P h o t o c e l l C i r c u i t . Only one photoelectric cell is used and it is the p h o to v o lt a ic type which acts as a source of current without the aid of an external e. m. f. The fundamental characteristic of this type of cell is the current, which is almost exactly pro
portional to the light intensity for low
r e s is ta n c e s in the external circuit, Fi g u r e 1 . Ph o t o e l e c t r i c Co l o r i m e t e r
rather than the e. m. f. as is usual in the ordinary sources of elec
trical energy. The cell used here is a photronic cell, Model 594, manufactured by the Weston Electrical Instrument Corp. (1).
I t is connected in series with a microammeter (MA, Figure 2) hav
ing 50 ohms resistance and reading up to 50 microamperes. A special scale permits an accurate estimation to 0.1 microampere.
L i g h t C i r c u i t . The source of energy is a 6-volt, 17-plate lead storage battery and the lamp used is a 6- to 8-volt, single
filament auto headlight bulb. The lamp, b, is connected to the battery terminals through a pair of resistances, Ri and /i2, in parallel, one for coarse and the other for fine adjustment. Across the lamp is connected a small voltmeter, V, reading up to 8 volts, for approximate adjustment of the resistances.
O p t i c a l S y s t e m . Below the lamp is placed a spherical metal reflector, a, with the filament a t its center of curvature, so th a t light rays are reflected back upon themselves to approximately double the intensity of the beam traveling upward through diaphragm, c, which defines the beam.
Above the filament a lens, d—diame
ter, ‘2.5 cm.; focal length, 5 cm.—is placed a t its focal distance, so that the beam striking it is rendered very nearly parallel. Another diaphragm, /, limits the parallel beam before it enters the tube containing the liquid.
The unabsorbed light then strikes the surface of the photocell exposed by the aperture of diaphragm, g. The beam is rigidly defined in order to eliminate as far as possible errors due to stray reflections from the sides of the tubes, which are never entirely regular; condensing lens effect of drops a t the tops of the tubes; re
flections from finger prints on the out
side of the tubes; and the diverging lens effect of the meniscus, an error which is here reduced to a minimum by having the diameter of the beam small in comparison with the diameter of the meniscus.
' - T u b e s . The tubes (Figure 3) are made from precision-bore tubing of clear glass and have optically plane, fused-on bottoms to prevent distortion of the beam. The inside height to the graduation mark is 100 mm. and the volume is 100 ml. These tubes can be made so th at the volume variation is a negligible fraction of a milliliter (100.04 ml. is the volume of each of the two tubes at hand). This provides a very convenient volume for ordinary work.
M e c h a n i c a l C o n s t r u c t i o n . On the bottom of each tube is cemented a brass ring into which a slot has been cut to coincide with a pin, so that the tube is always held in the same position.
A turntable, mounted on smooth bearings, carries the two tubes and successively rotates them into position in the light path.
A ratchet device with the two fixed points insures the correct position of the tube for a reading. At the base of the cylinder containing the turntable, a convenient slide permits the insertion of color filters just above the lens (e, Figure 2). When a filter is not used, a clear optical flat is kept in the slide to protect the lens and to keep out dust. A sliding door on the side of the cylinder opens to permit insertion of the tubes and when closed keeps out stray light and dust.
The lens, lamp, and reflector are all rigidly mounted in a light- alloy casting. The lamp socket is adjustable to permit accurate centering of the filament with respect to the lens, and the reflector can be focused by means of a screw, thus permitting realignment of the beam should a bulb have to be replaced.
A pair of adapters (Figure 3) can be mounted on the turntable, suspended from the shoulders th at hold the tops of the large tubes in fixed position. The adapters hold microtubes (5 ml.) so that the instrument can be used in cases where only very small volumes of liquid are available, as is often the case in biochemical and biological work.
O peration
The first and a very im portant step is cleaning the tubes and securing a photom etric balance. The tubes are first cleaned inside with sulfuric acid-dichrom ate cleaning solution and then rinsed very thoroughly w ith distilled water. The bottom s are occasionally washed with a soap solution free from abrasive, b u t ordinarily polishing them w ith lens paper is sufficient.
The tubes are filled to the mark with distilled water and then placed in position in the instrument. The sliding panel is closed and the light switch turned on. The coarse resistance is adjusted so that the voltage reading is approximately 3 volts. The instru
ment must then be allowed to stand for 2 or 3 minutes to permit the filament and the rheostats to warm up so that their resistance becomes fairly constant. Then the photocell switch is turned on and the intensity of the light adjusted by means of the coarse and fine resistances so that the microammeter reads 50 with tube I in position. Tube II is then quickly turned into position. The reading should be 50, when the needle has come to rest. It is necessary to turn I back into position to make certain th a t the reading is still 50. The first reading is the least reliable and in this work it was discarded and the mean of three readings on
tube II, between which the readings on I did not vary from 50, was taken as the correct value. If an exact balance is not a t
tained, for the sake of simplicity in handling the data, it is better to secure it by again polishing the bottoms of the tubes. The same procedure is followed in taking the reading of a colored solution in tube II. Tube I is filled with the blank solution and the microammeter always set a t 50. By having the blank solu
tion contain everything except the color-producing substance, errors due to the absorption of light by other substances in solu
tion are automatically eliminated. Setting the light intensity to a definite value with the blank makes it unnecessary to have a sensitive voltmeter, since an exact knowledge of the voltage ap
plied is not required.
