Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)[No abstract available]1153341343ACRL; Association of College and Research Libraries; CAPES; Association of College and Research LibrariesCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Cella N.

Da Silva W.J.

Lima C.A.S.

Marquezini M.V.

Prioli A.J.

Serra D.B.

Silva E.C.

Vargas H.

English

Repositorio da Producao Cientifica e Intelectual da Unicamp

ANALYST, MARCH 1990, VOL. 115 341 
SHORT PAPERS 
Photoacoustic Assessment of the in vivo Genotypical Response of 
Corn to Toxic Aluminium 
Maria Valeria Marquezini, Norbert0 Cella, Edson C. Silva, Dinah B. Serra, Carlos A. S. Lima* and Helion 
Vargas 
lnstituto de Fisica, Universidade Estadual de Campinas, Unicamp, 13100 Campinas, Sao Paulo, Brazil 
Albert0 Jose Prioli and William Jose da Silva 
lnstituto de Biologia, Universidade Estadual de Campinas, Unicamp, 13100 Campinas, Sao Paulo, Brazil 
An open-cell photoacoustic (OCPA) spectroscopic technique is described which can assess genetically rooted 
differences in the response of  live specimens of  corn (Zea Mays L.) to the toxic action of  aluminium. 
Specimens f rom a susceptible and a tolerant inbred line were examined. The differences in their OCPA in vivo 
leaf spectra were evident and an interpretation is given based on  the genetically controlled response of the 
inbreds t o  aluminium toxicity. A n  earlier discrimination among specimens is obtained with this technique 
than has been possible so far w i th  previous techniques. 
Keywords: Photoacoustic spectroscopy; open-cell photoacoustic detector; aluminium toxicity in plants; 
genetically induced tolerance to aluminium toxicity; corn leaf spectrum 
Several reports have appeared on the development of inbred 
lines of corn ( Z e a  mays L.) in which both genetic variability 
and tolerant genotypes occur in relation to  the toxic effects of 
aluminium. 1-5 Hence a non-destructive analytical method for 
evaluating the differences in plant response to aluminium 
toxicity would be of value. 
Photoacoustic (PA) spectroscopy has been shown to  be an 
increasingly important tool in plant science related analytical 
investigati0ns.h-1’ This paper discusses its use in connection 
with the assessment of the aforementioned differences. 
Although the interpretation of the data, as far as accounting 
for a response pattern is concerned, is still preliminary, the 
demonstration of the usefulness of P A  spectroscopy in the 
present context is conclusive. 
Experimental 
Specimens 
Details of the specimens used have been given elsewhere.4.5 
Two lines were chosen: L1037 (AP+-susceptible) and L922 
(AI3+-tolerant). The aim was to compare the in vivo PA leaf 
spectra while keeping the specimens continuously exposed to 
different concentrations of aluminium. 
Set-up and Procedures 
In t2i\io PA leaf spectra were recorded using the open-cell 
photoacoustic (OCPA) configuration described by da Silva 
et al. l 2  and Bento et al. 13 This acted as the detecting element in 
a laboratory-built PA spectrometer consisting of a 1000-W 
xenon arc lamp (Oriel), a Jarrell-Ash monochromator, a 
Princeton Applied Research (PAR) rotating blade light 
chopper, a PAR lock-in amplifier, a Hewlett-Packard x - y 
recorder and a suitably interfaced Commodore micro- 
computer for on-line data acquisition and processing. Modula- 
tion was set to 20 Hz and all spectra were normalised to a 
carbon-black reference. The aim was to detect significant 
spectral differences between seedlings from a given inbred line 
* To whom correspondence should be addressed. 
and those from an equivalent line differing from the former 
only by the fact that they had been grown in a nutrient solution 
to which 9 p.p.m. of Al3+ had been added. Experiments were 
carried using pairs of seedlings of either the L1037 or  L922 
line. One member of each pair was grown under a zero 
concentration of Al3+ and served as a control specimen; the 
other, grown under a concentration of 9 p.p.m. of A P + ,  
served as the experimental specimen. 
I I I I I I 
Wavelengthlnm 
400 500 600 700 800 
Fig. 1. Photoacoustic spectra for Al3+-susceptible L1037 specimens. 
A, Specimens grown in a nutrient solution to which 9 p.p.m. of A13+ 
