INDUSTRIAL AND ENGINEERING CHEMISTRY
436
Summary The analytical method devised for estimating the atmospheric aromatic contents consists of cooling a blend of the unknown air sample and the vapor of a fixed volume of methyl alcohol on one hand, and blends of air samples containing known concentrations of the aromatic distillates in question and the vapors of a fixed volume of methyl alcohol on the other, to -56.67" C. (-70" F.) and obtaining refractive indices of the condensates. This method is reasonably simple and gives a n accuracy
VOL. 12, NO. 7
within 15 parts per million for critical concentrations of 100 parts per million.
Acknowledgment The authors extend thanks to the Humble Oil and Refining Company for permission to publish their findings.
Literature Cited National Safety Council, 1926. (2) Yant, IT.P., Pearce, S. J., and Schrenk,H. H., U.S.Bur. Mines, R e p t . Inwstigations 3323, 1936. (1) Benzol Report,
Microscope Hot Stage for Determination of Melting Points Application to Carotenoid Pigments F. P. ZSCEIEILE
AND
J. W. WHITE, JR., Purdue University, Lafayette, Ind.
T
HE usual method of determining melting points by means
of aBerl block or a liquid bath surrounding a melting point tube does not give satisfactory results when applied to such compounds as the carotenoid pigments. The melting point of these compounds observed in this manner is really an indication of decomposition; it is not a sharply defined point and considerable material is required. Values for the melting point of alpha-carotene reported in the literature vary from 178.5" (11) to 188" C. ( 8 ) , a range of 9.5" C. For betacarotene the reported values vary from 177.8" (11) to 187.5"C. (IO),a range of 9.7" C. The range of any one determination is usually 1" C. or more. The sintering point is usually observed slightly below the melting range. A number of hot stages for melting point determination hare been described ( 1 , 3 , 4 , 5 , 9 ) ,but the apparatus discussed here includes refinements which add greatly to the precision of measurement. T o the authors' knowledge, this technique has not been applied previously to carotenoid pigments, to which i t is very well adapted. I t s use should add significance to the melting point as a physical constant in carotenoid chemist'rg.
Description of Apparatus The hot stage is constructed as shown in Figure 1. The two copper plates are placed on the stage of a polarizing microscope, and the sample is placed on a circular cover glass in the upper recess of the lorver plate. A4spring clip holds the cover glass in place, so that the crystals are directly over the 0.156-cm. (0.0625inch) hole passing through the axis of the copper plates. The sample is thus illuminated from below Iyith polarized light and is observed through the analyzing S i c o l prism of the microscope. The stage is heated electrically by means of a coil of 60 cm. ( 2 feet) of S o . 21 Xichrome wire having a resistance of 3.1 ohms. The core, machined from lava and baked to hardness, is recessed in the bottom of the lower block around the central hole. An annulus of lava is cemented under the heating element to enclose it within the block. Control of the heating rate is effected by a Variac variable transformer, Type 200-C. Heating rates of from 0.1" to 6.5" C. per minute can be maintained at any temperature from that of the room t o 250' C. The temperature within the apparatus is measured by a Leeds & Xorthrup portable precision potentiometer and an iron-constantan thermocouple. The hot junction of the couple, about 1 mm. in diameter, is located within 1 or 2 mm. of the sample. The leads are brought diagonally through the lower plate in a porcelain tube, cemented in place. Two holes lead from the outside to the central chamber for the passage of nitrogen through the apparatus. Figure 2 shom how the hot stage is insulated from the microscope stage by a Pyrex support on an asbestos plate. A Leitz
Periplan 20-power ocular and a Bausch & Lomb 4-power objective with a 38-mm. working distance provide suitable magnification and permit the objective t o be sufficiently distant from the hot stage t o avoid heat damage. A camera lucida makes it possible to observe simultaneously both the sample being melted and the galvanometer pointer. At the left center of the photograph is the Variac transformer, with voltmeter. To the right of the microscope is the potentiometer box; at the extreme right is a Dewar flask containing the cold junction of the thermocouple. In front of the Variac is a bubble counter to measure the flow- of nitrogen. The precision pinchcock in the foreground regulates the rate of gas flow. A micro combustion tube containing heated copper for removal of traces of oxygen from the nitrogen and a drying tube containing Dehydrite are not shown. With this equipment, the thermocouple voltage can be estimated to 1 microvolt, which corresponds to 0.02" C. with an iron-constantan thermocouple.
