Environ. Sci. Technol. 1992, 26, 153-159
Uptake of Organic Chemicals in Conifer Needles: Surface Adsorption and Permeability of Cuticles Lukas Schreiber" and Jorg Schonherr Lehrstuhl fur Botanik, Technische Universitat Munchen, Arcisstrasse 2 1, D-8000 Munchen 2, Germany
Rates of uptake of organics [ (2,4-dichlorophenoxy)acetic acid, triadimenol, lindane, bitertanol, and pentachlorophenol (PCP)] in needles from five different conifer species (Picea abies, Picea pungens, Pinus sylvestris, Abies alba, and Abies koreana) were investigated. Uptake (mole uptake vs time) from aqueous solutions was biphasic. The first phase is attributed to adsorption of the organics on the needle surfaces. The second phase represents uptake into the needles. Surface sorption and uptake into needles increased with lipophilicity of compounds and could be predicted from cuticlelwater partition coefficients. Specific surface areas of the needles, determined from monolayer sorption of PCP, varied by more than 1 order of magnitude between different plant species. Since differences in rates of uptake between needles from different species correlated well with specific surface areas, needles with large surface areas showed appreciably higher rates of accumulation of organics at equal environmental concentrations than needles having lower specific surfaces. Implications concerning foliar uptake of organics and exposure monitoring are discussed.
Table I. Physicochemical Properties of the Test Compounds
Introduction Organic chemicals of anthropogenic origin occur in substantial amounts in the atmosphere (1). Forest ecosystems have a large surface area (2) and act like filters for polluted rain and air. Volatile compounds can reach the living plant tissue via stomata. Nonvolatile substances dissolved in rain or fog are intercepted by the plant cuticle, which forms the interface between the plant and the environment (3). Before they can reach the living plant tissue, they must diffuse through the plant cuticle. The plant cuticle is a lipophilic polymer membrane, which covers the aerial parts of plants and limits passive water loss to the atmosphere (4). It consists of the insoluble bipolymer cutin (mainly composed of hydroxyalkanoic acid monomers) (5) and waxlike lipids (6),which are deposited within the polymer membrane and on the cuticle/atmosphere interface. Sorption capacity of cuticles (7) and their permeability (8) for lipophilic substances can be high. Diffusion through leaf cuticles represents the main route and the rate-limiting step in uptake for nonvolatile lipophilic environmental chemicals, which are deposited to leaf surfaces. Uptake of these chemicals via roots is small, because the mobilities of lipophilic substances in soils are limited (9). A number of publications have recently focused on plant uptake of organics (10-13) after long-term exposure. Numerous organics were found in leaves, but with the exception of ref 10, no attempts were made to identify leaf compartments where organics were located. In this type of exposure monitoring it must be tacitly assumed that organics are rapidly taken up into leaves and once taken up wiU stay there for a long time. Riederer (10) has argued that organic sorbed in various leaf compartments are mobil and can be lost to the atmosphere. In addition, translocation and metabolism will reduce the amounts of xenobiotics that can be found in leaves. All these factors must be considered before any conclusions about exposure can be drawn from amounts of xenobiotics associated with
Experimental Section Plant Materials. Needles from five different plant species were investigated. Healthy needles from the species Picea abies (L.) Karst., Picea pungens Engelm., Pinus sylvestris L., Abies alba Mill., and Abies koreana Wils. were collected from 5-10-year-old trees growing in a rural area 80 km from Munich. Chemicals. 14C-labeled chemicals were used in the experiments. 2,4-Di~hlorophenoxy[~~C]acetic acid (specific activity 2.04 X 10l2Bqmol-l) and [14C]lindane (specific activity 2.37 X 10l2Bq-mol-l) were purchased from Amersham Buchler, Braunschweig, FRG. Pentachloro[14C]phenol (specific activity 1.37 X 10l2Bq-mol-') was purchased from CEA, Grenoble, France. [14C]Triadimenol (specific activity 7.6 X loll Bqmol-l) and [14C]bitertanol (specific activity 6.3 X loll Bqmol-l) were gifts from Bayer AG, Leverkusen, FRG. Radiochemical purity was better than 99% in all cases and was checked regularly by radio-TLC. Compounds were dissolved in deionized water. Pentachlorophenol (PCP) and (2,4-dichlorophenoxy)aceticacid (2,4-D) are weak acids and solutions were buffered with citric acid (0.01 M). The pH was adjusted to 3.0 using KOH. Only the nondissociated species penetrate into the plant