MICROSCOPE HOT STAGE Use in Melting Point Determinations F. W. MATTHEWS, Central Research Laboratory, Canadian Industries Limited, McMasteruille, Quebec .i new design of hot stage uses an electrically heated copper block surrounded by a water cooling jacket. With improved electric control this provides rapid heating and cooling of the stage and reproducible heating at melting point.
M
AXY devices for the determination of melting point are
described in the literature. Recently hot stages for use on polarizing microscopes have been favored for precise work when only small amounts of material are available (8). The advantages and limitations of this technique are discussed by Jelley (4).
ing block is provided with a copper cover to enclose specimen recess. The hole in the cover is covered with a glass window (8 mm. square of Xo. 1microscope cover glass) to prevent convection air currents. Two eyes in the cover are provided, so that the lid may be lifted with a fork-shaped removable lifter. The heating block is supported inside the water jacket on three L-shaped Transite hlorks. A plug is inserted in the water chamber for
Klein ( 6 )described an electrically heated hot srage which eniployed a thermometer to measure the temperature. Kiethammer ( 7 ) described a similar stage but used a thermocouple and millivoltmeter to measure the temperature. The use of a polarizer and analyzer in the "crossed" position was described by Amdur and Hjort (1) and by Weygand and Gruntzig (10). Review articles on the design of microscope hot stages are given by Chamot and Mason (2)and Saylor (9). The latter gives a very brief description of a stage used by Stadinchenko for the study of the effect of heat on thin sections of oil-bearing rocks. The stage used water cooling to protect the microscope and was designed for operation a t a maximum temperature of 1000" C. Provision was made for the wide aperture required for higher magnifications. The necessity of the use of thermocouples with very fine wire and of a potentiometer circuit is pointed out by Saylor. The vertical illuminator described by M'eygand and Gruntzig (10) and by Pregl (8) allows a simplified stage construction to be used, inasmuch as it is not necessary to provide a hole through the stage for illumination of the specimen. A micromelting point hot stage having a number of refinements over those mentioned previously was described by Zscheile and White (12). This stage used a variable transformer for control of the heating current and thermocouples of fine wire for measurement of the temperature. Melting poiht measurements with this equipment were reported to be reproducible to 0.04" C. with certain substances. The main disadvantage of a stage constructed to this design was that the large co per block (weight approximately 2000 grams) was very slow to [eat and cool. Furthermore, very little control of the rate of heating was provided by the electric circuit and the protection provided for the microscope was not adequate for temperatures above 180" C. These are decided disadvantages in the routinr use of the stage in an industrial laboratory.
PLAN (COVERS REMOVED) A\
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The stage described below uses a heating block of smaller dimensions (weight approximately 100 grams) placed inside a water jacket. This allows faster heating and cooling of the stage and gives protection to the microscope a t temperatures well above the range of melting point determinations in routine organic work. An electric circuit provides good control of the rate of heating and of the temperature gradient a t the melting point.
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DESCRIPTION OF HOT STAGE
The stage consists of an electrically heated copper block surrounded by a jacket in which water may be circulated (Figure 1). The copper block is recessed in the top to provide a chamber for the specimen and on the under side to receive the electric resistance heating element. The element consists of 45 cm. (18 inches) of KO.26 gage (B & S) Nichrome wire wound on a Transite or mica form. The element is enclosed on the under side by a Transite cover and on the upper side by mica insulation. A V groove cut in the upper side of the block to the depth of the specimen recess permits the use of capillaries as specimen mounts. A short groove is cut in the floor of the recess to facilitate the use of tweezers in the placement and removal of cover glass mounted specimens. The axis of the block is drilled (diameter, 3/32 inch) to provide for observation of the specimen by transmitted light. The heat-
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Figure 1. Diagram of Hot Stage Scale shown is in inches
A . Top cover, brass B . Heater cover c. Water jacket, brass D . Water connections. brass E . Heater block F . Supports for E, Transite G. Heating coil form, Transite H . Heating coil cover, Transite I. Insulators, porcelain J . Heater leads K. Thermocouple leads
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Stage clamps, brass
M. Insulation, mica N . Specimen (between) glass P. Window (8 mm. sa), glass
Screws t o hold W ,brass Eyelets for lifting B , brass V groove for capillary, inserts T. Groove for thermocouple, leads U. Groove for tweezers W. Frame 0.004 inch shim, brass
