Plate count —

Large quantities of water are used in the food, drink, and pharmaceutical industries. This is frequently subjected to further treatment on entering the plant and is usually of very high quality. Plate counts are used widely in these industries to assess the effectiveness of this extra treatment. These industries also frequently 

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First, solutes with larger electrophoretic mobilities (in the same direction as the electroosmotic flow) have greater efficiencies thus, smaller, more highly charged solutes are not only the first solutes to elute, but do so with greater efficiency. Second, efficiency in capillary electrophoresis is independent of the capillary s length. Typical theoretical plate counts are approximately 100,000-200,000 for capillary electrophoresis. 

Luminol chemiluminescence has also been recommended for measuring bacteria populations (304,305). The luminol—hydrogen peroxide reaction is catalyzed by the iron porphyrins contained in bacteria, and the light intensity is proportional to the bacterial concentration. The method is rapid, especially compared to the two-day period required by the microbiological plate-count method, and it correlates weU with the latter when used to determine bacteria... 

Heterotrophic plate count n/a TT3 HPC has no health effects, but can indicate how effective treatment is at controlling microorganisms. HPC measures a range of bacteria that are naturally present in the environment... 

After packing and testing the column, one may find that the plate count or the peak symmetry is unsatisfactory. In this case some adjustments need to... 

Additional factors influencing column performance are the type and quality of the packing process, which mainly determines the theoretical plate count (N) of the column. In contrast to HPLC columns the efficiency of the separation itself is determined predominantly by the quality of the sorbent alone (pore... 

TABLE 9.7 Dependence of Theoretical Plate Count and Resolution on Sorbent Particle Size°... 

PSS uses the following formulae (22,23) to calculate plate count, asymmetry, and resolution. PSS uses test conditions that conform to the ISO/EN 13885 and DIN 55672 requirements for SEC sample testing. 

There are different ways to calculate the theoretical plate count of a column. PSS SEC column plate counts are measured using the so-called half-height method. Figure 9.9 shows a graphic representation of this test. 

The following formula is used to calculate the plate count ... 

Plate count should always be tested with a monodisperse sample of low molecular weight. Polymers can also be used, but they show much lower plate counts because their diffusion coefficients are much smaller than those of low molecular weight compounds. 

The reader should note that the value of the theoretical plate count depends on the sample chosen for testing. PSS always specifies on the PSS column quality certificate which sample has been used for measuring plate count and the exact test conditions. 

Using other methods for the calculation of plate count can result in different numbers, depending on peak shape. It should also be kept in mind that many other operational parameters, such as eluent viscosity, column temperature, flow rate, and injection volume, will influence the results of the plate count determination. 

For SEC separations of polymers, column efficiency is better characterized by specific resolution, R,p, and efficiency, T, than by theoretical plate count. Peak resolution, R is calculated according to (7) ... 

All packing materials produced at PSS are tested for all relevant properties. This includes physical tests (e.g., pressure stability, temperature stability, permeability, particle size distribution, porosity) as well as chromatographic tests using packed columns (plate count, resolution, peak symmetry, calibration curves). PSS uses inverse SEC methodology (26,27) to determine chromatographic-active sorbent properties such as surface area, pore volume, average pore size, and pore size distribution. Table 9.10 shows details on inverse SEC tests on PSS SDV sorbent as an example. Pig. 9.10 shows the dependence... 

After column packing, each column is tested for theoretical plate count, peak symmetry, resolution, pore volume, and back pressure. If one of these tests fails the column is removed from the production cycle. If a PSS SEC column is kept in storage for a longer time, it is retested for theoretical plate count, peak symmetry, resolution, pore volume, and back pressure prior to shipping to the customer to prove up-to-date column performance. 

FIGURE 9.12 Quality control chart of PSS SDV columns plate count (per m) measurements over a I0>year period 5% limits are also shown. 

Each SynChropak column is tested chromatographically to assure that it has been packed according to specifications. For SynChropak GPC columns, a mixture of a high molecular weight DNA and glycyltyrosine, a dipeptide, is used to evaluate internal volume and efficiency. The mobile phase used for the test is 0.1 M potassium phosphate, pH 7, and the flow rate is 0.5 ml/min for 4.6-mm i.d. columns. Minimum plate count values and operational flow rates are listed in Table 10.4 for 4.6-mm i.d. columns of all supports and the various diameters of the SynChropak GPC 100 columns. 

SynChropak CATSEC columns are evaluated similarly using a polyvinyl-pyridine standard of molecular weight 600,000 and cytidine. The mobile phase is 0.1 % trifluoroacetic (TEA) acid containing 0.2 M sodium chloride. Minimum plate counts are listed in Table 10.4. 