cuticle (14). The pK, values of 2,4-D and PCP are 2.73 and 4.73, respectively (15). Thus, at pH 3.0 the concentrations of the nonionized species are smaller than the analytical concentrations by factors of 0.35 (2,4-D) and 0.982 (PCP) (16). Cuticlelwater partition coefficients K,, were used as a quantitative measure of the lipophilicity of the substances (17) (Table I). There was no measurable loss of 14C02 from needles previously incubated with the four test compounds. Thus, the radioactivity associated with the needles is a quantitative measure of uptake. Determination of the Radioactivity. The radioactive concentrations of the donor solutions were determined by liquid scintillation counting (TRI Carb 2000, Packard,
0013-936X/92/0926-0153$03.00/0
substance
MW"
log Kowb
2,4-D triadimenol lindane bitertanol PCP
221 296 291 337 266
2.63 3.12 3.80 4.16 4.79
log KcWc log cgd 2.59 2.88 3.73 4.05 4.75
+0.26 -0.72 -1.66 -2.17 -4.35
" MW, molecular weight. log KO,,1-octanol/water partition coefficient: 2,4-D from ref 15; triadimenol and bitertanol from Bayer AG (private communication) (means of a and fl isomers); lindane from ref 24; PCP from ref 28. clog K,, cuticle/water partition coefficients from ref 17; lindane from our own determination. dlog co, water solubility (m~l-m-~): 2,4-D from ref 15; triadimenol and bitertanol from Bayer AG (private communication); lindane from ref 24; PCP from our own determination. leaves. We have, therefore, measured rates of uptake of organics into conifer needles as affected by properties of the organics and needles. Loss of organics contained in various compartments of the needles was also measured.
0 1991 American Chemical Society
Environ. Sci. Technol., Vol. 26, No. 1, 1992 153
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120 180 240 300 360 Time (min)
Figure 1. (A) Biphasic uptake of PCP in A . koreana needles from the aqueous phase within the first 2 h. (B) Uptake of PCP in needles of five conifer species from the aqueous phase. The first phase is calculated by extrapolating the y intercept of the regression line fied to the second phase.
Downers Grove, IL). Needles were oxidized in pure oxygen (oxidizer 306, Packard). The 14C02was trapped quantitatively and mixed with scintillation cocktail. Radioactivity of these samples was determined by liquid scintillation counting. The 2a error was 1% or less in all cases. Necessary quench corrections were made. Determination of Uptake from Aqueous Solutions. Aqueous donor solutions (4 mL), containing the 14C-labeled chemical ( molm3), were pipeted into 4.5-mL glass vials. Needles were cut at their bases using a razor blade. Four needles were incubated in each of the vials. The vials were closed with aluminum lined screwcaps and were shaken by inversion during experiments in the dark at 25 OC (*0.5 "C). At the end of the experiment the needles were blotted carefully with filter paper to remove adhering drops of donor solution before being oxidized. Distribution of radioactivity over the length of the whole needles was investigated in order to determine whether significant amounts of radioactivity could enter the interior of the needle through the cut edge at the needle base. After an incubation of 24 h in radioactive donor solution, the needles were cut in 3-mm slices and the radioactivity of the single slices was determined. The distribution of radioactivity was homogeneous from the base to the top with all compounds and conifer species. This indicated that an uptake of radioactivity through the cut edge at the needle base was negligible. However, to be on the safe side, N
154
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Table 11. Specific Surface Areas (A spec) Determined with PCP Adsorption and Projected Surface Areas (A),,, Calculated from Area-Length Correlations per I-mm Needle Length species
Aspeo mm2.mm-'
ci
APRY mrn2.mm-'
P. abies P. sylvestris P. pungens A. alba A. koreana
29.4 41.3 65.7 254.0 683.0
15.2 f3.9 f6.3 f22.3 f92.1
3 3 3 5 5
Aspee/Apro 9.8 13.8 21.9 50.8 136.6
the lower portions of the needles (3 mm) were discarded in all subsequent experiments. When uptake was plotted vs time, biphasic graphs were obtained for all compounds and species (Figure lA,B). As shown below (results of adsorption isotherms and desorption experiments), the initial rapid uptake can be attributed to sorption of chemicals to the needle surface, while the slow uptake is due to diffusion across the cuticle into the needle. The slope of the slow uptake (Figure 1B) is called flow ( m o l d ) . The flow is proportional to the area A (m2), across which uptake occurred, and to driving force Ac (m~lam-~), representing the difference between the concentration of the test compound in the donor and the receiver, respectively (14). The proportionality constant P (me-') is a mass conductance called permeance P = F/AAc (1) The slow uptake was always steady (r 1 0.99), and the donor concentrations were practically constant during the course of the experiment. Thus, F is readily obtained from the slope uptake vs time (Figure 1B). It is well established that uptake into leaves is limited by the permeability of the cuticle (14). In our experiments, the concentrations of solutes at the inner surfaces of the cuticles were not known. We therefore used the donor concentrations as the driving force, assuming the inner concentrations remained negligibly small during experimentation. This seems reasonable, because (i) the amounts taken up were very small, (ii) cells and organelles act as sinks for lipophilic materials, which prevents an increase in solute concentration in the cell wall near the inner surface of the cuticle, and (iii) the uptake was in fact steady and did not level off (Figure 1B). The two-dimensional, projected surface areas of needles Apro(Table 11) were obtained from arealength correlations as described in ref 18. Desorption Experiments. Desorption experiments were carried out, using PCP and the needles of A. koreana, to determine those amounts of radioactivity that were either sorbed superficially or taken up irreversibly into the needle interior. Needles were incubated for 1,17, and 20 h in radioactive donor solutions. After being blotted to remove adhering donor solution, they were incubated in vials containing 4 mL of sodium borate buffer (pH 9.0). At pH 9.0 PCP molecules are completely dissociated and the concentration of nonionized PCP in the solution is essentially zero (19). Desorption media were exchanged after predetermined periods of time. At the end of the desorption experiments, needles were oxidized to determine residual radioactivity not desorbed. Loss of PCP from the needles to the atmosphere after the aqueous solution had been removed was determined using the following experimental design. A constant flow of air (2.3 X IO4 cm3.h-l) was bubbled through a large volume (500 mL) of water. This airstream, saturated with water vapor, passed through two subsequent wash bottles (30 mL). Wash bottles and connections were made of
borosilicate glass. Needles, preincubated for 1h in aqueous PCP solution, were blotted carefully and placed in the first wash bottle. The second wash bottle contained cold (0 "C) cocktail, in which the PCP was quantitatively trapped. The scintillation cocktail was exchanged every hour and radioactivity was measured. The largest part of the PCP, lost to the vapor phase, was trapped in the cocktail. The amount of PCP sorbed to the glass walls of the desorption bottle was recovered by rinsing the apparatus with cocktail. After needles were desorbed for 25 h in the vapor phase, they were further subjected to desorption with sodium borate buffer as described above. Adsorption Isotherms. Adsorption isotherms of PCP to the waxy needle surfaces were determined using aqueous PCP concentrations from 10-~ to m ~ l . m - ~Concen. trations higher than m ~ l - mwere - ~ prepared by adding nonradioactive PCP (purity 299%)(Sigma, St. Louis, MO) to radioactive PCP. Uptake of PCP was measured with each concentration, as described above. A linear regression line was fitted to the slow phase of the biphasic uptake kinetic. T h e y intercepts of the regression line were taken as the amounts of PCP adsorbed to the needle surface (Figure 1B). A plot of the amount of PCP adsorbed (mol-mm-l) vs donor concentration ( m ~ l - m -resulted ~) in an adsorption isotherm, which was characterized by a weakly pronounced plateau and a subsequent multilayer adsorption. Adsorption of molecules from aqueous solution on solid surfaces can be described by the BET isotherm equation (20-22): X,Bc X = (2) (co - c)[l + ( B - N C / C O ) l
X (mol-mm-') describes the amount of molecules adsorbed at the surface of a 1-mm needle length at any donor concentration, X, (mol.mm-l) is the amount of molecules in is)the donor concentration, co the monolayer, c ( m ~ l . m - ~ ( m ~ l - m -is~ )the aqueous solubility of PCP at the temperature used (25 "C), and B is the dimensionless BET constant. Rearranging yields the following expression: C/CO = - +1- - B - l c (3) X(l-C/C,) X,B X,B co For reduced concentrations c / c o in the range of 0.01--0.2, a plot of (c/co)/X(l- c / c o ) vs c / c o gives a straight line. The slope of this plot corresponds to (B - l)/X,B and can be used to calculate the BET constant B. The intercept corresponds to l/X,B, from which the number of molecules in the monolayer X, can be calculated. Furthermore, if the size of the PCP molecule in the monolayer is known, the specific area of the needle