CRYOBIOLOGY

16, 272-286

( 1979)

Temperature Dependence of the Microwave Properties of Aqueous Solutions of Ethylene Glycol Between +15”C and -70°C JEFFREY

D. MACKLIS,* ERNEST

FREDERICK D. KETTERER,t G. CRAVALHO :, 1

AND

* Cryogenic Engineering Laboratory, Massachusetts Institute of Technology, bridge, Massachusetts 02139, t Harrison Department of Surgical Research the Department of Electrical Engineering, Vnioersity of Pennsylvania, Philadelphia, Pennsylvania 19104, and 1 Matsushita Professor of Mechnical Engineering in Medicine, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 INTRODUCTION

Certain difficulties involved in the microwave thawing of whole organs stored at sub-freezing temperatures have been discussed briefly in a previous paper (14). Problems are encountered both in the monitoring of organ temperature and in the strong temperature dependence of the tissue and perfusate electrical properties (2, 3, 7, 13, 19, 20). Thermal runaway, leading to the simultaneous appearance of hot spots and frozen regions, results from this temperature variation due to the higher microwave absorption of warmer tissue (1, 2, 6, 9, 20). In addition, depending both upon the frequencies used for the thawing and upon the electrical properties of the organ, interior focusing due to electric field heating or outside-in thawing due to magnetic field heating may result ( 12). The effects of cryoprotective perfusates upon tissue properties are important and must be considered in order to better control microwave thawing. In a previous work, the temperature Received August 18, 1978; accepted March 15, 1979. 1 Address reprint requests to Prof. E. G. Cravalho, Room 16-520, M.I.T., Cambridge, MA 02139. 272 OOll-2240/79/030272-15$02.00/0 Copyright 0 1979 by Academic Press, Inc. All rights of reproduction in any form reserved.

Camand

dependence of the electrical properties of 100% solutions of ethylene glycol, dimethyl sulfoxide (DMSO or Me$O), and glycerol were determined ( 14). The experimental system described by Noorchashm (15) was modified for improved accuracy, and his analytic solution of the electromagnetic fields within the system’s reentrant cavity was used. These allowed an accurate determination of the dielectric constant and loss tangent of the three chemicals from experimental measurements. Substances of known properties were measured to verify the system’s accuracy and the validity of the mathematical model. Due to the availability of commercial microwave equipment at 0.915 GHz and 2.450 GHz and due to the demonstrated usefulness of these frequencies for organ thawing (6, 9, 20), 1.5 GHz was chosen as the approximate frequency for investigation. Data obtained at 1.5 GHz are typical of those at either 0.915 GHz ‘or 2.450 GHz ( 19, 21). Results were quite reproducible over four to six experimental runs for each cryoprotectant. Similarities in the general temperature dependencies of the microwave properties of the Me2S0, ethylene glycol, and glycerol were demonstrated, but dis-

MICROWAVE

PROPERTIES

tinct differences were revealed. MeLIS exhibited the most active behavior, especially at room temperature, with dielectric constant values several times those of the other two chemicals. All three showed a steady decrease in dielectric constant value with decreasing temperature. Loss tangent temperature dependence varied dramatically, with maxima and plateaus occurring at different temperatures for each of the cryoprotectants. Me,SO and ethylene glycol were both shown to have appreciable microwave absorption below -60°C but ethylene glycol exhibited a smoother low temperature loss tangent slope. The result:5 of this preliminary study indicated that a more thorough investigation of aqueous solutions of cryoprotectants would be useful, both in the design of electrically appropriate perfusate mixtures and in the understanding of cryoprotectant phase equilibria. Although it is generally true that electrical properties of aqueous solutions show features that derive directly from those of the pure liquids (10, Chap. 7), little is known of the specific temperature dependencies of the dielectric constant and loss tangent of aqueous solutions of cryoprotectants. Therefore, a study

FIG. 1. Schematic of experimental cryoprotective chemicals.

