Influence of Mineral Nutrition on the Sensitivity of Tomato Plants to Hydrogen Fluoride D. C. MacLean, 0. F. Roark,' G . Folkerts, and R . E. Schneider Boyce Thompson Institute for Plant Research, Yonkers, N.Y. 10701
Materials and Methods Tomato plants grown in sand culture and provided with a complete nutrient solution o r solutions deficient in calcium, magnesium, or potassium were exposed to various concentrations of hydrogen fluoride (HF) for periods of three, five, or 64 days. In magnesium deficient plants, H F exposure resulted in a reduction in the dry weight of both tops and roots and a n increase in the severity of magnesium deficiency symptoms on basal lea\,es. Foliar accumulation of fluoride was suppressed by magnesium deficiency. The effects of HF exposure on calcium deficient plants were characterized by reduced top growth, increased foliar chlorosis, and an enhancement of necrosis on apical leaves. Fluoride accumulation was unaffected by calcium nutrition. The responses of potassium deficient plants to H F were limited to increases in F accumulation and in necrosis on apical leaves. The differential responses of the elements to H F exposure were ascribed to differences in their mobility within plant tissues, the stability of fluoride salts formed. and 'or their metabolic roles.
T
he nutritional status of plants is generally believed to be one of the factors that determine the susceptibility of plants to atmospheric fluorides. However, published experimental results on this subject are rare. Brennan, Leone, et al. (1950) demonstrated that deficient and superoptimal levels of nitrogen, phosphorus, o r calcium retarded fluoride uptake and subsequent injury of tomato plants, whereas optimal concentrations of these elements favored fluoride-induced damage. Conversely. fluoride accumulation by bean seedlings exposed to hydrogen fluoride (HF) was enhanced under conditions of phosphorus. potassium, or iron deficiency (Applegate and Adams, 1960). Adams and Sulzbach (1961) reported that a n acute exposure to H F resulted in obvious injury to leaves of nitrogen deficient bean plants. Although tissues of plants grown on phosphorus, potassium, iron, or calcium deficient media accumulated equivalent amounts of fluorine, they were not visibly affected. H F fumigation experiments by McCune, Hitchcock, er a/. (1966) showed that increased tip burn of gladiolus leaf blades was associated with potassium, phosphorus, and magnesium deficiency, and decreased tip burn was observed in calcium and nitrogen deficient plants. The effects of calcium nutrition and H F fumigation o n the fruiting of tomato plants were investigated by Pack (1966). Low calcium reduced the size of fruit, and, in combination with H F , fruit size was further reduced, resulting in partially or completely seedless fruit. Pack (1966) suggested that H F might interfere with calcium metabolism during fertilization. The experiments described herein were performed to test the influence of three essential cations-calcium, magnesium, and potassium-on growth, fluoride accumulation, foliar injury, flowering, and fruiting of tomato plants exposed to gaseous HF. 1 Present address: West Essex High School, West Caldwell, New Jersey 07006.
Tomato (Lycopersicon esculentum var. Bonny Best) seeds were germinated in soil. When the seedlings were two weeks old, soil was washed from the roots and three uniform seedlings were transplanted to 5-inch plastic pots containing washed (Robbins, 1946) flint-shot quartz sand. Cultures were maintained in a greenhouse and irrigated with one-fourth strength Hoagland's solution for one week following transplanting. Two seedlings were then removed from each culture, leaving one uniform plant per pot, which was automatically subirrigated twice daily with one of seven nutrient solutions. The nutrient regimes included complete (Hoagland's) nutrient solution and solutions containing calcium, magnesium, or potassium at one-twentieth and one-fourth the concentration of the complete solution. Nutrient solutions were prepared as described by McCune, Hitchcock, et al. (1966). In deficient solutions, a n equivalent amount of sodium was substituted for the element varied. When deficiency symptoms first became apparent on plants given the lowest nutrient levels (18-21 days after transplanting), all plants were transferred to Mylarcovered fumigation chambers (Hitchcock, Zimmerman, et ai., 1962) and exposed to HF o r filtered ambient air. Four separate fumigation experiments were carried out. Exposure to average concentrations of: (a) 0, 4.7, o r 12.0 pg. F/m.a for five days; (b) 0, 1.0, or 15.1 pg. F/m.3 for five days; (c) 0 or 8.9 pg. F/m.3 for three days; and (d) 0 or 3.1 pg. F/m.S for 64 days. (Control fumigations listed as 0 pg. F/m.S were known t o be less than 0.2 pg. F / m 3 . ) The two potassium deficient treatments were not included in the last