Chemical Instrumentation Edited by GALEN W. EWING, Seton Hall Univenity, So. Oronge, N. J. 07079
These articles are intended lo serve the readers o f ~ mJOURNAL s by calling allention to new developments i n ihe theory, design, or availability of chemical laboratory instrumenlation, or by presenting useful insights and ezplanalions of topics that are of practical imporlanee lo those who use, or leach the use of, modern instrumentalia and instrumental techniques. The editor invites correspondence from prospeclive contributors.
detector itself acts as a microcolumn. Micro-dsarption detectors arenondestructive and are subject t o other thermal effects such as thermal conductivity and heat capacity of the solvent. They therefore require accurate calibration with pure standards before they can be used to obtain acceptable quantitative data. Several companies manufacture thermal detectors.
Dr. Hupe Apparatebau
LIII. Liquid Chromatography DetectorsHANS VEENING, Deportment of Chemistry, Bucknell University, Lewisburg, Pa. 17837 curve. This peak shape is illustrated in Pigrue 30. Hupe and Bayer (31) have The principle of operation of thermal painted out thal the initial hypothesis by ~ ~ -workers that the signal should redetectors a180 known as ' L m i ~ r ~ - a d ~other tion" or "heat of adsorption" detectors semble t,he first derivative of the eoncendepends on temperature changes taking trat,ian profile, was not correct. The place due to the heat of sorption on an signal is non-Gaussian because heat is active solid surface. These detectors were transferred out of the cell by the eluent in first reported by Claxton (YO), and Hupe the direction of flow during the sorption and Bayeyer (Sl), and are now mannprocess. I n Figure 30 the distance AC is factured by a number of companies. the positive peak width; CE, the negat,ive Generally a portion of the dsorbent peak width; AE, t,ho tot,al peak widt,h; column packing is contained in ~1 small and DB, the iota1 peak height,. chamber s t the column outlet. A second These dotector can be used in applieachamber filled with an inert material such tions involving liquid-solid, liquid-liquid, as glitss microbeads is used to achieve a ion-exchange, and gel-permeation chromareference signal. This assembly is c a r e t,ography. They are able to sense any fully thermostatted. Bolh chambers contypeof component separable by these techtain matched thermistors (1 and 2 in Fig. niques. They have an advantage in that 29) which constitute the memuring arms of +,hey are ideally suited for determining a Wheatstone bridge. As the elr~ted optimum sepmation conditions since the sample is admrbed on the ~olid,a. local temperature change initiates a signal. A reverse thermal react,ion occurs when the sample is relemed later. The detector signal therefore, represents a rather unconvehional peak ~ h a p e which , is referred to as a. "different,in.l peak" due t,o its rough similarity to the derivative of the Gaussian
THERMAL DETECTORS
EFFLUENT
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STAINLESS STEEL SHELL
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Figure 29. Micro-odrorption detector utilizing matched thermirtorr, (1) ond (2). (Courtery of Dr. Hupe Apporatebw.1
Figure 30. Micro-adsorption detector response I injection; AC poritive peak width; CE negative width; AE totol peak width; and LIB total peak height. (Redrown from Hvpe and Bayer (311 with permission of The Journal of Chrnmot~~raphic Science.)
This detector has already been partially described in the previous seotion. It is oonstructed of stainless steel snd Teflon. The bridge voltage is adjustable between 0.1 and 10 V. The microvolt amplifier is chopper stabilized and has an input voltage for full recorder response of 100 pV and an attenuator with steps ranging from 0.1 to 10 mV. The instrument is marketed in the U S . by Ionics Resemch, Inc., and the total package includes the d e tector, a dc power supply, and a microvolt amplifier.
Jeolco Thermal Reaction Energy Detector The Jeolco thermal detector was the first one of its type to be made commercially avtvailahle. As in other thermal detectors, the method of measurement depends on temperature variations resulting from the migration of the sample between the stationary and mobile phase. The JLC-A detection system consists of a double thermistor within the detection column for detecting reaction heat, a controller, a. signal amplifier and a power supply. Optional packing materials can be used in the detection system. Recording sensitivity can be adjusted in seven steps from f l to f O.OOl°C, where temperature correspondv to full scale recorder deflection. The detection limit is to 10-7 mole, and the quantitative The detector error is claimed to be &3%. temperature is variable in eight steps from 10 to 50°C.