OF ETHYLENE

273

GLYCOL

of concentrations of more practical interest was warranted. Microwave property temperature dependencies of ethylene glycol exhibited features desired for microwave thawing. Absorption at low temperature, along with a relatively mild change with increasing temperature, would allow a fast and controlled thaw. In addition, ethylene glycol has been shown to be relatively non-toxic to whole excised rat hearts at such moderate concentrations as 3 M to 5 M (5). For these reasons, ethylene glycol was chosen for further study. This paper reports the results of this study of practical concentration solutions, discusses the physical implications, and makes recommendations as to desirable concentrations for use as a cryoprotecting perfusate. MATERIALS

AND

METHODS

The present study employed the same field theory analysis, mathematical model, computer software, and instrumentation described previously ( 14). The experimental system is briefly described here and is illustrated in Fig. 1. A microwave sweep oscillator provides a continuous wave output which is isolated from the sweep oscil-

system for the measurement

of electrical

properties

of

274

MACKLIS,

KETTERER,

AND

CRAVALHO

r--;--t

PROPORTIONAL TEMPERATURE CONTROLLER

I FIG. 2. Feedback

controlled

system for the stabilization

lator, sampled by a frequency counter, externally square wave modulated, and attenuated to the desired level. This signal excites the single mode of the reentrant cavity. Output from the cavity is detected by a crystal detector, isolated, and fed into a lock-in amplifier which extracts the signal from the noise and also provides the reference signal to the modulator. Output from the amplifier is displayed versus frequency on an X-Y plotter, showing changes in cavity resonant frequency bandwidth due to the sample. Temperature control and measurement are again provided by the system depicted in Fig. 2 ( 15). Liquid nitrogen allows feedback controlled cooling to -100°C and heating to +SO”C with total sample isolation. A thermocouple in direct contact with the sample is used for temperature measurement with a precision of 1 PV (approximately 0.025”C). Resonance curves are obtained at many fixed temperatures in the specific range. These yield resonance frequency and bandwidth data which are

and read-out

of sample

temperature.

used, along with empty cavity data, as inputs into the mathematical model. Dielectric constant and loss tangent versus temperature curves are thus computed. Conversion charts based on Noorchashm’s exact model for cavity parameters used in this study are shown in Fig. 3. These curves have the following functional dependencies :

t’ = s(W)> tan 6 = K*, E where E’ = dielectric constant, Af = resonance frequency shift, K = cavity constant, AB = bandwidth change, tan 6 = loss tangent. The function g exhibits a weak dependence on Ahf. A standard measurement procedure almost identical to that used in the lOO$% solution study was employed here. At least 3 full temperature range experimental runs were made on each of the eight aqueous solutions of ethylene glycol studied: 0 M ( distilled water), four runs; 1 M, six runs;

MICROWAVE

PROPERTIES

OF ETHYLENE

GLYCOL

275

FIG. 3. Approximate conversion curves for the determination of electrical properties as functions of ‘cavity resonance frequency shift and bandwidth where B’ = g (Ai) and tan 6 = KAB/d.

2 M, three runs; 3 M, three runs; 4 M, six runs; 5h1, four runs; 10 M, four runs; 100% ethylene glycol, two runs plus six from the earlier study. All results were repeatable with a -t 3% range for the dielectric constant and within a +- 5% range for the loss tangent. After uniform pre-experimental preparation of the reentrant cavity and electronics, resonance curves showing changes due to the sample were made at temperature intervals of approximately 1°C to 3°C throughout the range of +lS’C to -70°C. Close to the phase change, measurements were made every O.l”C to 0.5”C in order to detect any irregularities. During each measurement, temperature drift

was held typically to O.l”C or less. Between points, the cooling rate was kept at a fairly constant 1.5”C per min, with the exception of slower cooling near the freezing point. Once at a satisfactorily low temperature, the sample temperature was held constant for between 0.5 hr and 1 hr. After this “pseudo-storage” period, the sample was warmed while measurements were taken every 1°C to 3°C. Between frequency measurements, the sample temperature was increased at a controlled rate of approximately 1°C per min, except near the phase change, where warming was slower and where measurements were more frequent. The slow rate of temperature change

276

MACKLIS,

KETTERER,

contributed to the low level of observed hysteresis, although hysteresis was slightly greater at low concentrations than with the 100% solutions. Neither the total measurement time nor the “storage” time had any effect on the results obtained. RESULTS

In order to evaluate reproducibility and to link these data with those of the previous investigation, measurements using 100% ethylene glycol were repeated with new batches of the chemical (Baker Reagent Grade). The new data fall within approximately 1°C of the previously published curves (14). These results and various standard measurements at room temperature show good reproducibility and a continuity between the experiments. Figures 4 through 11 show the electrical properties of aqueous solutions of ethylene

FIG. 4. 100% ethylene

glycol:

temperature

AND

CRAVALHO

glycol as a function of temperature. Figure 4 gives the properties of 100% ethylene glycol as presented in the preliminary investigation; the new data are almost identical. The strange temperature behavior near 0°C of the loss tangent of 100% ethylene glycol can be attributed to the trace amounts of water in Baker Reagent Grade ethylene glycol. As will be discussed later, extremely small concentrations of water are capable of disrupting ethylene glycol’s hydrogen bonding, altering its dielectric constant. These very small amounts of water may not follow conventional phase equilibria curves, but they may rapidly crystallize out of solution. This produces a sample of higher purity ethylene glycol below 0°C. The data shown suggest that concentration strongly affects the temperature dependence of the microwave properties of

dependence

of dielectric

constant

and loss tangent.