FIGURE 13.21 A series of polymer additives using the infrared detector at 5.78 micron. Efficiencies were calculated using the last peak, Tinuvin P, and a plate count of 290,000 was achieved. 

Figures 13.25-13.28 show the ultrahigh resolution separations in chloroform of polystyrene standards, polytetramethylene glycol, urethanes and isocyanates, and epoxy resins, respectively. Multiple column sets of anywhere from two to six columns in series have been used for well over a year with no apparent loss of efficiency. The 500- and 10 -A gels can easily tolerate 15,000 psi or more. In fact, the limiting factor in the number of columns that can be used in series is generally the pump or injector in the FIPLC system. A pump capable of 10,000 psi operation should allow the use of a column bank of 10-12 50-cm columns with a total plate count of 500,000 or more.

FIGURE 13.24 A mixture of hydrocarbons and some polyethylene standards at I45°C. Column backpressure was approximately 5500 psi. Plate counts calculated on the hexane and heptane peaks yield 204,000 plates. 

FIGURE 13.25 Using chloroform as the solvent, a mixture of polystyrene standards were nicely separated on the 3-m set of columns. Run times here were 160 min. Plate count for toluene was calculated at 240,000 plates. The 500 MW Standard is separated nicely into its oligomers. 

Most size exclusion chromatography (SEC) practitioners select their columns primarily to cover the molar mass area of interest and to ensure compatibility with the mobile phase(s) applied. A further parameter to judge is the column efficiency expressed, e.g., by the theoretical plate count or related values, which are measured by appropriate low molar mass probes. It follows the apparent linearity of the calibration dependence and the attainable selectivity of separation the latter parameter is in turn connected with the width of the molar mass range covered by the column and depends on both the pore size distribution and the pore volume of the packing material. Other important column parameters are the column production repeatability, availability, and price. Unfortunately, the interactive properties of SEC columns are often overlooked. 

Column manufacturers normally provide basic information about their columns, such as plate count, particle size, exclusion limit, and calibration curve. This information is necessary and fundamental, however, it is not sufficient to allow users to make an intelligent decision about a column for a specific application. For example, separation efficiency, the dependence of separation efficiency on the mobile phase, the ability to separate the system peaks from the polymer peak, the symmetry of the polymer peak, and the possible interaction with polymers are seldom provided. 

Two linear columns from Showa Denko, Shodex SB-806M and Shodex SB-806MHQ, and two linear columns from TosoHaas, TSK GM-PWxl and TSK GM-PW, were evaluated. Prior to the evaluation, the number of theoretical plates for Shodex SB-806MHQ, SB-806M, PWxl, and PW was determined to be 15,100, 15,700, 11,390, and 4710, respectively, as per manufacturer inspection. The lower plate count of the TSK PW column is due to the larger particle size of this column. Two mobile phases, water with 0.1 M LiNOi and 50 50 methanol/water (v/v) with 0.1 M LiNOi, were used for each of the four columns. These four columns were new and only PEO and PVP were analyzed with these columns in this study. Waters Ultrahydrogel columns have also been used in this laboratory. However, Ultrahydrogel columns are exactly the same as the TSK GM-PWxl columns based on the calibrations curves supplied by the manufacturers and by the pyrolysis GC data discussed later. 

It is difficult to decide what should serve as adequate column quality parameters for describing the performance of a set of GPC columns. The two most common measures are plate count and resolution. While both of these can be useful for monitoring the performance of a column set over time, it is not generally possible to a priori specify the performance needed for a specific analysis. This will depend on the nature of the polymer itself, as well as the other matrix components. 

Most GPC columns are provided with vendor estimates of the plate count of the column and a chromatogram of a series of test peaks. These plate count estimates are usually obtained using small molecule analytes that elute at the total permeation volume (Vp) of the column. The Gaussian peak shape model... 

Scientific (Northbrook, IL) contain a silica support with a -y-glycidoxypropylsi-lane-bonded phase to minimize interaction with anionic and neutral polymers. The columns come in five different pore sizes ranging from 100 to 4000 A. The packing material has a diameter from 5 to 10 /cm and yields in excess of 10,000 plate counts. With a rigid silica packing material, the columns can withstand high pressure (maximum of 3000 psi) and can be used under a variety of salt and/or buffered conditions. A mobile phase above pH 8, however, will dissolve the silica support of the column (21). A summary of the experimental conditions used for Synchropak columns is described in Table 20.8. 

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