surface Aspec(Table 11) covered by PCP molecules can be calculated by multiplying the number of PCP molecules in the monomolecular layer with the required surface of the PCP molecule (23). Experimental Error and Statistics. Rates of uptake of a given compound and species differed considerably among individual needles. Coefficients of variation were -30% in most cases, sometimes even higher. This largely represents biological variability, because experimental errors were much smaller. In uptake experiments, needles were sampled at four time intervals (e.g., 1.5, 3.0, 4.5, and 6.0 h). At each sampling time, five replications with at least four needles were investigated. Each uptake experiment (uptake of one organic chemical in the needles of one plant species) was repeated two to four times. In desorption experiments, five replications with at least four needles in each case were investigated. Since results were reproducible in all cases,
replications of an experiment were combined. Unless stated otherwise, results are given as means with 95% confidence intervals (ci). Statistical calculations were performed using SPSS/PC+ statistical software (SPSS Inc., Chicago, IL). Results and Discussion
Time Course of Uptake from Aqueous Solutions. Uptake into needles of five different plant species was biphasic for all compounds, as shown for PCP in Figure 1A,B. The rapid first phase of uptake was completed after -30 min (Figure 1A). During the second phase (30-360 min), uptake continued at considerably slower rates that were constant throughout. The amounts of PCP taken up during the rapid phase per millimeter length of the needle at a constant donor concentration m ~ l - m - were ~) quantified by extrapolating the steady-state rates of uptake to zero time (Figure 1B). The waxy needle surfaces are not impermeable to PCP. Sorption on the surface and penetration into the needles occur simultaneously. The two processes can be separated, because penetration into the needles proceeds much slower than sorption to the surface. By extrapolating steady-state uptake (penetration) to zero time the amounts in the cuticles are substrated from the total amount of PCP associated with the needle. Thus, the y intercept is an estimate of the amount of PCP sorbed on the needle surface. Sorption on the needle surface depends on mixing of and diffusion in the donor solutions, while penetration into the needle is largely determined by the diffusion coefficient in the cuticles. Cuticles are solids and diffusion coefficients are smaller by up to 6 orders of magnitude than in liquids (14). This is the main reason why sorption and penetration can be separated. These y intercepts and the rates of uptake between 30-360 min increased in the order P. abies, P. sylvestris, P. pungens, A. alba, and A. koreana. Surface adsorption and rates of uptake of different compounds increased with increasing cuticle/water partition coefficient (Kcw)of the compounds in the order 2,4-D, triadimenol, lindane, bitertanol, and PCP. Adsorption Isotherms. The isotherms for all species had the same shape as demonstrated for the two species P. abies and A. alba (Figure 2A,B) and resembled BETtype isotherms (20-22). They can be interpreted as follows: In the lower concentration range the needle surface was saturated with a monomolecular layer of PCP molecules, resulting in the plateau of the adsorption isotherms. Further increase of the donor concentration leads to multilayer adsorption, which was reflecteed by the second steep slope of the adsorption isotherm. The data could be fitted successfully to eq 3 (Figure 3A,B). The amount of PCP molecules in the monolayer (X,) was highest for A. koreana and lowest for P. abies. The BET constant B was the same within experimental error for all needle surfaces (Table 111). These results are consistent with PCP sorption to the surface during the first 30 min (Figure 1B). If penetration into the cuticles would have been rate limiting during the rapid-uptake phase, the dependence on donor concentration of uptake should have been linear (14). Desorption. Only 81-98% of the PCP could be desorbed from needles during 24 h (Table IV). The fraction that could be recovered was smaller the longer the duration of the preincubation. After -4-6 h, the slopes of the desorption curves became zero (Figure 4A). This means that PCP inside the needle cannot be desorbed. This fraction is likely composed of PCP sorbed in lipid compartments of cells and ionized PCP in aqueous compartEnviron. Sci. Technol., Vol. 26, No. 1, 1992
155
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Figure 2. PCP adsorption isotherms of the needles of (A) P . abies and (B) A . a/ba. The solid lines are calculated BET isotherms according to eq 2.