experiment. Treatments were replicated four times, except in the 64-day fumigation, in which three replicates were used. Before, during, and after fumigation, lateral shoots were removed as they developed to maintain plants with a single stem. Growth rates were estimated by periodic measurements of the distance between the cotyledonary node and the terminal bud. Foliar injury was subjectively assessed by rating the extent of chlorosis and necrosis on each leaf. An injury index of 1 , 2 , 3 , and 4 was equivalent to no chlorosis, slight, moderate, and severe chlorosis, respectively; and 1, 3, 5 , and 7 to no necrosis, slight, moderate, and severe necrosis. The number of nodes per plant (8-9) at the time foliar injury was assessed was relatively less variable than plant height. Therefore, the ratings for the uppermost four leaves of each plant were defined as apical injury and the ratings for the remaining leaves as basal injury. Fluoride accumulation by vegetation was determined using the semi-automated method of analysis (Mandl, Weinstein, et al., 1966; Weinstein, Mandl, et al., 1963). Unless otherwise noted, surface fluoride was removed prior to analysis by washing for 30 sec. in Alconox-EDTA solution (i.e., 0.05% Alconox plus 0.05 disodium ethylenediamine tetraacetic acid, w/v) followed by two successive 15-sec. rinses in deionized water. Results
Visible symptoms of nutrient deficiencies. Foliar markings induced by nutrient deficiencies in the absence of HF are Volume 3, Number 11, November 1969 1201
Table I. Effect of Nutrient Treatment on Stem Elongation of Bonny Best Tomato Plants During and After Fumigation with HF Concentration Stem elongation (cm.). relative to Eloneation Postcomplete du;ing fumigation Nutrient varied medium fumigation elongation , . . 11,18a 56.02a None (complete) 1/20 7.47c 9.72~ Calcium 1/4 11.72a 49.88b Calcium 1/20 7.48~ 57.17a Magnesium 1i4 10.71ab 55.12a Magnesium 1/20 9.68b 50.91b Potassium 1/4 10.54ab 47.12b Potassium ~~~
~
~
~
a Means within a column followed by different letters are significantly different at the 5 % level (Duncan, 1955).
Table 11. Effect of Nutrient Treatment and HF Exposure (0 or 3 pg. F/m.3 for 64 days) on the Dry Weight of Tops and Roots of Bonny Best Tomato Plants ConcentraDry weight Dry weight tion (grams) of tops (grams) of roots tive to exposed to HF exposed to HF concentrations of concentrations of dete 0 3 0 pgi Nutrient varied medium F/m. F/m. F/m. None (complete) . , , 58.05 36.48O 5 . 1 5 4 . 9 6 8.61. 6.224 2.73. 1.44. Calcium 1/20 33.47. 25.31a'b 4 . 6 8 3.38 1/4 Calcium 4.42. 1.230 0.91. 1/20 5.84. Magnesium 45.35 23.41a.b 5 . 4 2 3.84* 114 Magnesium
;/gi
summarized as follows. Symptoms of magnesium deficiency were first apparent on the oldest leaves and progressed acropetally. Magnesium at one-fourth the complete level induced intercostal chlorosis on leaves from the oldest 2 or 3 nodes. O n plants supplied with magnesium at one-twentieth of that in the complete solution, symptoms progressed from mild chlorosis to severe intercostal chlorosis with intercostal necrotic lesions, and, finally, to desiccation and death. These progressive symptoms could be followed on a single leaf over a period of two weeks or by making comparisons of adjacent leaves along the stem. Symptoms of calcium deficiency were confined to apical leaves. One-fourth strength calcium resulted in a yellow-green color of expanding and young, fully expanded leaves. One-twentieth strength calcium resulted in marginal necrosis, intercostal chlorosis, and death of the terminal buds. Calcium shortage did not affect basal leaves. Potassium deficiency did not result in significant visible foliar markings. Occasionany, very mild marginal chlorosis of basal leaves was noted on plants cuftured on the medium with the lowest potassium concentration. Growth. Elongation of single-stem plants exposed to 0, 4.7, o r 12.0 pg. F/m.a for five days was measured a t three-day intervals throughout the experiment. N o significant effect of HF exposure on growth within a nutrient treatment was observed during fumigation or the post-fumigation period. Therefore, the data for control and HF exposed plants were pooled in the analysis of the effects of nutrient treatment I202 Environmental Seience & Technology
Table 111. Effect of Nutrient Culture and HF Concentration on the Development of Chlorosis on Leaves of Bonny Best Tomato Plants ConcenChlorosis ratings after a 5-day tration relative to exposure to HF concentrations Nutrient complete (pg. Fim.3) of ~varied medium 0 4.7 12.0 None (complete) ... 1.12 1.25 1.50 Calcium 1/20 3.88. 4.79 6.75.~~ Calcium 114 1.19 1.81 1.94 Magnesium 1/20 5.12. 4.44. 4.25. Magnesium 114 1.06 1.12 1.44 Potassium 1/20 1.19 1.31 1.38 Potassium 1 /4 1.25 1.12 1.75 h
Significantly different from plants in complete medium ( 5 2 level). Significantly different from nonfumigated plants ( 5 Z level).