Calibration of Apparatus A heating rate of 0.3" C. per minute was selected because it is slow enough to permit the sample and the thermocouple to attain the same temperature and i t is rapid enough to avoid an unduly long time for the determination. The thermocouple voltage may be kept balanced easily a t this heating rate. The voltage necessary to maintain a heating rate of 0.3" C. per minute at any desired temperature was determined by measurement of the block temperature increase with time for four voltages. Rates of heating were plotted against temperature for each voltage. The temperature a t which a given voltage heated the apparatus at the desired rate (0.3" C. per minute) was then plotted against that voltage. The result was a straight line from which can be read the voltage necessary to heat the block a t 0.3" C. per minute at any temperature from room temperature to 250" C. When a melting point is to be determined, the sample, which may be a single crystal if necessary, is placed in position and the apparatus is heated at 22.5 volts, corresponding to a current o,f 6 amperes, the upper limit of the heating element. Within 1.5 to 2.0° of the anticipated melting point, the voltage is set at the point necessary to maintain a heating rate of 0.3" per minute. The response to voltage change is sufficiently rapid t o allow maintaining solid and liquid phases as desired. Thus, if the sample does not decompose, several melting points may be determined with the same sample. Nitrogen should flow through the apparatus during the entire heating procedure. About 1.5 minutes before the expected melting point is reached, the flow of nitrogen is stopped to ensure temperature equilibrium. When anisotropic crystals are observed between crossed Kicol prisms, the melting point is observed as a sudden disappearance of
JULY 15, 1940
ANALYTICAL EDITION
double refraction. The time necessary for the complete determination is about 30 minutes.
TABLE I.
To test the reliability of the apparatus, melting points of purified samples of well-known compounds were determined covering the temperature range of 40" to 175" C. Results are reported in Table I. These determinations have a maximum difference of 0.13 " C. between successive crystallizations of the same material. The precision of successive determinations on fresh samples of these materials is *0.04" C. or less. I t is concluded that the melting points of these compounds, determined by this method, are not significantly different frcm those reported in the literature. Two determinations on fresh samples of potassium thiocyanate, heated a t different rates, 0.25" and 0.73" C. per minute in air, gave 175.31' and 175.29" C. Four months
437
Substance
CoMP.4RIsoS O F MELTISG POINTS
Reported Mplting Point
c.
p-Tolyl amine
42-43 (6) 4 3 . 7 (7)
p-Sitronniline
147.5 147.3 148.0 173 2
Potassium thiocyanate
(6) (2)
(7) (6.7)
Grade and Treatment Eastman, after treatment with bone black and recrystallization from ethanol After second recrystallisation from ethanol Eastman White Label After recrystallization from Pthanol
Observed0 Melting Point
c.
42.95 42.87
42.89 42.82 147.20 147.22
175.38 175.31 175.39
a Each observed melting point is ior a fresh sample in air. .ill samples were dried In a vacuum of 1.5 microns of mercury (measured by a Pirani gage) ior 4 hours a t room temperature.
INDUSTRIAL AND ENGINEERING CHEMISTRY
438
VOL. 12, NO. 7
.4 sample of beta-carotene was recrystallized four times from carbon disulfide-ethanol and dried 20 hours at 84" C. in vacuum. Three melting point determinations, made under nitrogen, were asfollows: 179.78", 179.81", and 179.81" C. In air, this sample melted at 178.51' C,.. 1.3" C. lower than under nitrogen. When oxygen was passed through the apparatus during a melting point determination, this sample melted at 168.63' C., 11.2' C. lower than under nitrogen. A sample of alpha-carotene, four times recrystallized from carbon disulfide-et,hanol and dried at room temperature in a vacuum of 10 microns of mercury (measured by a Pirani gage), melted at 184.43' C. under nitrogen. After 3 weeks' storage at 0" C. in vacuum, it melted at 184.37" C. under the same conditions. In air, this sample melted at 180.73" C., or 3.7" lower than under nitroge?.? Under oxygen, it melted at 172.78" C., or 11.6 C. loiver than under nitrogen.
