ANALYTICAL CHEMISTRY, VOL. 50, NO. 7, JUNE 1978
1017
Pulse Dampening System for High Pressure Liquid Chromatography Dominic A. Ventura and John G. Nikelly” Department of Chemistry, Philadelphia College of Pharmacy and Science, Philadephia, Pennsylvania 19 104
Mechanical reciprocating piston pumps are widely used in high pressure liquid chromatography (HPLC) owing to several advantages, including relatively low cost, constant flow that is independent of column backpressure and small internal volume which places no restriction on the size of the solvent reservoir. Unfortunately, the main disadvantage of this type of pump is that it produces a pulsating flow. Since most detection systems are flow sensitive, these pumps produce synchronous baseline fluctuations which tend to limit the sensitivity of the HPLC system. There are several ways to reduce flow pulsations ( I , 2). In most commercial instruments equipped with reciprocating pumps, flow pulsations are reduced with a dampening device, a flexible metal vessel or bellows, which takes up some of the energy from the positive stroke and releases it when the pump refills, thus helping to maintain a fairly even pressure in the system. Unfortunately, these devices are limited to working pressures below 1200 psig ( 3 ) ;moreover, they are relatively expensive, costing several hundred dollars. Another often used pulse dampening device is a combination flow-resistance and capacitance, comprising a 5-m length of 0.01-in. i.d. capillary tubing or fine needle valve (flow-restrictor) in series with a pressure gauge (Bourdon tube or diaphragm type) as ballast. Unfortunately such systems are susceptible to clogging by particulate matter and require considerably higher operating pump pressures compared to systems without a restrictor ( 4 ) . More important, this type of dampening does not seem to work well with pumps that have a relative low stroke rate such as the popular Milton Roy pumps which have 27 strokes per min and which work directly on the mobile phase (2). This paper describes the practical use of an inexpensive commercially available dampening device, a flexible hose connector, which substantially reduces or eliminates pressure and flow pulsations in the solvent and fluctuations in the baseline. The connector is designed mainly for industrial uses (for installing flexible loops to allow for thermal expansion, misalignment, intermittent flexing, static bends and vibration) but it is essentially a bellows, i.e., it stores about one half of the solvent moved out of the pump with each positive stroke and then restores it to the system during the suction stroke a t about 99% of the undampened surge pressure of the pump.
EXPERIMENTAL In addition to the usual fittings and capillary tubing, the complete dampener assembly includes a conventional pressure gauge (Bourdon tube, 0 to 3000 psig range) and an annularly corrugated, flexible stainless steel hose connector, as ballast, Swagelok part number SS-4HO-6S4, 25 cm long or SS-4HO-6L4,80cm long, Crawford Fitting Company, Cleveland, Ohio 44139. The connector is protected with a braided wire sheath and is capable of operating at pressures up to 2660 psig. The dampener assembly was used as part of an HPLC system comprising a synchronous reciprocating pump, “Minipump”, and UV Monitor, Model 1205 (Laboratory Control Division, Milton Roy Company, Riviera Beach, Fla. 33404). In some experiments, a commercially available dampening system was used, Laboratory Data Control, Model 709. Samples were injected with a Valco Sample Injection Valve, Series SVOV-6-1(Glenco Scientific,Inc., Houston, Texas 77007). The analytical column was 30 cm x 3.9 mm i.d., packed with 10-pm particles of reverse phase CIS Bondapak (Waters Associates, Inc., Milford, Mass. 01757). P r o c e d u r e . The dampening assembly, its components arranged in various configurations, Le., in various positions relative
A
6 0
B
h
Flgure 1. Pulse dampening system. Three basic arrangements of the connector: (A) Ballast closed at one end, (B) Flow-through ballast, and (C) Gauge directly connected to ballast
A
B
C
Apparatus.
0003-2700/78/0350-1017$01 .OO/O
D Flgure 2. Comparison of ballast effects. Sample, 10 pL of sulfadiazine solution, 10 p g per mL. Solvent, 10% MeOH in phosphate buffer, pH 7.07, 0.7 mL/min flow rate. Column, 25 cm X 4 mm i.d., reverse phase, Cle, 10-pm particle size. Detector sensitivity, 0.04 absorbance unit full scale, 1-mV recorder. (A) Without gauge or connector, (B) With gauge only, (C) With gauge and 25-cm connector, (D) With gauge and 80-cm connector
t o each other, was tested by noting the range of pressure pulses on the pressure gauge and the effect of the pulses on the detector output (recorder base line). Other variables considered and evaluated were the length or size of the flexible hose connector 0 1978 American Chemical Society
1018
ANALYTICAL CHEMISTRY, VOL. 50, NO. 7,JUNE 1978
30 RETENTION
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TIME, M I N .