NesterIFaust Heot of Adsorption Detector This unit is first charged with adsorbent which is the same or similar to that used in separating the sample components of the column. This insures that a thermal reaction will be detected and that the sample will not be retained too long to cause a.loss of resolution. The cell is assembled as shown in Figure 31 and then immersed with the amplifier chamher into an insulated water bath. Cohmn efluent is transported to the hot,t,om of the cell using Teflon capillary tubing and discharged from the top into a fraction eollect,or or waste container. The
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controls and meter provided an the electronic console. It is absolutely essential that a constant, surge free flow be available for operation of this detector. Since the thermistors are, by necessity, mounted directly in the stream, any short or long term flow rate changes will be detected and recorded. This detector is shown photograpbicdly in Figure 32.
Varian Aerogmph MicroAdsorption Detector The micro-adsorption detector marketed by Varian Aerograph is based an the Hupe instrument (Fig. 29) and has been described by Munk (36). A 3 X 10-ST temperature change in the detection thermistor gives a 1 mV sigrral at maximum sensitivity with a corresponding short termnoise (100 sec or less) of less than2%. The linear dynamic range is 2 X loa, and the cell volume is 9 pl. The detector and its associated water bath are shown photographically in Figure 33.
Figure 31. Cell assembly for the Nerkr/Faust heel of mdsorption detector. Column effluent ontors through theTeflon disc at tho boHom and emerges thmvgh mother identical disc at tho top. (Courtesy of Nerter/Faurt Manufacturing ~ o r p o r a d )
Figure 32. Norter/Faust heat of adsorption detector. lnrulokd housing on the left, elesIronic plug-in console on the right. (Photograph courtesy of Nerter/Foust Manufacturing Corporation.)
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Figure 33. r.,..,.r,, micm-adsorption detector. (Photograph courtesy of Varian Aemgraph.)
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Circle NI. IM an Realm' Service Card
Chemkal lnstrumentatlon FLUORESCENCE DETECTORS The use of fluorescence as a. technique for monitoring high speed LC seems promising. Several mmanufscturers are presently engaged in efforts to design floorescenee detectors. The technique has advantages, namely selecbivity for oertain
processes such as quenching, solvent flu; rescence, and turhidit,~. The technique seems particularly well suited, however, for monitorine " such hioloeicallv .> " active materials 8s proteins, porphyrins, vitamins, and plant pigments. Fluorescence-monitoring systems have been wed primarily in preparative scale separations of proteins using gel-permea tion columns (%, 55). No work has been reported in which fluorescence is used a8 a detection system for high speed, high pressure, LC, although one company (LDC) now markets a high performance detector useful for high resolution work. American Instrument Company manufactures a. Fluoro-Microphotometer, a filter fluorometer which can also be converted to a calorimeter or turbidimeter. This instrument has been used as a fluorescence monitor for streams in conjunction with the Technicon AutoAnalyzer. The flow cell is round and has 8. 6-mm light path. The instrument is of solid state circuitry and has recorder output facilities. Laboratory Data Control has recently placed its new high performance differential fluoroMonitor on the msrket. I t contains a. dual cell allowing for correction in hackeronnd fluorescence. The cell quinine sulfate. G. K. Turner Associates manufactures a flow attachment permitting continuous fluorescenw measurement of LC column efluents using a. Turner model 111 selfhalrtncing, double beam, filter fluorometer. This fluorometer measures the ratio of intensities of fluorescent light and a constant proportion of light from the source, tho.; compensating for floctulttions in light intensity. The instrument can he equipped with a wide variety of light sources (254 to 600 nm), including a mercury lamp as well as severd fluorescent lamps with special phosphors. A photo-
Figure 34. G. K. Turner model 1 1 1 Ruororneter with continuous flow attachment, and "robot limit controller" (on the right). (Photograph courtesy of G. K. Tvrner Asrocioter.)
