Adaptation of Photomultiplier Photometer to Beckman DU Spectrophotometer H. B. COLLIER A ND R. P. BARSCEIEL Department o.f Biochemistry, Univensity oSAlberta, Edmonton, Alta., Canada
I N ORDER to gain increased sensitivity and to permit the use - of narrower slit widths in flame photometry and reflectance measurements, the authors have adapted the Model 520M lineoperated photomultiplier photometer of the Photovolt Carp., 95 Madison Ave., New York 16, N. Y , to the Beckman Model DU spectrophotometer The Photovolt unit replaces the phototubes, amplifier, and meter of the Beckman, which is used simply as a monochromator with variable slit
transmittsncy readings on six solutions of sodium chromate are recorded. The average deviation of replicate readings is about 0.1% in transmittancy units. The linearity of response of the Photovolt unit is also illustrated in Table I. Various dilutions were made from a stock solution of reagent grade sodium chmmate, all in 0.05 M sodium hydroxide, and replicate transmittaney readings were made at 373 mp with search unit C (RCA photomultiplier tube lF'ZI), range X 100, and slit width 0.04 mm. For the various solutions the deviations
include the volumetric errors in making the dilutions. The gain in sensitivity that can be achieved in transmittmce and reflectance measurements is indicated in Table 11. This table gives a comparison of the slit widths required to give 100% readings on the transmittilnce scales of the two photometers when the tungsten lamp is used as light sonroe. The transmittanoemadingsweremaderr-ithn.aterina I-. Corexruvette and reflectance readings with a white Vitrolite standard. As the reflectance measurements were made with both the Beckman and Photovolt photometers adjusted for maximum response, these measurements can be used to make a direct comparison of sensitivity. It may be assumed that the amount of light striking the phototubes is roughly proportional to the square of the slit width; on this basis the increased photometric response for the Photovolt unit varies from shout 5X a t 700 m~ to 138X a t 600 mr. The results of flame photometry of B serum standad, diluted 1 to 12.5, in the Beckman model 10300 flame photometer attachment mith a propane-oxygen flame are given in Table 111. The same solutions were used in both photometers, and We slit widths
Figure 1. Attachment of Photovolt Search Unit to Beckman Phototube Housing MQ 0.
The chief advantages of this arrangement are the increased sensitivity and the greater range of light intensities that can be covered with the Soupdecade switch. Also, the line-operated amplifier uses standard radio tubes; serious delays have been experienced in replacing Beckman amplifier tubes. Minor conveniences are the absence of dry batteries and the ability t o cover the vhole visible spectrum with one phototube The Photovolt search units are adapted to the Beckman as f0llOW~S. The end plate of the Beckman phototube housing is taken off and the phototubes and amplifier tubes are removed. The end plate is then replaced by a plate which has a circular opening 1.5 inches in diameter to accommodate the window of the search unit. This opening is located directly oppopite the Beckman aperture and shutter. Upon the plate is mounted a clamp which holds the search unit in position and facilitates rapid changing of search units for various spectral regions. An annular rubber gasket is cemented around the opening in the plate t o prevent light leakage. The arrangement is illustrated in Figure 1.
0. 0,"""
I",
0.065 0.100 0.150
""
,
".
0.304 0.478
332: 33': 333: 338: 335 la?, igz, 191, 193,I W
i6ii
4730
0.707
M~ean
4710 4690
....b "Absorbancy ."..""".l.."..-".ll"""_l.". l = IozIdI. h b l e 11. SJit Widths for 100% Meter Readings in Transmittance and Reflectance Measurements, with Tungsten r T 4"L* E-.....,.. "D
Photovoltb Slit width.
Beckman' Wave Length, MP
Phototube
A.
400
B!?