(Table I). Elongation during fumigation was suppressed significantly in plants grown on media containing calcium, magnesium, or potassium at one-twentieth of that in the complete medium. Post-fumigation growth was inhibited more than 80% at the lowest calcium level. Calcium at one-fourth the complete level and both potassium treatments significantly reduced post-fumigation growth also. Magnesium deficiency did not affect post-fumigation elongation. The dry weights of tops (above ground plant parts) and roots, measured after a 64-day exposure to 0 or 3 pg. F/m.3, are given in Table 11. In this experiment, lateral shoots were not removed periodically, and the plants were allowed to branch. Exposure to HF did not affect the already severe inhibition of dry weight induced hy one-twentieth strength calcium or magnesium. In these plants the lack of calcium repeatedly killed terminal growth as new shoots developed, resulting in stunted plants having a witches'-broom appearance. Magnesium deficiency resulted in tall, spindly plants in which the reduction in dry weight may have been accentuated by desiccation and abscission of leaves from the lowest two or three nodes. H F significantly reduced the dry weight of tops of plants grown on complete nutrient and one-fourth strength calcium o r magnesium. Although one-fourth strength calcium resulted in a significant reduction compared to the complete nutrient, in both fumigated and nonfumigated plants, a significant effect of one-fourth strength magnesium was present only in H F fumigated plants. Inhibition of the dry weight of roots was found only in plants provided the lowest calcium and magnesium levels (one-twentieth strength). Although root growth of plants exposed to H F was, in all treatments, less than that of those provided filtered air, the only statistically significant reduction due to H F exposure was in plants cultured on onefourth magnesium. No visible symptoms of injury on roots were observed in treatments where root growth was inhibited. Foliar injury. Foliar injury assessed three days after a fiveday fumigation showed that H F exposure and the position of the affected leaves on the plant (apical rs. basal), significantly altered the amount and extent of chlorosis, necrosis, and foliar injury (the sum of chlorosis and necrosis ratings) . Chlorosis was significantly greater on plants grown on calcium or magnesium at one-twentieth of that in the complete medium (Table 111). The only effect of H F o n chlorosis was seen on plants provided the lowest calcium level. Chlorosis was significantly increased by exposure to 12.0 pg. F/m.3.
Table IV. Effect of Nutrient Culture and HF Concentration on the Foliar Necrosis Ratings of Apical and Basal Leaves of Bonny Best Tomato Plants
'1
Nutrient varied
Concentration relative tocomplete medium
None (complete) Calcium Calcium Magnesium Magnesium Potassium Potassium
1/20 1/4 1/20 114 1/20 1/4
. . .
--
Necrosis ratings of leaves after a 5-day exposure to HF concentrations (Mg. F/m.3)of 4.7
0
12.0
-~
Apical
Basal
Apical
Basal
Apical
Basal
1 .OOaa 13.00g 1.25ab 1.00a 1.00a 1.00a 1.00a
2.25abc 1.50ab 1.50ab 5.75e 2.00ab 1.00a 1.00a
1.75ab 16.00h 3.75abcde 1 .OOa 2.75abc 4.25bcde 3.50abcde
2.75abc 2 . OOab 3.75abcde 10.00f 2 , OOab 1.25ab 2.50abc
1 .25ab 19.75i 5 . OOcde 3 . OOabcd 5.OOcde 4.75bcde 5,25de
2 . OOab 2 . OOab 3.25abcd 11 .50fg 4 . OObcde 1 .OOa 1 .75ab
Any t w o means not followed by the same letter differ significantly at the 5 % level (Duncan, 1955).
Table V. Mean Foliar Accumulation of Fluoride by Leaves of Bonny Best Tomato Plants as Affected by Mineral Nutrition and HF Exposure ConcenFluoride content (p.p.m. dry weight) after exposure for tration 3 days 5 days 64 days relative to HF concentration (bg. F/rns3) comdete 8 . 9~ 0 15.1 0 3.1 Nutrient varied medium _. ~0 ~ 1 .o _ _ 196.2 131,3~ 171.8 103.5. 179.0 245.0. 211 .o Significantly differer from plants in c mplete medium ( 5 % level).