has been added; and B, specimens grown in Al”-free nutrient 
solution; the arrowed lines indicate intensities measured at the 
chlorophyll band peak with respect to the corresponding spectral base 
line. (a) Spectra of specimens exposed to the corresponding nutrient 
solution for 33 h; and ( b )  spectra of specimens after 57 h of exposure 
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View Article Online / Journal Homepage / Table of Contents for this issue
ANALYST, MAKCH 1990, VOL. 115 
0) 
m 
.z 1.0 
- 
2 
' Y  
+ I I  I 1 .  11 
T T  i T  
. ' 
4 
400 500 600 700 800 
Wavelengthinm 
Fig. 2 .  Photoacoustic spectra for A13+-tolerant L922 specimens 
(symbols as in Fig. 1) 
Results and Discussion 
Figs. l(a) and 2(a) and l(b) and 2(b) show representative PA 
leaf spectra for some selected specimens taken after continu- 
ous exposure to the corresponding nutrient solution for ca. 33 
and 57 h, respectively. The control and experimental speci- 
mens show essentially no spectral differences for the L922 
seedlings, in contrast to the differences observed for those 
from the L1037 line, even after exposure for only 30 h. It is 
apparent that for the latter the presence of AP+ significantly 
altered the in vivo optical response of the seedling leaves. This 
correlates well with a previous verification of phenotypical 
susceptibility involving radicular growth inhibition.5 Most of 
the spectral differences lie in the region dominated by the 
chlorophyll band, with a maximum at ca. 680 nm. In fact, a 
30% decrease in peak intensity was observed after exposure 
for 57 h. Again, no significant decrease was observed for the 
L922 line. The exposure time ( t )  behaviour of the ratio r( t )  = 
Sexperlmcntal/Scontrol, S being the appropriately normalised PA 
signal for each genotype, as measured at that peak, was also 
studied. The results are shown in Figs. 3 and 4. The data in 
Fig. 3 are consistent with a constant experimental value of 1.0 
for r ( t ) ,  even for long exposure times; this is consistent with 
the fact that L922 inbreds are known to show a high tolerance 
to AP+. In contrast, in Fig. 4, r ( t )  exhibits a steady decline at 
an approximately linear rate of 0.5 h-1, at least for values o f t  
down to 60 h. For longer exposure times, the apparent 
recovery of Sexperlrnenta] has still to be explained. 
In an attempt to interpret these data it should be remem- 
bered that under the experimental conditions used here the 
OCPA signal can be represented by S = A(3(h) avlk, where B(h) 
is the light absorption coefficient at wavelength h,  a the 
thermal diffusivity, k the thermal conductivity of the sample 
and A a constant. Now, taking a = (k/pc), which gives a in 
terms of the mass density, p, and the specific heat, c, of the 
sample, and considering measurements at a fixed value of A, 
say at the chlorophyll band peak, so that B(h) remains 
v) 1.5 
.- I 
q c , , , I  
15 35 55 75 
I? 0.5 
t/h 
Fig. 3. Plot of ratio of rclative intensities vprsus time for Ali+- 
tolerant L922 specimens (for details see text) 
rn 1.5 
.- 
c .- 
v) c + c .- 
15 35 55 75 
tlh 
Fig. 4. 
susceptible L1037 specimens (for details see text) 
Plot of ratio of relative intcnsities versus time for A13+- 
constant, we obtain S - (pc)-', an equation that expresses the 
dependence of the OCPA signal on the parameters of the 
sample that are relevant in this instance. 
On the basis of previous findings of genetic character,5 we 
can assume that for the L1037 seedlings exposed to Al3+ the 
volume of symplastic (intracellular) water decreases with 
time. As a result, the average heat capacity per unit volume at 
the plant leaf site where the in vivo OCPA measurements 
were made would increase. Such variations are not observed 
for the L922 specimens. By combining this information with 
the equation S - (pc)-l, it is possible to explain not only the 
t-invariance of Scontrol but also the observed decrease in the 
values of Sexpcrlmcnta1 for the L1037 specimens. However, in 
our view, a complete explanation of the behaviour of r(t)  is 
still required. To  this end the assumption could be made that 
such a decrease is somehow enhanced by the fact that the 
change in plant physiology results in a lower rate of 
