about 100 pg. of carbon and were measured with a precision of =t11.4%. All samples weighed between 250 and 400 mg. Included in the above errors are those inherent in detection of the carbon dioxide produced. One major source of error is caused by uncertainty in the COz background rate over the required 15-minute collection time. Normally in
bench-top operation a sample can be analyzed in 5 to 6 minutes, but in this case the furnace is 25 feet from the analyzer. These errors can be reduced, particularly for bench-top application, by burning larger samples and minimizing the length of the gas line between the furnace and analyzer. However, it is considered sufficient proof that this furnace is directly applicable for the
combustion of steels and refractory metals for carbon or sulfur analysis. ACKNOWLEDGMENT
The author thanks Frank F. Felber, Jr., for his guidance in the project, and W. E. Hatch and R. A. Norton for their help in the mechanical development. Work performed for the U. S. Atomic Energy Commission under contract No. AT(30-1)-2789.
A Gas Sampling and Injection Valve for Vacuum Service Martin Alperstein and Ronald STUDY
1. Bradow, Texaco Research Center, Beacon, N. Y.
of slow combustion reactions
@ NUT
A in the end gas of an Otto-cycle engine in progress in this laboratory uses a unique gas-chromatograph column introduction valve. In one phase of the experimental program hot, reacting gases are withdrawn from the engine through a special valve (1) and are rapidly quenched by expansion into a low-pressure receiver. The quenched gases a t pressures less than 40 torr are qualitatively and quantitatively analyzed by flame-ionization gas chromatography. The analysis system requires a gas handling apparatus that would maintain sample integrity while introducing the sample into a flowing carrier gas stream a t pressures up to 100 p s i Specific requirements for such a gas sampling valve include: capability of leak free operation with collection torr; low volume pressures of internal pressure drops; leak-free carrier gas circuit; no exposure of sample t o the atmosphere during switching; no materials which absorb sample or outgas to contaminate sample. Spool or linear valves are unsatisfactory because of their high pressure drops. When ports are increased in diameter, damage of the highly loaded edge seals occurs ( 7 ) . Rotary indexed face-sealed valves (4,6,8), which possess the advantage of large ports and variable loading of the seals, are commonly constructed with hard seals which either do not effectively seal the pressure extremes or quickly deteriorate on repeated switching. Systems of glass (2, 6) or metal stopcocks (3) either expose the sample to stopcock lubricant or do not have sufficiently rapid injection characteristics to be used with low-flow, high-resolution columns. Ideally, a valve meeting the system requirements should have the injection and variable seat loading characteristics of a rotary indexing valve and the low pressure sealing capability of a spool valve. A valve was designed and constructed using internal flow paths similar to the rotary valves but using easily replaced, 366
ANALYTICAL CHEMISTRY
A W E PORT PLATE
SEAL (6) TWO CONCENTRIC O-RINGS
INDEXING GUIDE
SURROUNDING O-RING GROOV
Figure 1.
VACUUM PORT
Low-pressure service six-port sampling valve
inexpensive O-rings as the face seals. The figure illustrates a typical design with this unique sealing configuration. The isometric assembly sketch shows a six-port valve; however, other porting arrangements should be equally feasible. As constructed, the smallest port passage used has a 1/16-inchdiameter and the valves were machined from 304 stainless steel. Each port seal consists of two concentric O-rings ( W o n A series 2-10 outer and series 2-5 inner) held in 0.047-inch deep recesses machined in the valve body. In assembly, the valve port plate slides easily over the valve stem, and is indexed t o the valve body so that all ports align a t valve motion extremes (in this case 60" movement). The valve port plate is loaded toward the valve body by variable spring tension so that the O-rings are squeezed approximately 0.009 inch. This deformation is sufficient to seal over the required pressure ranges, yet permits handoperated orientation of port positions.
In construction, it is not necessary to lap the valve port plate face; tolerances of 3tO.001 in. and =t are maintained. The port edges of the valve port plate are rounded approximately 0.005 inch to prevent O-ring cutting. The valve port plate is self leveling relative to the valve body and equalizes loadings on all O-rings. The valve illustrated is suitable for analyses of air components because the port seals could be surrounded by a vacuum environment formed by series 2-33 outer and series 2-15 inner O-rings. Such arrangements prevent dilution of sample by entrapped air in the valve port plate passages upon flow path orientation. This precaution is unnecessary for other analyses, and the surrounding O-rings and vacuum port are not provided in valves used for hydrocarbon analyses. The valve stem illustrated can be extended and keyed to the valve port plate for remote operation or immersion of valve body in a controlled-temperature air bath.
The described valves have been used satisfactorily for over two years in various analyses systems and have proven durable and dependable. Precision of repeated calibration gas sample analyses by flame ionization gas chromatography has been within 0.7%. No evidence of absorption or contamination by outgassing has been observed using either Viton A or silicone rubber O-ring seals. In addition to
satisfying the stated requirements, the valves are easy to disassemble for OCCaSiOnal cleaning Or for infrequent O-ring replacement.