Table 111. Parameters of the BET Equation Fitted to the Lower Portions of PCP Adsorption Isotherms for Needles of Selected Conifer Species. BET Constants B with 95% Confidence Intervals (ci) and Monolayer Capacities X, (molomm-I) Related to 1-mm Needle Length with 95% Confidence Intervals species
B
ci
4.4 5.7 4.1 7.1 5.6
f2.5 fl.1 11.5 f1.2 f1.8
X,, mol.mm-' 6.79 X 9.81 X 15.60 X 60.20 X 162.00 X
lo-" lo-" 10-l' lo-''
lo-''
ci f1.21 X lo-" f0.94 X lo-*' f1.50 X lo-" f5.30 X lo-'' 122.00 X lo-"
Table IV. Desorption of PCP from Abies koreaaa Needles. Fractions of PCP Irreversibly Sorbed in the Needle Interior and Desorbed within 24 h with 95% Confidence Intervals (ci) preincub, h
total desorption in 24 h
irreversible fraction in needle interior
1 17 20
0.98 f 0.02 0.90 f 0.01 0.81 f 0.02
0.02 f 0.001 0.10 f 0.006 0.19 f 0.012
ments having a relatively high pH (ion trap). After short uptake periods lasting only 1h (Figure 4A), 88% of the PCP associated with the needles were desorbed nearly instantaneously (less than 1min). This represents 156
Environ. Sci. Technol., Vol. 26, No. 1, 1992
c 13
1
2
c 15
020
c/c,
Concentration x l o 7 (mol I - ' )
P. abies P. syluestris P. pungens A. alba A . koreana
005
Figure 3. BET plots of the adsorption isotherms of the needles of (A)
P . abies and (B) A . alba. Coefficients of correlation of the linear regressions were 0.96 with P . abies and 0.98 with A . a/ba.
the fraction sorbed to the surface, because diffusion coefficients in waxes could not account for this rapid efflux (14).
Permeances. Penetration of organics across the cuticles can be quantified by permeances as defined by eq 1. Very large differences in rates of uptake (at constant driving forces) among compounds and conifer species were observed. Data obtained with 2,4-D and PCP show that the differences between plant species are fully accounted for by differences in specific surface areas of the needles (Figure 5A,B). If specific surface areas are used in calculating permeances ( P ) ,significant differences between plant species are not apparent in most cases (Table V). Permeances increased in order of increasing lipophilicity, that is, 2,4-D, triadimenol, lindane, bitertanol, and PCP. Thus, differences in rates of uptake between the four conifer species tested for a given compound were solely caused by differences in specific surface areas of the needles. For a given plant species, the differences in permeances between compounds were due to different lipophilicities of the compounds (Figure 6). This is seen from the excellent correlation between log P and log K,, ( r = 0.99) log P = 1.00 (4~0.12)log K,, - 13.15 (f0.44) (4) This excellent correlation between permeances and partition coefficients is somewhat surprising, since P is also a function of diffusion coefficients (D)and the path length ( I ) (14). It appears that either the effect of K,, on P was
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Figure 4. (A, top) Desorption of FCP from needles of A . koreana after different times of incubation in aqueous donor solution. The solid line was calculated by nonlinear regression from experimentalvalues. M o represents the total amount of radioactivity assoclated wlth the needles at the beginning of the desorption. M,is the amount of radioactivity desorbed after defined periods of time. (B, bottom) Desorption of PCP from needles of A . koreana after 1 h of incubation in aqueous donor solution and different times of desorption from the vapor phase. The solid line was calculated by nonlinear regression from experimental values. M,and M , are defined as in (A).
dominating the effect of D and 1 or the ratio D/1 was constant for all species. We have shown that rates of uptake (at constant driving forces) are proportional to specific surface areas between solutions and needles and partition coefficients. Since surface sorption per millimeter length of needle depends on the same variables, rates of uptake for all compounds and species are only a function of the amounts sorbed on the needles (Figure 7). Sorption might be high because a compound is very lipophilic or because of pronounced surface roughness; the effects on rates of uptake are just the same. This is evidence that both variables act independently and that surface adsorption is in fact the first step in foliar penetration. Desorption into Air. Since large amounts of organics can be adsorbed on needle surfaces from aqueous solutions within short periods of times, the question arises as to what happens when the water evaporates. Will the fraction sorbed to the surface be lost to the atmosphere if the compound is volatile? From our test compounds only PCP Pa at 25 had a relatively high vapor pressure [9.3 X "C, (24)].To test loss of PCP to the atmosphere, needles were incubated in aqueous PCP solution for 1h. After this
Figure 5. Correlations of the rates of uptake of (A) PCP (r = 0.99) and (6)2,4-D (r = 0.99) with the specific surface areas AaWcof the different needles.