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semhly is shown in Figure 34.
CONDUCTANCE DETECTORS ~~
multiplier tube is used as detector. Provision is made for direct hook-up to 0-10 mV recorders. The instrument is capable of detecting 1.5 parts/lOL1of quinine sulfate. Continuous flow attachments for the Turner fluorometer are available with three different size flow cells, 4, 10, and 19 mm (i.d.). The volume for the smallest cell is 0.5 ml and is available in barosilicate or qumte. A small recorder and a "Robot Limit Controller" for actuating external equipment at ppre-set levels of fluorescence are also available. The complete as-
LC detectors based on the principle of conductance consist of a flow cell containing s pair of metd electrodes. These electrodes are incorporated into the usual Wheatstone bridge circuit and measurement occurs by use of an alternating current. These detectors are sold by a number of manufacturers and have also been used for amino acid analyzers and gel-permeation techniques. They are simple to operate and easy to maintain and clean. Chromatronix, Inc., manufactures s.
new improved model CM-1A microflow conductivity cell and conductivity met,er with recorder output designed specifically for high resolution LC. I t provides continuous nand-meter disnlav. It nti-
-. ? w e 36. Chromatronix conductivity meter crotlow conductivity cell and LC ~ o l u m n
Figure Inc.1
35. Chmmotmnix MCC-75 microflow conductiviv, c e l l schematic.
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(Courtesy of Chmmotmnir,
hotogroph courtesy of Chromotronix, Inc.1
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volume and has a pressure rating of 500 psi. Full range selectivity from 1 to 1000 rmho is provided, and the limit of detection is 0.001 rrnho. The cell also contains a temperature compensating thermistor. The cell, the meter and a column are shown in Figure 36. The NesterFaust conductivity detector is of a three-electrode design thus compensating for changes in system plumbing. Dimension3 of the cell are 10 cm (length) and 2 mm (id.). The range of specific resistance which can he accommodxted is 10 to 100,000 ohm-em. The cell constant is 10 em-'. The range selectivit,y varies from 10 to lo5mho/cm. Sensitivity is one part in 10,000 change in conductivity. The total effluent stream passes through the flow-throngh cell, alt,hough the stream can he split. The schematic representation of the flow cell and associated circuitry is shown in Figure 37, the electronic plugin console is shown in Figure 38.
Figure 37. Schematic representation of the Netter/Fwst conductivity detector. (Courte3y Nerter/Faurt Manufacturing Corporotion.)
Figure 38. Nerter/Fourt conductivity plug-in conrole. (Photograph courtesy Nerter/Fourt Monufocturing C0~~orotion.1
RADIOACTIVITY AND POLAROGRAPHIC DETECTORS Radiometric techniques have not been used a. great d e d in liquid chromatogrnphie (Continued a page A758)
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Chemical Instrumentation detection devices. Their useful features such a? ultra high sensitivity, adaptability to gradient elation and the fact that they are nondestructive make tthis type of a detector potentially applicable to LC. Utilization of beta radiation using s u s pended scintillators in continuous flow measurement has been described by MeGuinness and Cullen in a previous article in this series ( 3 4 ) . Palarographic detectors are not presently marketed, although they are being developed in certain laboratories. They should be very useful in detecting such reducible substances as metal ions, pesticides, alkaloids, aldehydes, and ketones. The design of a high speed LC polmographie detector has recently been described in a naner bv Koen. Huher.
dimensions of 1 X 1 mm. A 1 cm2 mercury pool is used ss the reference electrode. Column effluent flows past the DME in s. chamber volume of only a. few cubic millimeters. A one-second drop time of t,he DME enables precise measurement of fast peaks. Since ourrent varies widely aver the lifetime of a drop, it is necessary to damp the signal hy means of RGcircnits. The detector was successfully applied to the ehrornxtographie separation of phosphate insecticides hearing a. nitro group. Concontralions of mole/l could br determined with a standard deviation of better than 2'%. An aqueous mixture containing ethanol, acetic acid, sodium hydroxide, and potassium chloride was used a3 the eluent, and isooctane served as the stationary phase.