Slit width. mm. Trsnsmittance 0.051 0.026
Readings on the Beckman photometer can he estimated to 0.1% Y""
Y l Y C
700
Red
RSlW
mm.
xXI00 10
0.014 0.018
0.030
XI00 XI0
0.60
XI
0.35 0.60 0.40
x1
0.050
on the transmittance scale. This is not possible with the Photovolt meter because of the shorter scale and the lanceshaped pointer. However, the tip of the microammeter pointer is bent through 90' to give a knife-edge, and in this way the readings can be estimated to 025% on the transmittance scale. The precision of photometric readings is indicated in Table I, where 30
".-I_.
,1", " 1
493 492 491 493 491
XI x1
0.018 0.015
0.155 0.052 0.051 0.185
For transmittance: range 1.0, 2000-megohm resimtor, a-itivity control fully counterdockarise. for reEecta?oe; !awe 0.1.l0.000-megohmjesistor sensitivity control i .. b Photomultiplier tube C
1030
V O L U M E 2 4 , NO. 6, J U N E 1 9 5 2
1031
Table 111. Slit Widths for 50% Readings in Flame Photometry of Serum Standard“ Element Na K Ca
Concentration, Me./L. 11.5 4.08 0.40
Slit Width, Mm. Beckmana Photovoltc 0.128 0.018 0.174 0.084 0.80 0.081
Wave Length,
M@ 388 766 554
a Beckman model 10300 flame photometer: air pressure 20 Ib./sq. inch, ox gen 16 inches of water, propane 1 cm. of isopropyl alcohol. f Range 0.1, 10,000-me ohm resistor, sensitivity control 3 turns from clockwise limit; blue t u b e for N a and Ca, red tube for K. 0 For X a : tube 1P22, range X 10; for K: tube 1P22, range X 1: for Ca: t u b e 1P21. range X 1.
.
..
__
.
-
required to give 50% readings on the transmittance scales of the photometers were determined. Only a slight increase in photometric response for potassium was achieved, even with the infrared-sensitive search unit F (RCA photomultiplier tube lP22). The wave lengths of the potassium lines a t about 768 mp are, unfortunately, near the limits of the spectral response curves for both the 1P21 and 1P22 tubes. The gain in sensitivity cannot be estimated directly from the slit widths recorded in Table 111. Gilbert, Hawes, and Beckman ( 1 ) state that in flame photometry the line energy passed by the monochromator is proportional to the first power of the slit width. However, with the authors’ instrument and under their conditions, the energy reaching the phototube (corrected for flame background) was found to be approximately proportional
to the second power of the slit width. The relationship varied somewhat with the element and its concentration. Dilution of the serum standard solution to give the same photometric response in both photometers (same slit width and same meter reading) indicated increases in response of about fivefold for potassium and one hundredfold for calcium. -4 several hundredfold increase for sodium has been observed repeatedly, but no satisfactory explanation for this can be offered. In flame photometry of sodium, potassium, and calciuni with the Photovolt photometer it is possible to use the following dilutions: serum 1to 50, urine 1 to 100, red cells 1to 200. At these dilutions the solutions do not require deproteinization and can be qrayed directly into the flame. There is no mutual interference of potassium on the determination of sodium in serum or of sodium on the determination of potassium in red-cell hemolysates. Positive mutual interference is observed in the following: potassium on sodium in red cells; sodium on potassium in serum and urine; sodium on calcium in serum. The precision and accuracy of these flame photometric determinations, and the effect of various other factors, are now under investigation and will be the subject of a further communication. LITERATURE CITED
(1) Gilbert, P. T., Jr., Hawes, R. C., and Beckman, 1, O., A 1 ~ ~ ~ .
CHEY.,22,772 (1950). RECEIVEDfor review J u n e 2 5 , 1951. Accepted Decenibei 20, 1951. b y a grant from t h e Yational Research Council of Canada.
Aided
Homogeneous Precipitation of Barium Sulfate by Hydrolysis of Sulfamic Acid W. F. WAGNER AND J. A. WUELLNER Unit-ersityof h’entucky, Lexington, h7y.