Q. R. S.
V O L U M E 20, NO. 11, N O V E M B E R 1 9 4 8
1113
CONSTANT
"OLThCL TRANSFORMER
STEPDOWN
TRANSFORMER
HEATING ELEMENT
loo" TO 24"
Figure 2. Eleotric Heating Circuit
bringing electric heater and thermocouple leads to the heating block. Porcelain tubes are used for the element leads which are brought to insulated terminals fastened t o the water jacket, on which an electric plug is mounted (not shown in Figure 1). The lid and base are drilled with holes to correspond to the axial hole in the heating black. These holes may be somewhat larger (0.125 inch) to allow for a cone of illumination and observation. Two stage clips on the water jacket allow the hot stage to be centered on the light path of the microscope. All metal parts are chromium plated to lower heat radiation and to add to the corrosion resistanoe and sppessance of the stage. The electric heating circuit is shown in schematic dmgram in Figure 2. The voltage regulator of the constant voltage transformer type is required if the stage is t o be used a t constant. temperatun. The u4e of a stepdown transformer following the varixblc autotransformer (Varix) changes the approximate
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Stenton Aves., Philadelphia 44, Pa.). The e.m.f. from the couple may be measured with a potentiometer such as Leeds & Northrup No. 8657C or 8662. With the former the temperature may be estimated to 10.4" C. with the latter to +O.lo C. The higher accuracy mould be reduired only in the most precise work. The thermocouple junction may be used as a clip t o hold the cover glasses enclosing the specimen, The dimensions of the junction are such that the field observed can conveniently include the junction of the thermocouple. By the clip action the junction is held in goad thermal contact with the cover glass. To lessen the lass of heat from the thermacauple junction by conduction along the leads, a turn around the bcated black is made in a groove cut for this purpose. The cold junction is kept in an ice bath which should be agitated before final readings are made. Alternatively the junction may be kept in the 25" circulating system used for the water jacket, or a t the terminals of the potentiometer. With a potentiometer equipped with compensating circuik, this correction can be applicd electrically. OPERATION OF HOI' STAGE
The microscope required for melting point measurement need only be a simple stand which may be adapted for use with polarized light by inserting & Nicol prism below the condenser lens and a. second prism above the object lens. Provision should be made for one of the p r i s m to be rotated to the "crossed" position with respect t o the other. Satisfactory results-were obtained by the use of Polaroid attachments for obtaining this crossed position. A 3 X objective and 15X eyepiece give suitable magnification for mast purposes. The aperture of the optical system is low in the design given but adequate for a 3 X objective. For higher magnifications this should be increased hy tapering the hole in the heater lid outward and upward at an angle-of about 15"and by enlarging the boles in the cooling cell to 0.375 inch. Adequate protection is given for closer working objectives by the water cooli n s rdl. The nireat.ion of the mwTt.ive
6u6~
heating element wire (B & S No. 32, 10.5 ohms per foot) available a t the time the heater was constructed would have required a very long wire to be mounted in the heater recess. An ammeter OSA is connected in series with the heating element. The voltage regulator and stepdown transformer are refinements which can be eliminated for less critical work. The controls m y .~ be mounted in a small panel nrhch provides switches for the heating circuit, microscope lamp, and water circulation pump (Figures3and4left). TEMPERATURE MEASUR13MENT
The temperature of the specimmen is measured with a thermocouple made from No. 30 B & S gage iron and constantan glass-insulated wire (supplied by Leeds & NorthrupI, Roeklsnd &
Figure 4. Hot Stage (Covers Removed on Stage of Microscope)
1114
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ANALYTICAL CHEMISTRY
the more usual sizes present in the laboratory. The small groove cut in the base of the specimen recess facilitates the placing and removal of specimens. With substances having high vapor pressure there is a tendency for the sample to sublime out of the field in such a mount. This difficulty has been met by previous workers by the use of calcium oxide casein cement (6) to seal the cover glasses. The usual melting point capillary was found satisfactory. This can be plugged with glass wool or sealed in the case of substances which sublime very readily.
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5 120CI MAXIMUM TEMPERATURE.