The photronic cell has an inherent objectionable defect which may be classified as a “ fatigue" effect, and can produce an appreciable error unless th e proper care is observed in operation. W hen the cell is first exposed to light its reading is high, b u t after a very short tim e it drops to a constant value. As long as th e solution is dilute and transm its alm ost as much light as th e blank, the fatigue effect is negligible, b u t when the solution is concentrated, it is necessary to w ait for about a m inute before taking the reading of the blank. This delay m ay allow the voltage across the lam p to change, since th e b attery is continuously discharging and the reading m ay be in error b y as much as 1 yua. This voltage change m ay be reduced greatly by using a b attery with a high am pere-hour capacity or by using two batteries connected in parallel. However, for work of the highest accuracy, it is necessary to determ ine th e approxim ate concentration of the unknown and then m ake u p a standard of th a t concentration.
Setting the light intensity to th e correct reading of the stan d ard, th e reading of the unknown is taken and the relationship between the readings and th e concentrations is given by Equation 5.
Fi g u r e 3 . Tu b e s a n d Ad a p t e r s
In case solutions to be studied m ust be m ade up with a colored reagent or contain colored substances which do not interfere by chemical reaction, color filters m ay be used to eliminate their absorption. If a color filter having th e same spectrophotometric transmission curve as th e interfering colored substance be inserted in the light path and the blank set to a reading of 50 jua as before, th e absorption observed will be due almost entirely to th e substance whose concentra
tion is to be measured. I t is thus unnecessary to know the concentration of the interfering colored substance so long as it remains below the lim it of the depth of color of th e filter.
When the microcells are used, the reading of the blank is set a t 25 rather than 50 because a sm aller area of the photronic cell is exposed and sufficient intensity to cause full scale de
flection would discharge the b attery so rapidly th a t constancy of the blank reading could be m aintained only with great difficulty.
JULY 15, 1935 ANALYTICAL EDITION 283 C a lib r a tio n
T he instrum ent m ust be calibrated for the q u antitative determ ination of a substance by means of a given color reac
tion. Two m ethods m ay be used for handling the data, the choice depending upon the accuracy dem anded and the tim e available. T he first, and simpler, m ethod is a plot of am m eter readings against concentrations (Curve I, Figure 4) or a curve of th e decrease in am m eter readings against concentrations (Curve II, Figure 4) gives a positive slope
Fi g u r e 4 . Ca l i b r a t i o n Cu r v e s
and m ay be preferable. In either case, th e concentrations m ay be read directly from the graph. Using this m ethod, however, precludes th e possibility of eliminating the error due to th e fatigue effect m entioned previously. The second m ethod of handling th e d a ta is based on two assumptions:
th e validity of B eer’s law and proportionality of current production to light intensity.
Beer’s law in its m ost familiar form is:
U = /o-10-“ « (1)
taking logarithms
log I c — log / o — acx
Assuming direct proportionality of current to light intensity,
R = K I (2)
where R is the ammeter reading. Since the depth of solution x is constant (100 mm.) a and x may be combined into another con
stant k
log - kc (3)
l i e
A plot of log R<,/Rc against c should give a straight line whose slope is k. T h a t such is th e case, a t least w ithin cer
tain lim its, can be seen from Figure 5. Plotting the curve and evaluating th e slope m akes it possible to calculate the concentration from the relation:
1 . 50 ...
C = k g R ^
From this th e relationship between two concentrations and their readings follows:
Ci = log 50/fli
c, log 5 0 /f t w
A table of the values of log 50/R for values of R from 50 to 10 a t intervals of one-tenth was m ade up and much tim e saved in the calculations.
The first step in studying a given color reaction is to deter
mine w hat the blank solution m ust contain. I t is advisable to choose a fixed am ount of th e color-developing reagent, or reagents, of sufficient q u antity to give a large excess for all concentrations th a t will be encountered. If the excess be very large, the am ount used up in th e reaction will be neg
ligible in comparison and th e variation in percentage excess will be small, so th a t the system will more closely approxi
m ate Beer’s law conditions. If the blank solution is found to show a measurable absorption, it m ust be used as the blank in tube I, b u t if it showrs no absorption or a filter can be found which will m atch its absorption, distilled w ater m ay be used in tube I and thereby greatly simplify th e procedure.