MICROWAVE

FIG. 5. 10 :M ethylene

glycol:

PROPERTIES

temperature

OF ETHYLENE

dependence

aqueous solutions of ethylene glycol. The temperature dependencies of both the dielectric constant and loss tangent differ in peak values, peak value temperatures, appearance of discontinuities, and changes in the shape of the curves. These variations with ethylene glycol concentration occur fairly smoothly with definite patterns in the temperature shifts and in the general behavior of the curves. Concentration greatly affects the temperature dependence of the dielectric constant in these solutions. Pure water, 1 M ethylene glycol, and 2 M ethylene glycol exhibit similar behavior with temperature, with a shift in the “discontinuity” temperature. At 0°C the dielectric constant of water falls dramatically; this results from the crystal lattice formation of the phase change. A solution of 1 M ethylene glycol exhibits a freezing point depression of

of dielectric

277

GLYCOL

constant

and

IOSS

tangent.

approximately 2.O”C (4); solid water is in equilibrium with the 1 M solution at this temperature. Consequently, the 1 M solution’s dielectric constant falls almost 90 units between -2.O”C and -2S”C. With the 2 M solution, rapid dielectric constant change occurs between -5°C and -7S”C. Since its freezing point depression is approximately 4.3”C (4), it follows that these low concentration solutions follow relatively “ideal” behavior with the dielectric constant temperature dependence. By the time a 3 M concentration of ethylene glycol is reached, the dielectric constant behavior is not so ideal. Although the freezing point is depressed to -7°C (4), the temperature of the steepest dielectric constant decline lies around -10°C. A distinct phase change is no longer present; rather, it occurs over a wider range in temperature. The dielectric

MACKLIS,

275

FIG. 6. 5

M

ethylene

glycol:

KETTERER,

temperature

AND

dependence

constant of 4 M ethylene glycol falls most rapidly between 0°C and -25°C; the freezing point depression of 10.4”C falls in the middle of this range. The dielectric constants of pure ethylene glycol, 10 M ethylene glycol, and 5 M ethylene glycol exhibit similar temperature dependencies without any rapid changes of slope. Ethylene glycol, which freezes at -11.5”C (4), h as its maximum value of 12.8 at +15”C. Similarly, the dielectric constant of the 10 M solution declines from 26.5 at +15”C, while that of the 5 M SOlution falls off from approximately 33 at f15”C. The freezing point depressions of 10 M and 5 M solutions are 14.6”C and 47.7”C, respectively. The magnitude of the dielectric constant at + 15°C continuously increases with decreasing ethylene glycol the concentration depenconcentration; dence is strongest between 2 M and 5 M,

CRAVALHO

of dielectric

constant

and loss tangent.

and it weakens toward both concentration extremes. Values of both the dielectric constant and loss tangent at +15”C and -65°C are shown in Table 1. The dielectric constant at -65°C has a different concentration dependence. With the exception of 100% ethylene ,glycol, which has a magnitude of approximately 1.6, the dielectric constant values at -65°C show a minimum near the 3 M concentration. The value increases both above and below this concentration. The dielectric constant of the 3 M solution is 1.9, whereas those of both 10 M ethylene glycol and pure water are greater than 3.0. An analysis of the family of loss tangent versus temperature curves uncovers a superposition of the effects of distilled water and ethylene glycol similar to that found in the dielectric constant data. The 3 M solution displays this joint effect to an ob-

MICROWAVE

PROPERTIES

OF ETHYLENE

TABLE Dielectric

GLYCOL

279

1

Constant and Loss Tangent Values for Aqueous Solutions of Ethylene Glycol at $15 and -65°C

r

c’

tan F,

7. 4 M ethylene

‘C

Temperature dependence of the microwave properties of aqueous solutions of ethylene glycol between +15 degrees C and -70 degrees C.

CRYOBIOLOGY 16, 272-286 ( 1979) Temperature Dependence of the Microwave Properties of Aqueous Solutions of Ethylene Glycol Between +15”C and -70°C...
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