COMPLETE LIQUID CHROMATOGRAPHS I t seems appropriate to include a description of the complete LC units which are now available from severd manufacturers. The detectors included in these units have already been described. All of the liquid chromatographs which are now on the market include as their basic equip-
Figure 39. DuPont 820 liquid chmmotogroph. Photograph courtesy of E. I. duPont d e Nemoum and Campony.1
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Chemical Instrumentation .
ment, solvent reservoirs, a pumping system, sample injectors, and columns. A vast and sometimes confusing o ~ t i o nof detectors is usually available at extra cost. Recorders are bv and l a n e not included in
of delivering flow rates from 0.01 to 5.8 ml/min. A~vacuum degassing chamber, stainless steel columns, and a manud fraotion collector are part of the basic unit. The instrument rtceommod~testhe DuPont precision photometer (previously described) as its detector. Provision has been made for inclusion of other detector systems. This chromatograph is very versatile and can he adapted to various modes of LC. The Jeolco JLC-5AH amino acid amlyzer is capable of handling 12 samples suoeessivelv. I t is a useful instrument for large slale separations of amino acids in biologicaUy active samples such as food stuffs, pharmaceuticals, and physialogicsl fluids. I t consists of apumping compartment which houses five buffer reservoirs, two bufferpumps, two buffer exchangers, a. ninhydrin system, a ninhydrin washing liquid exchanger and a display panel far an elution flow system. I t is an extremely complex instrument which houses two columns, a special detection pump, water baths, samplers and column selectors. The instrument is supplied with the visible double beam detector already described. The analyzer can be sutomatically operated up to 48 hr and during this time can be programmed by means of a punched taDe. The unit is shown in Fieure 40.
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Figure 41. Varion Asrograph LCS-1000 ion exchange liquid chmmotograph. (Photograph courteq of Varian Aerogroph.1
top console. Included are pumps, buffer and gradient reservoirs, columns, and a UV photometer. A strip chart recorder is also provided. Any of the Varian Aerograph detectors previously described ($6) (differentialrefmatometer, UV photometer or micro-adsorption) can be incorporated into the unit. A wide range of column dimensions is avdahle m d separation of nucleotides can he performed in 1 hr or less. The chromatograph is pictured in Figure 41. The Waters ALC-100 liquid chromatograph is of modular design and includes a solvent reservoir, a pumping system (0 to 160 ml/hr, 1000 psi). The detectors which can be incorporated in this instrument are the Waters differential refractometers and the LDC UV monitor. The system can handle aqueous and organic solvents and is capable of preparative scale chromatoera~hvwithout modificsi tion. The unit is shown in Figure 42. The manufacturer has also recently placed a new, low cost, model ALCJOI high resolution chromatograph on the market. The instrument is optimised for operation with narrow bore columns and high - linear velocities. U
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Fioure 40. Jeolco JLC-SAH amino acid on01 " lyzer. (Photqroph courtesy of Jeolco (U.S.A.1,
Inc.1
NesterlFaust manufactures a model 1200 modular liquid chromatograph. The instrument includes a single or multiple system of pumps (0.6 to 9.6 ml/min) with a maximum attainable pressure of 2500 psi, as well as a solvent reservoir, a. fraction collector, and a wide variety of columns. The chromatograph can accommodste any of the Nester/Faust detectors already described. The company has recently also placed a low cost (model 1240) high resolution, high speed liquid chromatograph on the market. The Varian Aerograph LCSlOOO ion exchange liquid chromatograph is a high pressure, high resolution ion exchange system useful for quantitative separation of water soluMe ionizable compounds. All components are contained in a bench-
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Figure 42. Watem ALC-100 liquid chmmatograph. IPhotograph courtesy of Waters Arrociotes, I n 4
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REFERENCES (SO) C ~ ~ x v oG.. n , J . Chromnloo., 2 , 136 (19591. ( 8 1 ) HDPE, I L P . nxn ~ A Y Z R ,E., J . Gar Clwomntoa.. 6, 197 (1907). (SBI Tomnsr, hf., A N D Pamrp, L. J.. J . C h w moloo., 20, 299 (1965). (55) Escx, C.I.. nun TnmsL, A. L.,Federation Proooodin~s.Paper No. 1085, April, 1907. ($4) MCGCINNEIS,E. T.,AX" CL.LL%N, It.C.. ,I. CEIE?~. ED.,47, A9 (1970). (561 K o w , J. G.. H u m s , .I. B. K., POPFE,H.. A N D DEN BOEP, G.. J . Chmnalou. Sci.. 8 , 1 0 1 11U70> ."-\.".",.
