum. By controlling both the pH of the reaction medium and the feed ratio of the two monomers, the series of copolymers listed in Table II was prepared, the analysis being by the titration, with O.IN NaOH, of a suspension of the copolymer in O.IN HCl to determine the AAc content of the copolymer. 

Acrylamide (AM) 99%, and chitosan (CS) 99% from Aldrich Chemical, 2,2 -azo-bis(2-amidinopropane) dihydrochloride (V-50) from Wako Chemicals, and itaconic acid 98.5% from Acros were used. N, N -methylene-bis-acrylamide or NMBA (Scientific Polymer) was recrystallized from a methanol solution. 

---