Detector refractive index detector

Model RR/066 351 and 352 pumps models 750/16 variable-wavelength UV monitor detector 750/11 variable filter UV detector, MPD 880S multiwave plasma detector, 750/14 mass detector 750/350/06 electrochemical detector refractive index detector HPLC columns column heaters, autosamplers, pre-columns derivatization systems, solvent degassers, preparative HPLC systems... 

A common cause of baseline drift is a slow elution of substances previously adsorbed on the column. A column cleanup procedure may be in order, or it may need to be replaced. This problem may also be caused by temperature effects in the detector. Refractive index detectors are especially vulnerable to this. In addition, a contaminated detector can cause drift. The solution here may be to disassemble and clean the detector. 

Figure 4.22 High temperature size-exclusion liquid chromatography of an engineering plastic, poly (phenyl sulfate). Column, SSC GPS-3506, 50 cm x 8 mm i.d. eluent, 1-chloronaphthalene flow rate, 1.0 ml min-1 column temperature, 210 °C detector, refractive index detector.

Figure 8. Gel filtration of ethylated (/ -0-4)-(/ -/ )-DHP 16. Solid line Ethylated (/ -0-4)-(/ -/ )-DHP 16 after removal of low molecular weight fractions. The column was calibrated with (/ -0-4)-(/ -/ ) lignin substructure model trimer 6 (molecular weight 642) /3-0-4 lignin model dimer 1 (molecular weight 348) and polystyrenes of molecular weight 9000, 4000 (void), 2200 (indicated by A). Column Sephadex LH-20, 1.1 x 48 cm. Eluent DMF, 13.5-14.4 ml/hr. Detector Refractive index detector RI-2 (Japan Analytical Industry Co., Ltd.).

The UV detector, fluorescence detector, electrolytic detector, refractive index detector, and other detection systems must be checked for repeatability of response signal, sensitivity, and stability. 

Sterols are generally monitored with either a refractive index detector or a UV detector. Refractive index detectors are generally rather insensitive and therefore UV detectors have generally been used where limits of detection are critical. The limits of detection at 254 nm for lanosterol, jS-stigmasterol and cholesterol have been reported to be approximately 600 ng and for ergosterol 40 ng however, at 282 nm the limits of detection for the latter are reduced to 700 pg, putting the range of detection in the same order as that reported for GLC with flame ionisation detection (Colin et al., 1979). 

This type of measurements can very elegantly be realized online by coupling several detectors at the end of the SEC column such as a concentration detector (refractive index detector, spectrophotometric detector, etc.) and an absolute detector measuring the molar mass or related property of the separated species such as laser light scattering detector or capillary viscometer detector. These modern sophisticated separation systems allow not only the separation of the analyzed species but also their very detailed analysis and characterization as concerns the MMD or PSD, as well as other structural and compositional characteristics of simple polymers, co-polymers, etc. A schematic representation of a procedure of SEC data treatment from an experimental chromatogram to the final MMD or PSD data is shown in Figure 8. 

Additional detectors available for HPLC analysis include fluorescence detectors, high-sensitivity diode-array detectors, refractive index detectors, and electrochemical detectors. 

Ultraviolet spectrophometric detectors, refractive index detectors and conductivity monitors are used to monitor liquid-phase compositions. 

Hplc techniques are used to routinely separate and quantify less volatile compounds. The hplc columns used to affect this separation are selected based on the constituents of interest. They are typically reverse phase or anion exchange in nature. The constituents routinely assayed in this type of analysis are those high in molecular weight or low in volatility. Specific compounds of interest include wood sugars, vanillin, and tannin complexes. The most common types of hplc detectors employed in the analysis of distilled spirits are the refractive index detector and the ultraviolet detector. Additionally, the recent introduction of the photodiode array detector is making a significant impact in the analysis of distilled spirits. 

Chromatographic conditions elution with 50 50 methanol/water solvent at the rate of 1.5 ml,/min through a DuPont Zorbax ODS column using a Waters R-401 Refractive Index Detector. 

Select the detector. To acquire molecular weight distribution data, use a general detector such as a refractive index detector. To acquire structural or compositional information, employ a more selective detector such as an ultraviolet (UV) or infrared (IR) detector. Viscometric and light-scattering detectors facilitate more accurate molecular weight measurement when appropriate standards are not available. 

Degassed and preswelled Bio-Gel P-6 and Sephacryl S-200 were packed in self-made glass columns (70 X 1.5 cm 140 X 1.5 cm) and equilibrated for 20 hr with H20(dest.) -t- 0.002% NaN3 to prevent microbial growth. The mass of eluted fractions was detected with a differential refractive index detector (Waters 403 RI, sensitivity 8). 

FIGURE 16.23 Inulln isolated from small (—). medium ( ) and large (A) tubers separated on P-6 (140 X 1.5 cm) flow rate 0.33 ml/min eluenf. H20(dest) + 0.002% NaNa mass detection Waters 403 R differential refractive index detector, sensitivity 8X applied sample solution volume I ml of a 20-mg/ml aqueous inulin solution. 

A more difficult criterion to meet with flow markers is that the polymer samples not contain interferents that coelute with or very near the flow marker and either affect its retention time or the ability of the analyst to reproducibly identify the retention time of the peak. Water is a ubiquitous problem in nonaqueous GPC and, when using a refractive index detector, it can cause a variable magnitude, negative area peak that may coelute with certain choices of totally permeated flow markers. This variable area negative peak may alter the apparent position of the flow marker when the flow rate has actually been invariant, thereby causing the user to falsely adjust data to compensate for the flow error. Similar problems can occur with the elution of positive peaks that are not exactly identical in elution to the totally permeated flow marker. Species that often contribute to these problems are residual monomer, reactants, surfactants, by-products, or buffers from the synthesis of the polymer. 