--8.5 8'o m
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Flgure 6. Correlatlon of the permeances (P)of the flve test compounds with cuticle/water partition coefficient K,. log P = 1.00 (It0.12)log Kcw- 13.15 (f0.44); r = 0.99.
treatment, 98% of the total PCP associated with the needles was sorbed on the surface (Table IV).When these needles were flushed with moist air, only 11%of the PCP was lost to the atmosphere during 25 h, while 89% of the PCP, initially adsorbed to the needle surface, continued to penetrate into the needle (Figure 4B). Thus, organics Environ. Sci. Technol., Vol. 26,No. 1, 1992
157
Table V. Permeances of Lipophilic Test Compounds in Needles of Selected Conifer Species. Permeances P with 95% Confidence Intervals (ci) and Means of the Permeances of One Compound in the Different Needles with Coefficients of Variation (cv)
P, m-s-'
species
P. abies P. syluestris P. pungens A. alba A . koreana
ci
2,4-D 3.51 x lo-" 3.78 X 2.24 x lo-" 3.11 X 2.71 X lo-"
f0.5 X 10-l' f0.5 X lo-" f0.8 X f1.4 x lo-" f0.6 X
3.07 X lo-" (cv 20%)
P. abies P. pungens A. alba A. koreana
P. sylvestris P. pungens
Triadimenol 4.00 X lo-" 7.00 X 10-l' 5.58 x lo-" 2.54 X 4.78 X 10-l' (cv 40%) Lindane 5.01 X 4.23 x 10-lo 4.62 X
P. abies P. pungens A. alba A. koreana
-
f0.7 X f 1 . 2 x 10-10
(cv 12%)
Bitertanol 4.36 X 7.95 x 10-10 6.84 X 4.23 X 5.85 X
P. abies P. syluestris P. pungens A . alba A. koreana
*0.9 x 10-11 f0.4 X 11.7 X lo-" f0.9 x 10-11
5.00 x 3.30 x 6.63 x 3.90 x 4.20 x 4.61 X
f3.0
X
12.7 X f 2 . 1 x 10-10 f1.6 X
(cv 32%)
PCP f2.5 fo.7 f1.3 fi.6 f1.7
1049 1045 1049 1049 10-09
x 1049 x 10-05 X
lo*
x 1049 X
lo*
k v 28%)
A
-15
r
t
-16
t
-17
1
-18
"
n
0
c -
D
1
V
-13
-1 2
Abies Abies Picea Pinur Picea
koreona aiba
pungens sylvestris abies
-"0
-1-
log Adsorption (mol mm
)
Flgure 7. Correlation of the rates of uptake of the five test compounds In needles of the five plant species with their rates of adsorption to the needle surfaces. log rate of uptake = 1.07 ( f 0 . l ) log adsorption - 4.1 (fl.O);r = 0.98.
sorbed to the lipophilic needle surfaces exhibit little tendency to volatilize. Summary and Conclusions. Uptake of lipophilic organics and penetration into the cuticles was preceded by sorption to the waxy surface of the needles. Surface adsorption was a very rapid process, while diffusion into the cuticle proceeded much more slowly. The amounts sorbed on the surface and the rates of penetration into the cuticle differed greatly between compounds and plant 158
Environ. Sci. Technol., Vol. 26, No. 1 , 1992
species. Two variables can fully account for these differences. The amounts sorbed on the surface of the needles are proportional to the specific surface areas between solutions and needles and to the cuticle/water partition coefficients. Penetration into the needles proceeds from the organics sorbed on the surface. The amounts that penetrated per unit time and specific surface area (permenaces) were constant for all species. These two determinants of sorption and penetration (specific surface area and cuticle/water partition coefficient) acted independently. For a given compound (i.e., constant partition coefficient),all differences between species in sorption and penetration were caused by differences in specific surface area. For a given plant species (Le., constant specific surface area), all differences in sorption and penetration were due to differences in partition coefficients. Specific surface areas can easily be determined if the BET constant for surface sorption of a model compound of known space requirements (we used PCP) is known. Cuticle/water partition coefficients may be calculated from molecular connectivity indexes ( I 7). With these variables known, permeances for our five test conifers can be calculated for compounds not included in our study, provided they are within the range of partition coefficients used in this study (Table I). The size of the water droplets affects the rates of penetration for two reasons: (a) The smaller the droplets, the higher the ratio surface area/volume. Since lipophilic organics tend to accumulate at the air/water interface of the droplets, the average concentration of these substances in the water can be larger by orders of magnitude than predicted by the Henry constants (25). This effect increases with decreasing