Gas Knowledge

Chromatography, also known as chromatography. According to its separation principle, there are methods such as adsorption chromatography, partition chromatography, ion exchange chromatography and exclusion chromatography. Adsorption chromatography is the use of different adsorbent adsorption capacity of the material to be separated, eluted with a solvent or gas to separate the components. Commonly used adsorbents are alumina, silica gel, polyamide and other substances with adsorption activity. Partition chromatography uses the separation coefficient of the separated substances in the two phases to separate the components. One of the phases is a liquid, which is coated or bonded to a solid support and is called a stationary phase; the other phase is a liquid or gas and is called a mobile phase. Commonly used carriers include silica gel, diatomaceous earth, silica-magnesium adsorbent and cellulose powder. Ion exchange chromatography utilizes the ion exchange potential of the separated substance on the ion exchange resin to separate the components. Commonly used cation and anion exchange resins with different strengths, the mobile phase is generally water or a buffer solution containing an organic solvent. Exclusion chromatography, also known as gel chromatography or gel permeation chromatography, utilizes the difference in the molecular weight of the material to be separated and the degree of penetration on the filler to separate the components. Commonly used fillers include molecular sieves, dextran gel, microporous polymers, microporous silica gel or glass beads, etc. Water or organic solvents can be used as the mobile phase according to the nature of the carrier and the sample. The separation methods of chromatography include column chromatography, paper chromatography, thin layer chromatography, gas chromatography, and high performance liquid chromatography. The solvent used for chromatography should not react chemically with the sample, and a solvent with higher purity should be used. The temperature during chromatography refers to operation at room temperature unless otherwise specified by gas chromatography. For the detection of each component after separation, the method specified in each monomer shall be used. Column chromatography, paper chromatography or thin layer chromatography are usually used to separate colored substances according to their color bands. Some colorless substances can be inspected under 245-365nm ultraviolet light. Paper chromatography or thin layer chromatography can also be sprayed with developer to make it develop color. Thin-layer chromatography can also be used to examine the thin-layer silica gel with fluorescent substances, using the fluorescence quenching method. When using paper chromatography for quantitative measurement, you can cut or dig out the chromatographic spots, dissolve the component with a solvent, and then use spectrophotometry or colorimetry to determine it. You can also use a chromatographic scanner to measure directly on paper or thin-layer plates It can also be measured directly on a paper or thin-layer board with a chromatographic scanner. Column chromatography, gas chromatography and high performance liquid chromatography can be detected by various detectors connected to the outlet of the chromatography column. Column chromatography can also collect the effluent separately and measure it by a suitable method. The chromatographic tube used for column chromatography is a hard glass tube with uniform inner diameter and a narrowing at the lower end. The lower end is plugged with cotton or glass fiber, and the tube is filled with an adsorbent. The size of the chromatographic column, the type and amount of adsorbent, and the flow rate during elution are all in accordance with the regulations in each monomer. The particles of the adsorbent should be kept as uniform as possible to ensure a good separation effect. Unless otherwise specified, particles with a diameter of 0.07-0.15mm are usually used. Attention should be paid to the effect of adsorbent activity or adsorption force on the separation effect. Dry method of adsorbent filling: Add the adsorbent to the chromatography tube at one time, vibrate the wall to make it sink uniformly, then slowly add the mobile phase used to start chromatography along the wall, or add a piston to the lower end of the chromatography tube. Add an appropriate amount of mobile phase and unscrew it to slowly drop out the mobile phase, then slowly add the adsorbent from the top of the tube to make it evenly wet and sink, forming a moderately tight adsorption layer in the tube. During the operation, sufficient mobile phase should be kept on the adsorption layer. Wet method: Mix the adsorbent with the mobile phase, stir to remove air bubbles, slowly pour into the chromatography tube, and then add the mobile phase to wash the adsorbent attached to the wall of the tube to make the surface of the chromatography column flat. As soon as the mobile phase used for packing the adsorbent flows down from the chromatographic column naturally, the liquid level will be equal to the column surface, that is, the sample solution is added. Unless otherwise specified, the sample should be dissolved in the mobile phase used for chromatography, and then slowly added along the wall of the chromatography tube. Be careful not to tip the adsorbent. Or dissolve the sample in an appropriate solvent. Mix it with a small amount of adsorbent, and then evaporate the solvent to make it loose; add the adsorbent mixed with the sample to the prepared chromatography column. If the sample does not dissolve in common solvents, the sample and the appropriate amount of adsorbent can be ground and mixed in a mortar and then added. Unless otherwise specified, the elution capacity of the mobile phase is usually changed according to the size and proportion of the mobile phase. The effluent is collected separately and the effluent is collected separately until the content of the effluent is significantly reduced or no longer contained. Variety and proportion of mobile phase. During the operation, sufficient mobile phase should be kept