For acrylate polymers with higher levels of carboxylic acids, THF can be modified by the addition of acids such as acetic, phosphoric, or trifluoroacetic. Levels as high as 10% acetic acid are considered acceptable by most manufacturers for their styrene/DVB columns. If such a modified mobile phase is used, it may need to be premixed rather than generated using a dynamic mixing HPLC pump because on-line mixing often leads to much noisier baselines, particularly when using a refractive index detector. 

Refractive index detectors. These bulk property detectors are based on the change of refractive index of the eluant from the column with respect to pure mobile phase. Although they are widely used, the refractive index detectors suffer from several disadvantages — lack of high sensitivity, lack of suitability for gradient elution, and the need for strict temperature control ( + 0.001 °C) to operate at their highest sensitivity. A pulseless pump, or a reciprocating pump equipped with a pulse dampener, must also be employed. The effect of these limitations may to some extent be overcome by the use of differential systems in which the column eluant is compared with a reference flow of pure mobile phase. The two chief types of RI detector are as follows. 

A common error is to confuse the GPC distribution with the weight distribution. The response of a refractive index detector is proportional to the mass of polymer. The GPC elution volume (V) typically scales according to the logarithm of the degree of polymerization (or the logarithm of the molecular... 

Thin-layer chromatography (TLC) is used both for characterization of alcohol sulfates and alcohol ether sulfates and for their analysis in mixtures. This technique, combined with the use of scanning densitometers, is a quantitative analytical method. TLC is preferred to HPLC in this case as anionic surfactants do not contain strong chromophores and the refractive index detector is of low sensitivity and not suitable for gradient elution. A recent development in HPLC detector technology, the evaporative light-scattering detector, will probably overcome these sensitivity problems. 

Sodium dodecyl sulfate present in hydrophilic ointments has been determined by TLC on silica gel with flame ionization detection, which was considered better than the colorimetric method. TLC is preferred to HPLC in this case due to the low sensitivity of the refractive index detector that makes difficult the analysis of small amounts of sodium dodecyl sulfate [284]. 

If the mixture to be separated contains fairly polar materials, the silica may need to be deactivated by a more polar solvent such as ethyl acetate, propanol or even methanol. As already discussed, polar solutes are avidly adsorbed by silica gel and thus the optimum concentration is likely to be low, e.g. l-4%v/v and consequently, a little difficult to control in a reproducible manner. Ethyl acetate is the most useful moderator as it is significantly less polar than propanol or methanol and thus, more controllable, but unfortunately adsorbs in the UV range and can only be used in the mobile phase at concentrations up to about 5%v/v. Above this concentration the mobile phase may be opaque to the detector and thus, the solutes will not be discernible against the background adsorption of the mobile phase. If a detector such as the refractive index detector is employed then there is no restriction on the concentration of the moderator. Propanol and methanol are transparent in the UV so their presence does not effect the performance of a UV detector. However, their polarity is much greater than that of ethyl acetate and thus, the adjustment of the optimum moderator concentration is more difficult and not easy to reproduce accurately. For more polar mixtures it is better to explore the possibility of a reverse phase (which will be discussed shortly) than attempt to utilize silica gel out of the range of solutes for which it is appropriate. 

The flow sensitivity of a detector will also be one of the factors that determines the long term noise and thus will influence the sensitivity or minimum detectable concentration of the detector.lt is usually measured as the change in detector output for unit change in flow rate through the sensor cell. Again, the refractive index detector is the most sensitive to flow rate changes. 

The refractive index detector, in general, is a choice of last resort and is used for those applications where, for one reason or another, all other detectors are inappropriate or impractical. However, the detector has one particular area of application for which it is unique and that is in the separation and analysis of polymers. In general, for those polymers that contain more than six monomer units, the refractive index is directly proportional to the concentration of the polymer and is practically independent of the molecular weight. Thus, a quantitative analysis of a polymer mixture can be obtained by the simple normalization of the peak areas in the chromatogram, there being no need for the use of individual response factors. Some typical specifications for the refractive index detector are as follows ... 

Detection of the these types of compounds are sometimes difficult as many components of cosmetic products are aliphatic, do not possess a UV chromophore and are not easily reacted to give fluorescent derivatives. Providing the concentration of the component of interest is sufficiently high, then a refractive index detector is often used. If... 

The normalization method is the easiest and most straightforward to use but, unfortunately, it is also the least likely to be appropriate for most LC analyses. To be applicable, the detector must have the same response to all the components of the sample. An exceptional example, where the normalization procedure is frequently used, is in the analysis of polymers by exclusion chromatography using the refractive index detector. The refractive index of a specific polymer is a constant for all polymers of that type having more than 6 monomer units. Under these conditions normalization is the obvious quantitative method to use. 

The choice of detector is often crucial to the success of a particular HPLC method. A number are in routine use, including the UV, fluorescence, electrochemical, conductivity and refractive index detectors, and each has particular advantages and disadvantages, details of which can be found elsewhere [2-4],... 

This classification is concerned with whether the detector monitors a property of the solute (analyte), e.g. the UV detector, or a change in some property of the solvent (mobile phase) caused by the presence of an analyte, e.g. the refractive index detector. 

Adsorption chromatography using small particle silica or alumina has also been employed in the separation of biologically meaningful substances. Phospholipids, for example, have been separated on silica (38). One of the big problems for such substances is detection, since many of the compounds are not U.V. active. Generally, the refractive index detector is employed for isocratic operation, and the moving wire detector for gradient operation. Formation of U.V.-active derivatives is also possible (39). 

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