None (complete) Calcium Calcium Magnesium Magnesium Potassium Potassium $1
. . .
1/20 1/4 1/20 1i4 1i20 1/4
2.2 1.8 4.3 1.5 1. o 1.0 1.5
Necrosis ratings revealed a relationship between nutrient treatment, HF exposure, and the position of necrotic leaves on the plant (Table IV). The lowest calcium and magnesium levels resulted in significant necrosis of apical and basal leaves, respectively. on plants that were not exposed to HF. Necrosis on apical leaves of plants cultured o n one-twentieth strength calcium became more intense with each increase in H F concentration. Leaves of plants supplied one-fourth strength calcium did not exhibit increased necrosis when exposed to 4.7 pg. F/m.3, whereas a moderate, but significant increase in terminal leaf necrosis was induced o n plants exposed to 12.0 pg. F/m. 3. The appearance of necrosis was such that the contributions from each variable could not be separated to determine if H F enhanced the calcium deficiency symptoms or if calcium shortage favored HF-induced injury. Necrosis induced by magnesium deficiency generally was confined to the lower leaves. Necrosis induced by onetwentieth strength magnesium o n basal leaves was intensified by exposure to either 4.7 or 12.0 p g . F/m. This necrosis clearly resembled magnesium deficiency symptoms and was not similar to HF-induced markings produced o n younger leaves. Plants grown on magnesium at one-fourth the complete complement did not show significantly increased necrosis of basal leaves; however, exposure to 12.0 pg. F/m.3 resulted in significant increases in necrosis o n apical leaves. The higher HF concentration induced apical necrosis on all potassium deficient plants. At the lower H F concentration, a significant amount of apical necrosis was found only o n plants
6.6 6.9 7.6 4.9 5.6 5.4 4.8
38.6 37.6 48.2 39.6 38.6 40.3 34.0
254.4 263.2 254.6 216.8" 171.60 301.40 310.70
9.3 9.6 13.5 5.3 8.1 , . .
...
233.1 273.9 223.9 197.8 173,7~
... ...
grown with one-twentieth strength potassium. In general, the severity of necrosis o n basal leaves was not affected by fumigation or by calcium or potassium shortage. Fluoride accumulation. Post-fumigation analyses of tomato leaflets for fluoride showed that exposure to filtered ambient air for 3, 5, o r 64 days did not result in significant fluoride accumulation. Deficiencies of the three cations elicited different responses with respect to fluoride accumulation (Table V). I n plants exposed to 15.1 pg. F/m.3 for five days, accumulation of fluoride was decreased hy magnesium deficiency, increased by potassium deficiency, and unaffected by calcium deficiency. The accumulation of fluoride was similarly decreased by onefourth strength magnesium in plants exposed to 3.1 pg. F/m? for 64 days and by one-twentieth strength when exposed to 8.9 pg. F/m.3 for three days. The only significant effect of calcium deficiency o n fluoride accumulation was a reduction in plants grown at one-twentieth calcium and exposed to 8.9 pg. Fim. 3 . After one fumigation experiment (0, 1.0, and 15.1 pg. F/m. for five days), tomato plants were fractionated into three groups: leaflets; stems; and the petiole fraction, which included the petioles, petiolules, and rachises. Analyses of the unwashed stem and petiole fractions showed that the amount of fluoride accumulated in these tissues was negligible and not affected by nutrient culture or H F exposure. Roots were not analyzed. Flowering and fruiting. The indeterminate nature of flower production characteristic of the Bonny Best variety and within treatment variability made effects on flowering difficult to Volume 3, Number 11, November 1969
1203
assess. However, no flowers o r fruit were produced on plants grown on media containing calcium o r magnesium at onetwentieth the concentration of the complete medium. The time of flowering and the number of flowers produced o n plants in all other nutrient treatments were not affected by exposure to HF at 4.7 or 12.0 pg. F/m. a for 5 days or to 0 or 3.1 pg. F/m.3 for 64 days. Fruit production by plants cultured on complete nutrient media was not affected by H F during the 64day period. External appearance and locule and seed development appeared normal in all fruits regardless of treatment. Discussion A discussion of the interacting effects of H F exposure and mineral nutrition must also take into consideration some of the pertinent factors of mineral nutrition in relation to plant growth and development. Plant nutrition is complex, and interactions between various nutrients essential for plant growth have been documented. When HF-induced responses in plants deficient in a given nutrient are considered, it should be understood that these responses may be due to a nutritional imbalance peculiar to that element, rather than to the deficiency of that element per se. Thus, it is possible that the observed interactions between nutrient deficiency and fluoride exposure may be