RATE OF HEATING I ~ M I N . RATE OF HEATING 5;/MIN.
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Figure 6.
8 R A R OF HEATING 57MIN 01
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Figure 5.
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Rate of Heating a t Various Heating Currents
Water is circulated in the water jacket from a reservoir which is maintained at 25' =t 0.1' C. This provides the instrument with a definite ground level temperature. Under these conditions a given electrical current will give a reproducible temperature gradient and maximum temperature. The instrument may also be used as a constant temperature stage, the temperature of which can be adjusted by changing the heating current. The constancy of the thermostat will depend on the temperature of the circulating water and fluctuations in the line voltage. Using a voltage regulator on the heating current and circulating water constant to =t0.2' C. the temperature of the stage was constant to 1 0 . 2 ' C. The use of thermostatically controlled cooling water is also a refinement which may be eliminated for less critical work. Such a system is, however, often available in an organic laboratory for refractive index, viscosity, or density determinations. The heating rates possible are shown graphically in Figure 5. At the highest heating current the stage can be heated to 200 C. in less than 5 minutes. As the temperature of the stage iS a balance between the heating effect of the electric current and the cooling of the circulating water, the stage cools quickly when the heating current is reduced. Figure 6 shows the maxicurrent and the current mum temperature reached for a given necessary to give a rate of heating of 1 or 5 per minute a t any O
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desired temperature. This graph should be constantly a t hand when the stage is in use. In normal operation the stage is heated quickly to determine an approximate melting point if this information is not already known. A second sample is then mounted and the stage is heated quickly to about 10' below the approximate melting point. At this point the current is set to give the desired rate of heating a t the melting point. This factor may conveniently be chosen at 1' per minute except in the case of substances which decompose rapidly near the melting point, where a higher gradient m y be desirable. As the substance starts to melt, the heating current is reduced and can be adjusted to hold the specimen a t the melting point and cause crystals to grow or melt with temperature changes of less than 0.1 C. With some pure, dry specimens this temperature can be reproduced to 0.1' C. The determination of this equilibrium temperature, where crystals exist in a pool of melt, is the true melting point of the substance. Such a measurement, which is accomplished with the apparatus described, is not possible with most of the previously described methods for routine melting point determination. O
CALIBRATION OF APPARATUS
The thermocouple used and the potentiometer were calibrated a t the benzoic acid setting point. The benzoic acid setting point was found with a benzoic acid cell supplied and certified by the National Bureau of Standards (N.B.S. cell No. 1, setting point 122.352' =t 0.003' C.). The thermocouple reading of this cell given by Leeds & Northrup potentiometer LX 399,686 was 122.37' C. The correction factor was therefore -0.02' C. Suitable samples for the calibration of melting points to 0.1 C. are not easily obtained. In lists given for this purpose (2, 3), the temperature is often stated to better than 1O and no directions for adequate purification are included. The samples were proO
V O L U M E 20, NO. 11, N O V E M B E R 1 9 4 8 Table 1. Calibration Melting Point Substance Bennoia add
Anthraquinone
Melting Point.
C.
122.374 * 0.002 215.6 0.3
*
Found, * C. 122.4 215.8
$ded by the National Bureau of Standards. The results given in Tahle I show that the melting point determined by the method described is accurate to 10.2' C. on an absolute scale. The reproducibility is about half this figure. ACKNOWLEDGMENT
Acknowledgment is made to Canadian Industries Limited for permission to publish this paper. The author wishes to thank J. H. Michell for his interest and help in the develo-pment of this instrument and C. P. Saylor, Natio;oal Bureau i f Strrndar.ds, for the cdihration standards used in t hlis work. LITERATURE C1T E D (1) Amdur, I., and Hjort, F,. V.. IND. Ewe. G x ~ M . ANAL . ML, 2.
259 (1930).
1115 (2) Chamot, E. M., and Maaon, C. W.. "Handbook of Chemical Microscopy," 2nd ed.. Vol. I, p. 198, New York, John Wiley & Sons, 1938.