droplet size. (b) If the volume of a sessile droplet on a needle is constant, the time needed until half the amount of an organic originally present in the droplet has penetrated into the needle (the half-time) can easily be calculated. This half time depends on the permeance and on the ratio droplet volume/A,pec. This ratio decreases with decreasing size of the droplet (14),and the half-time is directly proportional to this ratio. This means that the concentration of pollutants decreases more rapidly in small droplets than in larger ones. This dependence on the ratio volume/contact area (which is not constant) prevents us from calculating half-times of penetration simply as a function of permeance. Permeances of our test compounds ranged from -1 X to 2 X m/s. If droplets are hemispherical and have a radius of 1 pm, half-times would range from -104 s to 14.4 h. For droplets 10 times that size, halftimes would be 10 times longer (14). Thus, depending on permeance and on droplet size, penetration can be rapid or it may take days before significant amounts can be taken up into needles. These considerations indicate that rapid fluctuations in concentrations of airborne contaminants are not likely to result in rapid fluctuations in leaf concentrations. Even though very rapid uptake of large amounts of organics into conifer needles is possible only under favorable conditions (very small droplets of fog loaded with organics having a high permeance being deposited on needles having a large specific surface area), uptake of specific amounts of organics into needles will occur from long-lasting air pollution. The ecotoxicological consequences of this uptake will depend on the fate of the organics in the needles and their phytotoxicity. No problems will arise if the material is metabolized to nontoxic compounds in rates comparable to the rates of uptake. If the rates of uptake are larger than rates of metabolism, phy-
totoxicity could develop. Under this condition, conifers having large specific surface areas would show damage earlier and the damage might be more severe, because with these species, surface adsorption as well as rates of uptake would be larger. Organics that are not readily metabolized will tend to accumulate in needles. Depending on the duration of exposure and rates of metabolism and translocation, as well as life expectancy of the needles, internal concentrations in the vital tissue may reach threshold levels for nonspecific toxicity (26) to develop. We do not know what concentrations of organics are actually required for nonspecific toxicity in conifers, but an important aspect should be pointed out. In nonspecific toxicity it is the total concentration of all organics that counts, and this may reach high levels much more readily than the concentration of individual compounds. A general description of foliar uptake as presented here and elsewhere (27)will eventually enable prediction of accumulation of all organics in leaves of plants. Acknowledgments
We gratefully acknowledge stimulating discussions with Dr. M. Riederer (Lehrstuhl fur Botanik, Technische Universitat Munchen, Arcisstrasse 21, D-8000 Munchen 2). Registry No. 2,4-D, 94-75-7; PCP, 87-86-5; triadimenol, 55219-65-3;lindane, 58-89-9; bitertanol, 55179-31-2. Literature Cited Helmig, D.; Muller, J.; Klein, W. Chemosphere 1989, 19, 1399-1412.
Schulze,E. D. In Encylopedia of Plant Physiology; Lange, 0. L., Nobel, P. S., Osmond, C. B., Ziegler, H., Eds.; Springer-Verlag: Berlin, 1982; Vol. 12B, pp 615-676. Schonherr,J.; Kerler, F.; Riederer, M. Dev. Plant. Biol. 1984, 9,491-498.
Schonherr,J. In Encyclopedia o f Plant Physiology;Lange, 0. L., Nobel, P. S., Osmond, C. B., Ziegler, H., Eds.; Springer-Verlag: Berlin, 1982; Vol. 12B, pp 153-179. Holloway,P. J. In The Plant Cuticle; Cutler, D. F., Alvin, K. L., Price, C. E., Eds.; Academic Press: London, 1982; pp 45-85. Baker, E. A. In The Plant Cuticle; Cutler, D. F., Alvin, K. L., Price, C. E., Eds.; Academic Press: London, 1982; pp 139-165.
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Received for review July 2, 1990. Revised manuscript received April 15,1991. Accepted July 29,1991. This work was supported by the Projektgruppe Bayern zur Erforschung der Wirkung von Umweltschadstoffen (PBWU).
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