The following aspects should be noted when using the UV method:
1 Calibration and verification of the instrument:
1.1 Wavelength Due to the influence of temperature changes on the mechanical part, the wavelength of the instrument often changes slightly. Therefore, in addition to the full calibration calibration of the instrument used, the measurement wavelength should be corrected before the measurement.
1.2 Absorption The accuracy of the absorbance can be determined by a solution of potassium dichromate in sulfuric acid.
1.3 Slit width Select the slit width of the instrument, and the absorbance of the test sample should be no longer increased when the width of the slit is reduced. For most varieties of the Chinese Pharmacopoeia UV method, 2nm seam can be used. Wide, but for some varieties such as penicillin potassium and sodium absorbance inspection requires 1nm slit width or narrower, otherwise its absorbance at 264nm will be low.
1.4 The capacity instrument and analytical balance used for the verification of the balance and glass instruments shall be verified. If there is a deviation, the correction value shall be added.
1.5 The absorption cell paired with the quartz absorption cell also absorbs ultraviolet light. The absorption cell of 1 em is in the range of wavelength 220-270 nm, with air as 100%, its light transmittance is often less than 100%, and each absorption The pools may not be identical, so an absorption pool pairing test should be performed before each measurement. The method is to take a dry and clean absorption tank, and put into the blank solvent for measurement, use one absorption tank as a blank, measure the absorbance of other absorption pools by using the wavelength used in the measurement, and finally select the one with the least absorption as the pairing, and determine And record the absorbance of other absorption tanks, the actual absorbance of the test solution should be the difference between the absorbance of the absorption tank (when there is a blank solvent). In order to reduce the error, a pair of cups is often used to measure several samples (which also reduces computational trouble), but care should be taken to adequately wash the liquid.
2 Solvent requirements: Due to the formation of hydrogen bonds, dipolarization, etc. between the solvent and the solute molecules, the solute absorption wavelength can be shifted. Generally, polar solvents have a greater influence than non-polar solvents, and care should be taken when selecting solvents. Before measuring the test sample, check whether the solvent used in the test sample has an absorption peak near the wavelength used for the test sample, and whether it affects the determination of the test sample. For each measurement, the same batch of solvent should be prepared by using the same manufacturer and the same batch of reagents.
3 Inspection: 3.1 Solution preparation Weighing should be carried out in accordance with the provisions of the Pharmacopoeia. When the preparation solution is prepared, the number of dilution transfer should be as small as possible, and the volume taken during transfer dilution should generally be not less than 5 mL. The content of the test sample should be weighed 2 parts. If it is the reference comparison method, the reference substance should also be weighed 2 parts. The absorption coefficient check should also weigh 2 parts of the test sample and operate in parallel. The deviation of the average value for each result should be within 4-0.5%. One sample may be taken for identification or inspection.
3.2 Determination Unless otherwise specified, the absorbance of several points shall be tested within the specified absorption peak wavelength of 4-2 nm to check whether the absorption peak wavelength position of the test sample is correct, unless otherwise specified. The maximum wavelength of the absorption peak should be within 4-1 nm of the wavelength specified by the variety. Otherwise, the authenticity, purity and accuracy of the wavelength of the sample should be considered. When taking the absorption cell, take the fingers on both sides of the frosted glass surface. The amount of the sample solution is preferably 4/5 of the volume of the pool. When using the volatile solution, it should be covered. The transparent surface should be wiped clean from top to bottom with a mirror paper. The solvent should be free from residual solvents. The light-transmissive surface remaining in the pool can be wiped with a lens paper with a blank solvent and then wiped with a dry tissue paper. When the absorption tank is placed in the sample chamber, it should be noted that the direction of each insertion is the same. Rinse with detergent and water after use, dry and keep dust-proof. If the absorption pool is not easily washed, it can be slightly soaked with fuming sulfuric acid + nitric acid (3:1) mixture, and then washed for use. If cleaning with potassium chromate cleaning solution, the absorption tank should not be soaked in the cleaning solution for a long time. Otherwise, the potassium chromate crystal in the cleaning solution will damage the optical surface of the absorption cell and should be washed thoroughly with water to prevent potassium chromate adsorption. On the surface of the absorption pool.
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