The colored solution on which th e m ost extensive investi
gation was carried out was ammoniacal cupric sulfate [Cu- (NH3)1SOt ]. I t was chosen because it is a relatively poor colorimetric system for visual study and hence would give the instrum ent as rigid a test as possible and still remain within the limits where comparison w ith visual methods could be made. Ammonia solutions exhibit an appreciable absorption in the visible region; hence it was necessary to use as a blank an ammonia solution of the same strength as th e standards in w’hich an excess of very large magnitude was used. Less extensive studies on two other systems gave comparable results, so th a t the results which follow m ay be considered as typical, b u t it m ust be remembered th a t m any colorimetric reactions give solutions of the order of one hundred times as intense a coloration a t a given concentra
tion and the sensitivity to concentration change is there
fore of the order of one hundred times as great.
R esults
The stock copper solution was prepared from reagent grade CuSOvSHjO by dissolving 39.282 gram s in a liter of solution a t 20° C. Then 100 ml. of this solution were diluted to a liter to give a solution which contained 1 mg. of copper per ml.
Fi g u r e 5 . Ca l i b r a t i o n Cu r v e s
This was then checked by electrolytic deposition and was found to be exact. The te st solutions were m ade u p over a range of 5 to 100 p. p. m. of Cu * the proper num ber of milli
liters of th e C u +'r solution (1 mg. per ml.) being added to 25 ml. of 15 M am monium hydroxide and diluting to a liter.
T hus in th e m ost dilute solution th e am ount of am monia present wras 1200 times th a t required for the reaction
Cu++ + 4 NH, - Cu(NH„),++
and in th e m ost concentrated, 60 tim es the theoretical am ount. The blank was m ade by diluting 25 ml. of th e same ammonium hydroxide solution to a liter.
Following th e procedure outlined under “ operation,” th e following results were obtained w ith the C u(N H 3)1++ solu
tions of concentrations as indicated.
Ta b l e I C o p p e r,
p . p . m . 1 2 - R
---3 M e a n 50-R lo g 50/ R
5 4 6 .8 4 6 .8 4 6 .8 4 6 .8 3 . 2 0 .0 2 9
10 4 3 .8 4 3 .7 4 3 .7 4 3 .7 6 . 3 0 .0 5 9
15 4 0 .9 4 0 .9 4 0 .9 4 0 .9 9 .1 0 .0 8 7
20 3 8 .0 3 8 .0 3 8 .0 3 8 .0 1 2 .0 0 .1 1 9
25 3 5 .4 3 5 .3 3 5 .3 3 5 .3 1 4 .7 0 .1 5 1
35 3 0 .8 3 0 .8 3 0 .8 3 0 .8 1 9 .2 0 .2 1 0
50 2 5 .1 2 5 .2 2 5 .1 2 5 .1 2 4 .9 0 .2 9 9
75 1 8 .1 1 8 .1 1 8 .1 1 8 .1 3 1 .9 0 .4 4 1
100 1 3 .8 1 3 .8 1 3 .8 1 3 .8 3 6 .2 0 .5 5 9
INDUSTRIAL AND ENGINEERING CHEMISTRY These d ata are plotted in Figures 4 and 5. From Figure
5 it can be seen th a t the linear relationship predicted by Equation 3 holds over a fairly wide range of concentrations, the value of k being 0.00599.
In repeating the above with two new sets of solutions, the values of k obtained were 0.00603 and 0.00597, respec
tively. A reliable average of these values is k = 0.0060.
A solution, whose concentration was unknown to th e ob
server, was placed in tube II and gave a reading of 44.7 jua.
From the curves of Figure 4, this means a concentration of 8.2 p. p. m. Or, the following calculation gives
log 50/44.7 0.0486
0.0060 0.0060 8.1 p. p. m.
The correct concentration was 8.0 p. p. m. In comparison with visual methods, by means of Xessler tubes in a roulette com parator (2), a solution containing 8 p. p. m. can be dis
tinguished only with difficulty from 7 or 9 p. p. m., and with certainty from 6 or 10 p. p. m.
Acknowled gm en ts
The authors are indebted to Frederick L. Brown of the Rouss Physical Laboratory for m any helpful suggestions and to Robert H. K ean of this laboratory for assistance in stu d y ing some of the early models of the colorimeter. Our thanks are due th e American Instrum ent Company, Washington, D. C., for building the apparatus and for assistance in solving some of th e problems of design.
L ite ra tu re Cited
(1) W eston E le ctrical I n s tru m e n t C o rp ., N e w a rk , X , J ., T ech n ical D a ta , B-1001-A.
(2 ) Y o e a n d C ru m p ler, I n d . E.vg. C h e m . , A nal. E d ., 7 , 7 S (1935).
R r.c E iT c o A p ril 16, 1935.