(3) Clwk, E. P., "Semimicro Quantitative Organic Analysis," p. 21. New York. Academic F'res%. 1943. (4) Jell&, E. E., in Weissberger, A,, "Physic a1 Methods of Organic Chemistry," Vol. I, p. 4515, New Yoi.k, Interscience Press, 1945. (5) Klein. G., Mikroohemie Pregl-Festschvift, 192 (1929). (6) Kofleer, L.. and Hilhek, H., MikroJlemie. 9. 38 (19:31); 15, 242 (1934). (7) Niethammer. A.,IEid.. 7,223 (1929). (8) Pregl, F., "Quantitative Organic Microanalysis," 4th English ed., ed. by J. Grant, p. 185, Philadelphia, P. Blakiston's Son Co., 1946. (9) Saylor. C. P., "Temperature, Its Measurement and Control in Science and Industrv." D. 673. New York. Reinhold Puhlishing Corp., 1941 (10) Weygsnd. C . , and GrUntaig. W., Mdkrachemie, IO. 1 (1932). (11) Zsoheile, F. P., and White, J. W., IND.EN& CHEM.,ANAL.En., 12,436 (1940) R e c e m ~ oJune 9, 1947
Detection of Cyanides and C . E. HUBACH, Alcohol Tar Unit L P O T A S S I U M ferrocyanide hss been used extensively in in the European countries and to a smller degree United States for the precipitation of excessive quantities Of iron in wine. This treatment, known in the wine industry &s blue fining, is used because of ita effectiveness in removing iron which is exceedingly troublesome, if allowed to remain in the wine. Compounds are formed with the tannin and coloring matter p r e s b t , which gradually precipitate and cause cloudiness after filtering and bottling. The use of potassium ferrocyanide is aeeompanied by considerable danger to consumers of wine because an excess may remain in solution if the treatment is not supervised hy a competent chemist. Decomposition into hydrooymic acid takes place according to the equation Fe(CN)e----
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specified apparatus. No acid need he added, as the acids naturally present in wine are sufficient. Sulfuric acid liberates hydrocyanic acid from ferrocyanides; therefore, its use would obscure the test far free hydrocyanic mid if ferrocyanides are also present in the sample. y .
.
. B
,
+ ZHQD+Fe(CN)sHzO--- + OH- + H C N
This action is appreciable even in solution in distilled water and is accelerated in acid solutions as the pH decreases (3). The acids of wine, therefore, favor the formation of free hydrocyanic acid. The method proposed by Gettler and Goldhaum for the detection and estimation of microquantities of cyanide (B) is exceedingly well suited for the detection of free hydrocyanic acid in wine. It provides a characteristic and delicate test. The Prussian blue formed is specific for hydrocyanic acid and is concentrated an a suitably chosen small area of contrasting white filter paper; the faintest blue is readily discernible. It can also he used, when modified ss described below, for the detection of soluble ferrooyanides in wine and insoluble ferrocyanides in sediment from wine. In testing wine for the presence of free hydrocyanic acid, no preliminary treatment is required. The method is employed as described except that aeration should take place a t room temperature instead of at 90" because the hydrolysis of ferrocyanide, accelerated a t the higher temperature, would produce enough free hydrocyanic acid to give a positive test. Five or 10 ml. of the wine are placed in the aeration tube which is eonneoted to the
Figure 1. Test Papers Stains 1 2 and 3 produced in 4-mm. Range r e p r e d i.0, 3.0, and 5.0 microgram of pqtassium feirooyanide and 0.4, 1.3, and 2.2 nucroarams of hydrocyanic acid. reweotiuely. strlna A i F,
ana 7 nrori,irori
in
eanaa
To detect soluble potassium ferrocyanide remaining in wine after treatment or to prove the presence of insoluble iron ferrocyanides in the sediment from the bottom of wine tanks after the treatment, use is made of the fact that hydrocyanic acid is liberated quantitatively from soluble and insoluble ferrocyanides by sulfuric acid in the presence of cuprous chloride a t temperatures near 100' C. ( 1 ) . To the sample in the amation tube remaining from the test for free hydrocyanic acid, 10 to 15 mg. of powdered cuprous chloride and 1 ml. of a 20% solution of sulfuric acid in water are added. Cuprous chloride acts as a catalyst and is continuously regenerated; therefore the quantity used is not a critical factor. A fresh disk is placed between the flanges and the test is performed as described in the original article (a). Numerous tests were made on very dilute aqueous solutions of potassium ferrocyanide of known strength and also on samples of
