Refractive index detection polymer analysis

The separation was carried out on a TSKgel Amide-80 column 4.6 mm i.d. and 25 cm long with a mobile phase consisting of a 80% acetonitrile 20% water mixture. The flow rate was 1 ml/min and the column was operated at an elevated temperature of 80°C. The saccharides shown were 1/ rhamnose, 2/ fucose, 3/ xylose, 4/ fructose, 5/ mannose, 6/ glucose, 7/ sucrose and 8/ maltose. The analysis was completed in less than 20 minutes. These types of separations including other biomonomers, dimers and polymers are frequently carried out employing refractive index detection. 

Detection is also frequently a key issue in polymer analysis, so much so that a section below is devoted to detectors. Only two detectors, the ultra-violet-visible spectrophotometer (UV-VIS) and the differential refractive index (DRI), are commonly in use as concentration-sensitive detectors in GPC. Many of the common polymer solvents absorb in the UV, so UV detection is the exception rather than the rule. Refractive index detectors have improved markedly in the last decade, but the limit of detection remains a common problem. Also, it is quite common that one component may have a positive RI response, while a second has a zero or negative response. This can be particularly problematic in co-polymer analysis. Although such problems can often be solved by changing or blending solvents, a third detector, the evaporative light-scattering detector, has found some favor. 

Detection in 2DLC is the same as encountered in one-dimensional HPLC. A variety of detectors are presented in Table 5.2. The choice of detector is dependent on the molecule being detected, the problem being solved, and the separation mode used for the second dimension. If MS detection is utilized, then volatile buffers are typically used in the second-dimension separation. Ultraviolet detection is used for peptides, proteins, and any molecules that contain an appropriate chromophore. Evaporative light scattering detection has become popular for the analysis of polymers and surfactants that do not contain UV chromophores. Refractive index (RI) detection is generally used with size exclusion chromatography for the analysis of polymers. 

The second method, using refractive index (RI) detection, is carried out using a resin-based polymer column. Sucrose elutes first from this column, followed by glucose, fructose and then sorbitol. This type of column is generally more robust than the amino-bonded column and if handled well will last much longer however, it is around three times more expensive. The method has been collaboratively tested for the analysis of sugars and sorbitol in fruit juices by the IFU. The HPLC conditions are given below. 

The equipment widely used for the detection of carbohydrates in the HPLC method is the differential refractive index (RI) detector. The principle involved in this detection depends on the continuous measurement of the variation of the RIs of the mobile phase containing the samples with little or no chromophores such as carbohydrates, lipids, and other polymer compounds that do not absorb UV light. RI detection method presents high degree of reproducibility and is very convenient for the analysis of polysaccharides. However, other detectors such as evaporative light scattering detector and pulsed amperometric detector have been used for the detection of polysaccharides [100]. 

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