indirect, rather than direct, effects. The metabolic patterns of leaves differ with age. In young leaves synthetic reactions and growth predominate, whereas in older leaves, photosynthesis is the dominant function, and little growth occurs. Mineral nutrients are transported via the transpirational stream to all leaves; however, retransport from older leaves to actively growing tissues prevents the accumulation of toxic concentrations. The rate of retransport is a function of the mobility of a mineral. The location of mineral deficiency symptoms on the plant is also a function of mobility. Magnesium and potassium are relatively mobile elements, and, when the external supply is limited, retransport occurs and deficiency symptoms first appear o n basal leaves. Calcium, on the other hand, is relatively immobile, and deficiency symptoms are first apparent on apical leaves. HF-induced foliar injury was limited to apical leaves, and was observed in calcium, magnesium, and potassium deficient plants. However, the increased injury to apical leaves in calcium deficient plants was more than additive, and, in magnesium deficient plants, injury to basal leaves was markedly increased by H F exposure. Thus, for both calcium and magnesium deficient plants, foliar injury resulting from the nutrient-HF treatments was related to the relative mobility of these cations and was most pronounced in tissues in which the nutrient shortage was most acute. Although potassium shortage was the least effective mineral deficiency tested, fluoride uptake was enhanced in potassium deficient plants, whereas fluoride accumulation was generally suppressed by magnesium deficiency and unaffected by calcium deficiency. The observed enhancement of fluoride accumulation by potassium deficiency is in agreement with the data of Applegate and Adams (1960) for bean seedlings, and the absence of a relationship between calcium deficiency and
1204 Environmental Science & Technology
fluoride uptake is in accord with the findings of Pack (1966) and Applegate and Adams (1960). However, Brennan, Leone, et a/. (1950) found that optimal levels of calcium increased fluoride uptake, and deficient and superoptimal levels suppressed its accumulation. The response of plants to H F fumigation is not only a function of the concentration of the pollutant and the duration of exposure, but also of the rate and pattern of fluoride absorption, translocation, and accumulation. Therefore, any external or internal factors that affect one or more of these processes can also alter the sensitivity of plants to HF. In this investigation it was shown that potassium deficiency did not affect the sensitivity of tomato plants to HF, whereas calcium and magnesium deficiency rendered plants more susceptible to HFinduced growth inhibition and foliar injury. These differential responses of the nutrients may be explained by considering their metabolic roles. Calcium and magnesium ions are required as cofactors in some essential enzymic reactions, and, when unavailable, these reactions cannot proceed. Potassium ions are also involved in essential enzymic reactions, but as activators rather than cofactors. When the amount of available potassium is low, the reactions are not completely inhibited, but usually proceed at reduced rates. The relative stability of fluoride salts formed within plant tissues is another factor to account for the potassium effect. K F is relatively soluble and, when the external supply of potassium ions is limited, dissociation can occur readilk. resulting in available potassium ions. CaF2 and MgF2, on the other hand, are relatively stable salts and, when calcium or magnesium ions are in short supply, the effective concentration within the plant is further reduced due to the relative insolubility of these salts. Literature Cited Adams, D. F., Sulzbach, C. W., Science 133,1425-6 (1961). Applesate. H. G.. Adams, D. F., Phyton (Buenos Aires) ‘14, 111-20 (1960). Brennan. E. G.. Leone. I. A.. Daines. R. H.. Plant Ph\-siol. 25, 736-47 (1950). Duncan, D. C., Biometrics 11, 1-42 (1955). Hitchcock, A. E., Zimmerman, P. W., Coe, R . R., Contrib. Boyce Thompson Inst. 21, 303-44 (1962). McCune, D. C . , Hitchcock, A. E., Weinstein, L. H., ibid. 23, 295-9 (1966). Mandl, R. H., Weinstein, L. H., Jacobson, J. S., McCune, D. C., Hitchcock, A. E., in “Technicon Symposia, 1965,” L. T. Skeggs, Jr., Ed., 270-3, Mediad Inc., New York, 1966. Pack, M. R., J . Air Pollution Control Assoc. 16, 541-4 (1966). Robbins, W. R., SoilSci. 62,3-22 (1946). Weinstein, L. H., Mandl, R. H., McCune, D. C., Jacobson, J. S., Hitchcock, A. E., Contrib. Boyce Thompson Inst. 22, 207-20 (1963). Receired for review February 24, 1969. Accepted August 21, 1969. This work was supported in part by Grant No. AP-00189 from the National Air Pollution Control Administration, US. Public Health Service. 0. F. R. was in the NSF High School Teacher Research Participation Program (G W-1692), 1967.
