Peak capacity reduced

Cooling loads are most significant on hot summer afternoons when electric utilities typically have annual peak loads. Lower solar heat gains lead to reduced peaks, which reduce peak demand charges and power plant capacity requircliiclits. 

As mentioned earlier, high-speed separation is necessary to carry out fast, comprehensive 2D HPLC. The polymer monoliths have not been employed in such 2D HPLC, probably because permeability of polymer monoliths is not high enough to allow fast elution of the second dimension (2nd-D) in simple 2D operation, and the gradient cycle at the 2nd-D cannot be so fast to allow online 2D operation without reducing peak capacity at first dimension (lst-D). 

The proposed estimate has several limitations. When taking into account the limited orthogonality of investigated 2DLC modes, the practical peak capacity is reduced approximately to half. It needs to be also emphasized that a full separation power of the first LC dimension is realized only when the number of collected fractions exceeds its peak capacity (Murphy et al., 1998). If the number of fractions analyzed is low, the achievable chromatographic peak capacity suffers. 

It has been argued that in a typical 2DLC proteomic experiment, with only a limited number of fractions submitted for analysis in the second LC dimension, chromatographic peak capacity is less than 1000. This value is considerably lower than the expected sample complexity. Additional resolution is offered by MS, which represents another separation dimension. With the peak capacity defined as the number of MS/MS scans (peptide identifications) accomplished within the LC analysis time, the MS-derived peak capacity was estimated to be in an order of tens of thousands. While the MS peak capacity is virtually independent of LC separation performance, the complexity of the sample entering the MS instrument still defines the quality of MS/MS data acquisition. The primary goal of 2DLC separation is to reduce the complexity of the sample (and concentrate it, if possible) to a level acceptable for MS/MS analysis. What is the acceptable level of complexity to maintain the reliability and the repeatability of DDA experiments remains to be seen. 

A reduced peak capacity in one domain may be counterbalanced by an increased peak capacity in another domain. If we know the average peak width of a chromatographic separation and the gradient duration, we can calculate the maximum number of peaks that can be separated. (Note peak capacity does not mean that this number of compounds in a sample will be separated they may still co-elute). That means we can operate between two limits (1) a peak capacity of zero representing a flow injection analysis and (2) a minimal required peak capacity that defines the peak capacity to separate all compounds in a given mixture. Unfortunately, especially in the early stages of drug... 

In general, LC peak capacity can be reduced if it is counter-balanced by an increased MS peak capacity in the orthogonal mass domain. Increasing the dimensions in LC/MS separations is another option to increase overall peak capacity in a given time, but comes at a cost of increased complexity of instruments and data evaluation. In the mass domain, highly resolving mass spectrometers round out the choices. Nevertheless, every throughput optimization step should be viewed in relation to... 

An example of a fast, well-executed separation is shown in Figure 8. A 2-mm i.d. 3 cm 3.5- J,m column was used in a rapid, 1-min gradient. In order to maximize the peak capacity, a high flow rate was used 2mL/min. To reduce the column backpressure, the separation was carried out at 60°C. A fast detector sampling rate was employed to keep up with the high speed of the separation. The injection was coordinated with the arrival of the gradient at the top of the column in order to eliminate the... 

The reports from the integrator consist of retention time and sample amount for each integrated peak. These are transmitted to a small computer. The integrator has the capacity to process up to 250 peaks in a run. However, because of the limited memory space of the computer, we had to decrease the number of peaks processed. Chromatographic runs with more than 150 peaks were reduced to 150 peaks by elimination of those with the smallest area. The reduced reports were then stored on tape. 

The approaches described above are designed to increase peak capacity and thus reduce peak overlap. A radically different approach involves accepting component overlap as inevitable and directing attention at numerical rather than physical peak resolution. Numerical resolution allows the recovery of analytical information but not the recovery of purified components. 

In analytical applications of liquid chromatography the most common causes of peak asymmetry are mixed mechanisms of retention, incompatibility of the sample with the chromatographic mobile phase, or development of excessive void volume at the head of the column. In preparative applications of liquid chromatography and related techniques, column overload can also contribute to peak asymmetry. The causes of severe peak asymmetry in analytical applications should be identified and corrected because they are frequently accompanied by concentration-dependent retention, non-linear calibration curves and poor precision. In addition, peak asymmetry can significantly compromise column efficiency leading, in turn, to reduced resolution and lower peak capacity (see sections 2.5 and 2.6). 

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