The authors have never observed a recrystallization of either alpha- or betacarotene from the melt after fusion, following a reduction in temperature. Visual observations on the color of the fusion product' indicate t'hat oxidation occurs in the presence of oxygen. Under nitrogen, no visible bleaching occurs for many hours after melting; in oxygen the melt is bleached colorless within a few minutes after fusion; and in air an intermediate amount of bleaching occurs. However, chemical change occurs, even when fused under pure nitrogen, as shown by the fact that the absorption maxipa of beta-carotene in the visible region were shifted 100 A. toward shorter rrave lengths. Severtheless, the melting point of separate samples of beta-carotene as observed under nitrogen by this method, even though accompanied by chemical change, may be determined within the precision of the instrument, +O.0Zo C.
FIGURE 2. MELTING POIXT APPARATUS
Summary A microscope hot stage is described, with which it is pos-
lo
1
I
-
i
9-
1
'
VFROM OLTAG 2 E2 5R JTDOU C1E4D5'1
-180
.....,..............'.........,......*.*.*......... T I T
-
LIP
UP
M P
sible to determine the melting point of a single crystal as a sharply defined physical constant of high precision. The crystal is surrounded by an inert atmosphere and the heating rate is carefully controlled. The melting point may be observed as a sharp change in t'ransmission of polarized light. The temperature range available is from room temperature to 250' C. The precision of the measurement is +0.02" C. lIelt,ing points of known standards were determined with a precision of =t0.04" C. This or higher precision has been attained wit,h alpha- and beta-carotenes.
Literature Cited -176.0°C,,
11
9.440
20
-1755
-
-175.0
-loo
-
-174.5
-
74.0
-
-20
I 1
FIGCIAE 3.
1
l
I
l
I
h l E L T I S G P O I X T OF
I
I
I
l
I
POTASSIUM T H I O C Y . 4 X A T E
n Q
K
0 I-
z
w
cn W
0
w
h m d u r , I., a n d H j o r t , E. V., ISD.ENG.CHmi., Anal. Ed., 2, 259 (1930). "Chemical Engineers' Handbook", New York, McGraw-Hill Book Co., 1934. Clevenger, J. F., IND.EKG.CHEW,16, 854 (1924). D u n b a r , R . E., I b i d . , Anal. E d . , 11, 516 (1939). F u c h s , L., Mllikrochim. A c t a , 2, 317 (1937). H o d g m a n , C. D., "Handbook of Chemistry a n d Physics", 22nd ed., Cleveland, Ohio, Chemical R u b b e r Publishing Co., 1937. I n t e r n a t i o n a l Critical Tables, Vol. I, New York, hlcGraw-Hill Book Co., 1926. K a r r e r . P.. a n d Walker, O., Helv. Chim. Acta, 16, 611 (1933). Kofler, L., a n d Hilbck, H . , MikTochemie, 9, 38 (1931); 15, 242 (1934). Miller, E. S . , Botan. Gaz., 96, 447 (1935). Shrewsbury, C . L . , Kraybill, H . R., a n d Withrow, R . B., IND. ESG. C H E X . , ! m a l . Ed., 10,253 (1938). Strain, H . H . , "Leaf Xanthophylls", p. 45, Washington, D. C., Carnegie Institution of Washington, 1938. PRESENTED before the Division of Biological Chemistry a t the 98th Meeting of t h e American Chemical Society, Boston, Mass. Based on a section of a thesis submitted t o the faculty of Purdue University by J. W. White, Jr., in partial fulfillnient of the requirements for the degree of master of science, August. 19.39.