LITERATURE CITED
(1) dperstein, M., Bradow, R. L., Rev. Sci. Instr. 36, 1028 (1965). (2) van de craats, F., Chim. A ~ 14, 136 (1956).
(3) Fredericks, E. M., Brooks, F. R., ANAL.CHEM.28, 297 (1956). (4) Hausdorf, H. H., “Vapor Phase Chromatography,” p. 377, Butterworths, London, 1957. (5) Hill, D. W., Hook, J. R., J. Sci. Instr. 37, 253 (1960). (6) Percival, W. C., ANAL.CHEM.29, 20 (1957). (7) Pratt, G. L., Purnell, J. H., ANAL. CHEM.32, 1213 (1960). (8) Watson, E. S., Bresky, D. R. (to ~ Perkin-Elmer ~ . Corp.), U. S. Patent 2,757,541 (Aug. 7, 1956).
Quinhydrone as Quantitative Standard for Electron Spin Resonance Spectrometry of Biological Systems Giuseppi Narni,
A
H. S.
Mason1, and lsao Yamazaki2 University of Oregon Medical School, Portland, Ore.
of substances such as peroxylamine disulfonate, diphenyl-picryl-hydrazyl, powdered coal, charred dextrose, copper sulfate, manganese sulfate, vanadyl sulfate, and synthetic ruby, have been used as standards of unpaired electron concentration in electron spin resonance spectrometry. None has proved altogether satisfactory for several reasons (2,s), and the general problem of determination of spin concentration by ESR for paramagnetic substances in solid, liquid, and gaseous states, through a broad temperature range, remains to be solved. I n a relatively narrow aspect of the problemdetermination of free radical concentration in aqueous solutions of neutral pH a t room temperature, as in the case of free radical generation from substrates by oxidative enzymes-it is essential that the unknown and standard solutions occupy identical space in the spectrometer cavity, that the free radical character of the unknown arise from a structure similar to that of the standard, that the dielectric constants of the standard and unknown solutions be similar, and that pH and ionic strength of the unknown be similar to that of the standard so that the replacement of one by the other in the sample cell by a flow method cannot produce residual changes. Solutions of peroxylamine disulfonate in mildly alkaline solution are useful as standards of spin concentration under these conditions because the compound is readily estimated by optical spectrometry; however, the peak-to-peak width of the ESR absorption lines is very narrow and difficult to integrate. I n this study, we have employed peroxylNUMBER
‘To whom inquiries should be addressed. * Present address, Departplent of Biophysics, Hokkaido University, Sapporo, Japan.
PEROXYAMINE DISULFONATE
p-SEMIOUINONE
Figure 1. Integrated signals of peroxylamine disulfonate in 0.01 N Na2C03, and quinhydrone in 0.2M phosphate buffer, pH 7.5; each solution approximately 5 m M
amine disulfonate as a primary standard, against which we have determined the concentration of p-benzosemiquinone in equilibrium with quinhydrone. This has given us an estimate of the reliability of quantitative ESR spectrometry. I n addition, the concentrations of p-semiquinone a t neutrality proved to be at convenient levels for ESR spectrometry of aqueous solutions, and this system appears to offer a readily available, easily prepared, and reasonably stable aqueous standard for enzymological work. EXPERIMENTAL
Apparatus. A Varian V-4500 xband ESR spectrometer was used with 100-kc. field modulation. The Varian quartz flat cell, 0.25 mm. internal thickness, containing 0.075 ml., was mounted in a fixed position in the cavity for optimal signal strength and maximum stability of the cavity and sample geometry. Sample solutions were placed in the cell by means of a syringe attached a t the bottom, and, with each replacement, the cell was very carefully flushed in
order to avoid contamination. All experiments were done with a field sweep speed of 4.3 gauss per minute, a t a field modulation amplitude of 0.2 gauss. The spectrometer response time was 0.3 second, and the full scale recorder response time was 1.0 second. Field strengths were monitored with a Varian F-8 fluxmeter and a Hewlett-Packard 524C electronic frequency counter and klystron frequencies with a HewlettPackard K532B frequency meter. Reagents. Potassium peroxylamine disulfonate prepared by the procedure of Harvey and Hollingshead ( I ) was twice recrystallized, dried with methanol, and stored in a freezer a t -2OO C. in sealed, evacuated tubes. The compound was standardized by iodometric titration, and used immediately for ESR measurements as a solution in 0.01N Na2C0a;only freshly standardized solutions were used for this purpose. Quinhydrone (Eastman Co.) was recrystallized to m.p. 170-1°, and finely ground in a mortar before making solutions. It was observed that oxygenfree buffer solutions gave rise to relatively stable concentrations of free radicals, and all solutions were accordingly prepared by shaking the powdered VOL. 38, NO. 2, FEBRUARY 1966
367