Figure 3. Comparison with commercil pulse dampener. Sample: 10 /.LLmethyl xanthines and methyl uric acids, 10 yg/mL each. Mobile phase: 12% MeOH/buffer pH 4.7,1.1 mL/min flow rate, 1100 psig pressure. Column: 30 cm X 3.9-cm i.d. packed with lOy, /.L& Bondapak, reverse phase; 0.02absorbance unit full scale. (A) With Model 709 pulse dampener, LDC. (B) With flexible connector, 80 cm long
and the operating pressure of the HPLC system.
RESULTS AND DISCUSSION Several variables were considered in evaluating the dampening system. The most important of these factors appears to be the way the connector is arranged in the system, e.g., in series, in parallel, flow-through, etc. There are, however, only three basic arrangements, and these are shown schematically in Figure 1. It was found that when the ballast was closed at one end (arrangement A in Figure 1) or when it was used as a flow-through connector (arrangement B), the depulsing effect was relatively small. When arranged in configuration C, the gauge and the ballast forming a "T" branch of the flow system, the recorder baseline fluctuations are sharply reduced to essentially background, pulseless levels. This is illustrated in Figure 2 which shows the comparative depulsing effects of the pressure gauge and flexible connector. T h e bottom recording representing the combination of
pressure gauge and 80-cm connector shows a baseline that is virtually pulseless a t a detector sensitivity of 0.04 absorbance unit full scale, 1 mV recorder. It should be noted that the addition of the flexible connector increases the total mobile phase volume in the HPLC system, by 20 to 40 mL, thus increasing the time required to reach equilibrium when the mobile phase is changed (about 10 min.). Another serious drawback is the additional time required, when starting up, to reach the desired operating pressure. Consequently, it was found practical, whenever the system is not in use or when the stop-flow injection procedure is being used, to isolate the ballast volume from the rest of the system by using a shut-off or needle valve between the connector and the solvent line. This reduces or eliminates the time required to reach operating pressure and allows rapid changeover of the solvent system. (When changing the mobile phase between immiscible solvents, e.g., water to hexane, an intermediate solvent is used; more important, the pressure gauge may be disconnected to allow the flexible connector to be flushed through with the new mobile phase. The complete mobile phase change-over takes only a few minutes.) The system was used a t pressures from 500 to 1500 psig. For pressures below 800 psig, it made no difference which connector length was used, while above 800 psig, the longer of the two connectors, 80 cm, was somewhat more effective in reducing the pressure pulses. In fact, the pulse-dampening effect obtained with the long connector a t operating pressures above 1000 psig is virtually the same as the effect obtained with a commercial dampening system. This is demonstrated in Figure 3 which compares chromatograms A and B, A obtained using an LDC model 709 dampening system costing approximately $700, and B using the 80-cm Swagelok flexible connector (with pressure gauge); in all other respects the experimental conditions were identical. (The test mixture in this case was an actual clinical sample of theophylline with metabolites and other endogenous xanthines.)
LITERATURE CITED (1) L. R. Snyder and J. J. Kirkland, "Introduction To Modern Liquid Chromatography". Wiley-Interscience, New York, N.Y., 1974, pp 101-102. (2) J. N. Done, "Idealized Equipment Design for HPLC", in "Practical High Performance Liquid Chromatography", C. F. Simpson, Ed., Heyden & Son, Inc., New York, N.Y., 1976, pp 71-2. (3) Instruction Manual, Pulse Dampener Model 709, Laboratory Data Control Division, Milton Roy Company, Riviera Beach, Fla. 33404. (4) K. K. Stewart, Anal. Chem., 49, 2125-2126 (1977).
RECEIVEDfor review December 27, 1977. Accepted March 6, 1978.
Mass Spectrometer Ion Multiplier Noise Filter Based on an Analog Delay Device D. L. Doerfler and I. M. Campbell* Department o f l i f e Sciences, Faculty o f Arts and Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 75260
Ion multipliers are inherently noisy devices, particularly when high sensitivity is demanded of them. In mass spectrometry, where ion multipliers are used commonly to quantitate ion abundances during scans of ion mass/charge ratio, noise can be especially disturbing. The noise problem becomes critical in processing data from fast-scanning mass spectrometers coupled to gas chromatographs. T o discriminate between true ion multiplier signal maxima and spurious, noise-associated maxima in computerized combined gas
chromatograph/mass spectrometers, noise filters have to be included either in the hardware of the interface or in the processing software. Software noise filters operate by summing successive, appropriately weighted and digitized values of the input signal. They have the advantage of being versatile, of being free from drift and distortion, and of being programmable to any desired degree of accuracy and sophistication (1-3). Software filters, however, tend to be time-consuming and frequently a compromise has to be struck
0003-2700/78/0350-1018$01.00/0 0 1978 American Chemical Society