chlorophyll production14 and thereby lowers its concentration 
in the plant leaves. 
For medium values o f t  (say, >60 h) the data for the L1037 
specimens appear to be consistent with the hypothesis that for 
any given concentration the impairing effects of A13+ will 
eventually show a tendency to subside or stabilise. Photo- 
acoustically, the proof of this would be Scxperirnentdl tending to a 
constant value. The observed trend in r ( t )  for this range of t 
values is consistent with this, particularly in view of the fact 
that, under such long and continued biochemical stress, 
fluctuations in chlorophyll content and morphological changes 
that can produce scattered results in the OCPA measurements 
are to be expected. However, at present we cannot fully 
explain the increase in r ( t )  (shown in Fig. 4) for t  values outside 
the ranges considered so far. One possible explanation would 
be an apparent adaptation response that appears to set in 
around and above ca. 60 h of exposure of susceptible 
specimens.5 The associated modifications in the plantules 
radicular system may enhance a recovery mechanism capable 
of offsetting, at least in part, the impairing effects of 
aluminium toxicity. The data presented here are still prelimi- 
nary, hence further studies are underway to ascertain whether 
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ANALYST, MARCH 1990, VOL. 11.5 343 
this correlates with the apparent recovery of the P A  response 
of the specimens. 
Conclusions 
It has been demonstrated that OCPA spectroscopy is a 
valuable analytical tool for the in vivo assessment of the 
genotypical response of corn plants to aluminium toxicity. The 
differences in the visible light OCPA spectra of the 
experimental and control specimens of either susceptible or  
tolerant inbred lines are very conspicuous for the former but 
hardly discernible for the latter, over the entire range of the 
visible spectrum. 
To the best of our knowledge this is the first report 
describing the use of an optically triggered thermoacoustic 
open-cell analytical technique for assessing differences that 
are deeply rooted in genetically controlled plant behaviour. 
To date the discrimination has relied on measurements 
associated with AP+-induced differences in radicular growth. 
This requires fairly long observation times leading to un- 
wanted effects that make i t  difficult to  interpret the results. In 
comparison, the technique described here can produce an 
accurate discrimination of responses at a far earlier stage 
among plant specimens that still appear normal. 
M. V. M. and N.  C.  acknowledge scholarships from the S2o 
Paulo Government Agency FAPESP (M. V. M.) and the 
Federal Government Agency CAPES (N. C.). We also 
acknowledge the support obtained through the co-operation 
between the Buro de K. F. A.  Julich (FRG) and Unicamp 
Photoacoustic Laboratory (H. V.) in the form of the donation 
of the Commodore microcomputer system. 
1. 
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References 
Foy, C. D.,  in Carson, E. W., Editor. “The Plant Root and Its 
Environment,” University Press, Charlottcsville, VA, 1974. 
p. 601. 
Rhue, R. D., Grogan, C. O., Stockmeyer, E. W., andEverett,  
H. L., Crop Sci., 1978, 18, 1063. 
Garcia, 0.. and Silva, W. J . ,  Cienc. Cult. (Brazil), 1979, 31. 
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Magnavaca, R. ,  f h D  Thesis, University of Nebraska, 1982. 
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Vargas, H., and Miranda, L. C. M.. Phys. Rep., 1988. 161,43. 
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Prioli, L.. and Silva, W. J . ,  Pfunt Sci. Letf. ,  198.5, 38, 47. 
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da Silva. M. D., Bandcira, I. N., and Miranda, L. C. M.,  
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Paper 9l00433E 
Received January 26th, 1989 
Accepted September 12th, 1989 
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Photoacoustic Assessment Of The In Vivo Genotypical Response Of Corn To Toxic Aluminium

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Photoacoustic Assessment Of The In Vivo Genotypical Response Of Corn To Toxic Aluminium

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