(36) MUNI, M. N., P w e r No. 84, Pittsburgh Confrrenoe on Anslytioal Chemistry and Anplied Speotroseopy. Clevelsnd, Ohio. hlsroli. 1970.
IBtornturo and b~mhuresfrom several manufaelnrors were used ns reference m a t e h l . The nnthor wishes to t,hsnk these companies for t,heir goneroils supply of photogritphs and line drawings. The names and addresses of these manufacturers w e listed below. The anthor also wishes to acknowledge The Petralenm Research Fund (Grant No. 3516l3) and the National Science Foundation (Grant GP-8938) for partial support in the preparation of this article. American Instrument Company, Tne. 8030 Georgia Ave. Silver Spring, Md. 20910 Barher-Colmnn Company (now Nuclonr-Chicnm Cornoration) 2500 Harbor Blvd. Fullerton, Calif. 92634
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Canal Industrial Corporation (Canallcn) 563.5 Fisher Lane Rockville, Md. 20852 Cary Instruments, A Varian Subsidiary 2724 South Peck Rd. Monrovia, Calif. 91016 Chromstronix, Ine. 2743 Ninth St. Berkeley, Calif. 94710 Coleman Instruments Div. The Perkin-Elmer Corporation 42 W Madison St. Maywood, Ill. 60153 E. I. duPont de Nemours m d Company, Ino. Instrument and Equipment Division Wilmingt,on, Del. 19898 E C Apparatus Corporation 7.55 St. Marks St. Universit,y City, Philadelphia, Pa. 19104 Dr.-Ing. K:P. Hupe Appamtebau 75 Kmlsruhe 51 Lange Stresse 25, Germany (Ionies Research, he.-USA distributor) Instrumentation Specialties Company, Inc. 4700 Superior Lincoln, Neb. 68504 Ionics Research, Inc. (USA distributor for Dr. Hupe) P.O. Box 8412 Houston, Tea. 77004 Jeolco (U.S.A.) Inc. 477 Riverside Ave. Medford, Mess. 02155 Laboratory Data Control (LDC) 42 Shelter Rock Rd. Danbury, Conn. 06810
LKB Instruments, Inc. 12221 Parklawn Dr. Rockville, Md. 20852 Nester/Faust Manufacturing Corporat,ian 2401 Ogletown Rd. Newark, Del. 19711 Nuclear-Chicago Corporation (Barber-Colman) 333 East Howard Ave. Des Plaines, Ill. 60018 Philips Electronic Instruments (Pye Unicam) 750 South Fulton Ave. Mount Vernon, N. Y. 10550 Phoenix Precision Instrument Company, Inc. 3803-05 North Fifth St. Philadelphia, Pa. 19140 Pye Unieam Ltd. (Philips Electronic Instruments) York St. Cambridge CB1 2PX, England Schoefiel Instrument Corporation 24 Booker St. Westwood, N. J. 0767.5 Technicon Corporation Tarrytown, N. Y. 10591 G. K. Turner, Associates 2524 Puleas Ave. Palo Alt; Calif. 94303 Varian Aerograph 2700 Mitchell Dr. Walnut Creek, Calif. 94598 Waters Associates, Inc. 61 Fountain St. Framingham, Mass. 01701 Carl Zeiss. Inc. 444 Fifth Ave. New York, N. Y. 10018