H E slow hydrolysis of sulfaniic arid to furnish sulfate ion is a Tbasis for the determination of barium as barium sulfate by homogeneous precipitation. Oberhauser and Urbina ( 3 )reported that a normal solution of sulfamic acid a t 96-97’ C. was 24% hydrolyzed after 15 minutes. The advantages of homogeneous precipitation, as pointed out by Willard and coworkers ( 4 ) are especially desirable for the formation of barium sulfate precipitates. Homogeneous precipitation yields a coarse crystalline precipitate that contains fewer coprecipitated impurities, requires little or no digestion, filters readily, and is easily washed free of adsorbed ions. Willard ( 4 ) reported the separation of barium from calcium by the hydrolysis of sulfamic acid. The separation of barium, strontium, and calcium by homogeneous precipitation methods using the hydrolysis of dimethyl sulfate in methanolwater solutions has been reported by Elving and Van Atta (8). The use of sulfamic acid might be preferable from the standpoint of cost and simplicity of procedure, by avoiding mixed solvents. CHEMICALS AND SOLUTIONS
The sulfamic acid used in the majority of experiments was Eastman Kodak practical grade or Du Pont technical grade. Results, using these grades of reagent, mere compared to results using a sample of the practical grade, which was purified according to the procedure given by Butler, Smith, and Audrieth (I), and a sample of rea ent grade sulfamic arid obtained from the G. Frederick Smith C%emical Co. The results were comparable regardless of the grade of sulfamic acid em loyed. The standard solutions of barium ch1orid)e were made from the anhydrous salt, which was frepared by heating bIerck reagent grade barium chloride dihy rate to constant weight. The solutions containing the cations used to study possible interferences were prepared by dissolving accurately weighed amounts of the C.P. chlorides in distilled water. The solutions of phosphoric acid and nitric acid were prepared by diluting accurately measured portions of the concentrated C.P. acids.
EXPERIMENTAL
At room temperature, barium sulfate slowly precipitates from a solution containing barium ion and sulfamic acid. On the steam
Table 1. Determination of Barium Ba Taken, h f g . 171.2
171.2
?io. of Detns. BR Found, N g . Sulfamic Acid Procedure 15 171.2 Standard deviation 0.38 Standard Sulfuric Acid Procedure 3 171.4 Standard deviation 0.13
Table 11. Determination of Barium in Presence of Foreign Ions Ions Added
Barium
Mg.
MU.
.vg.
None None Xone None None 500 O c a ’ + 200 O c a + + 100 O c a + ’ 20 O c a + ’ 20 O c a + + 40 O c a + + 40 O c a + + 20 o c a + + 500 0 Ca;;+ 500 O F e 500 O F e + + 200 O F e T a i 100 O F e + + 7 100 O F e + + + 40 O F e + + + 100 0 M g + + 40 O M g + +
100.0 100.0 25.0 10.0 10.0 100.0 100 0 100.0 100.0 100.0
-0.3 0.0 -0.1 -0.3 -0.1
10.0
10.0 10.0 1.0 100.0 100.0 100.0 100.0 10.0 10.0
100.0 100.0
Error
f0.6
4-0.3 +0.2 -0.6
4-0.3 f0.6 -0.3
-0.2
-0.2 +0.7 +0.5 -0.3 +o. 1 -0.2 -0.1 +0.; -0.2
Ions Added
Bariuin
Error
Mg.
Mg.
Mg.
40 10 500 200 100 40 100 50 100 40 20
100 40 40 10
5 1
10
10
1 10
0 0 0 0 0 0 0
Mg++
Mg’+ HsPOa HaPo? HaPOi Hipoi HNOj 0 “0s 0 “08 0 HNOa 0 HNOi 0 NaNOa 0 NaNOa 0 Nay03 0 Sr+’ 0 Sr++ 0 Sr++ 0 Sr+T 0 Sr++ 0 Sr++ 0 Sr++
10 0 1 0
fO.1 +I7 5