on the adsorption layer. Paper chromatography uses paper as a carrier and uses a single solvent or mixed solvents for partitioning. That is to say, using water or other substances contained in the paper as the stationary phase, the mobile phase is used for the development of distribution chromatography. The filter paper used should be even and flat in texture, have a certain mechanical strength, must not contain impurities that can affect the chromatographic effect, and should not work with the developer used, so as not to affect the separation and identification effects, and can be used after special treatment if necessary. After the sample is chromatographed, the ratio shift value (Rf) can be used to indicate the position of each component (ratio shift value = ratio of the distance from the center of the origin to the center of the chromatographic spot and the distance from the center of the origin to the front of the mobile phase), due to the influence of the ratio shift value There are many factors, so the comparison of control substances under the same experimental conditions is generally used to determine their similarities and differences. When identifying as a monomer, the color (or fluorescence) and supply of the main chromatographic spot displayed by the sample should be mixed with the spectral spot of the main color displayed by the control (standard) sample or the test-control substance (1: 1) The main chromatogram spots displayed are the same. As a quality index (purity) inspection, a certain amount of sample may be taken, and after expansion, the number of chromatographic spots or the intensity of color (or fluorescence) of the impurities displayed by each monomer shall be inspected according to the provisions of each monomer. For the content determination, the chromatographic spots can be cut off and eluted, and then measured by an appropriate method, and can also be measured by a chromatographic scanner. 1. The chromatographic cylinder used in the downward method is generally a round or rectangular glass cylinder. The cylinder has a ground glass cover, which should be able to be sealed. The cover has a hole, which can be inserted into a separatory Funnel to add a mobile phase. At the top of the near-cylinder, there is a glass tank set up by a bracket as a mobile phase container. A glass rod is used in the tank to support the chromatographic filter paper to sag naturally to avoid siphoning of the mobile phase along the filter paper and the solvent tank. Take appropriate chromatographic filter paper and cut it into a paper strip of appropriate size according to the direction of the fiber filament, at an appropriate distance from the upper end of the paper strip (so that the upper end of the chromatographic paper can be sufficiently immersed in the mobile phase in the solvent tank, and the spotting baseline can be in the solvent tank (Several centimeters below the glass support rod on the side) Use a pencil to draw a little baseline. If necessary, the lower end of the chromatographic paper can be cut into a zigzag shape to facilitate the dropping of the mobile phase. Dissolve the sample in an appropriate solvent to make a certain concentration of solvent. Use a micropipette or a microsyringe to draw the solvent and place it on the spotting baseline. The solution should be added point by point. After each point addition, let it dry naturally, dry at low temperature or blow dry with warm air flow. The diameter of the sample point generally does not exceed 0.5 cm, and the sample point should generally be round. Place the upper end of the spotted chromatographic filter paper in the solvent tank and press it with a glass rod so that the chromatographic paper sags naturally through the glass support rod on the side of the tank. The spotting baseline is a few centimeters below the support rod. Before the start of chromatography, the chromatographic cylinder is saturated with the vapor of the solvent specified in each monomer. Generally, a plate containing a mobile phase can be placed at the bottom of the chromatographic cylinder, or a filter paper strip soaked in the mobile phase can be attached to the chromatographic cylinder. Place on the inner wall for a certain period of time, as the solvent evaporates to fill the cylinder with saturated vapor. Then add the mobile phase to immerse the filter paper in the solvent tank. After the mobile phase expands along the filter paper by capillary action to a specified distance, remove the filter paper and mark the leading position of the mobile phase. Once the mobile phase is scattered, the chromatographic spots are detected according to the specified method. 2. The ascending method chromatography cylinder is basically similar to the ascending method except that the solvent tank and bracket are removed, and a plug is inserted into the hole on the cap of the chromatography cylinder. A glass hook is inserted into the plug to hang the spotted chromatography filter paper on the hook. Chromatography filter paper is generally about 25cm long, and the width depends on the needs. If necessary, the chromatography filter paper can be rolled into a cylindrical shape. The spotting baseline is about 2.5cm from the bottom edge, and the spotting method is the same as the descending method. Add an appropriate amount of mobile phase into the chromatographic cylinder and place it. Once the mobile phase vapor is saturated, lower the hook, so that the chromatographic filter paper is immersed in the mobile phase by about 0.5cm. The mobile phase rises along the chromatographic filter paper by capillary action. Unless otherwise specified, it generally expands to After 15cm, take it out to dry and inspect it according to the prescribed method. Chromatography can be carried out in one direction, that is, one-way chromatography; it can also be carried out in two directions, that is, first developed in one direction, taken out, and after the mobile phase has completely evaporated, turn the filter paper 90 °, and then use the original mobile phase or another flow Exhibition. It can also be unfolded multiple times, continuous development or radial chromatography.