S.A. DOS SANTOS, R.F. DA SILVA, M.G. PEREIRA, J.C. MACHADO, C.F. MACHADO, F.M. BORÉM, V.M. GOMES AND O.A.O. TONETTI
Dos Santos, S.A., da Silva, R.F., Pereira, M.G., Machado, J.C., Machado, C.F., Borem, F.M., Gomes, V.M. and
Tonetti, O.A.O. (2009), Seed Sci. & Technol., 37, 776-780
Research Note
X-ray technique application in evaluating the quality of papaya
seeds
S.A. DOS SANTOS1a, R.F. DA SILVA1b, M.G. PEREIRA1c, J.C. MACHADO2e, C.F. MACHADO3,
F.M. BORÉM2f, V.M. GOMES1d AND O.A.O. TONETTI2g
1
2
3
Universidade Estadual do Norte Fluminense Darcy Ribeiro (UENF), Av. Alberto Lamego, 2000, 28013-600,
Campos dos Goytacazes, RJ/Brazil
(E-mail: 1a [email protected]; 1b [email protected]; 1c [email protected]; 1d [email protected])
Universidade Federal de Lavras (UFLA), Caixa Postal 3037, 37200-000, Lavras, MG/Brazil
(E-mail: 2e [email protected]; 2f [email protected]; 2g [email protected])
Dow AgroSciences, Rod. Anhanguera km 344, 14680-000, Jardinópolis, SP/Brazil
(E-mail: [email protected])
(Accepted May 2009)
Summary
The objective of this work was that to investigate the application of X-ray technique in evaluating the quality
of papaya seeds and in establishing an experimental protocol. The work was carried out with papaya seeds
of the Brazilian’ hybrid UENF/CALIMAN 01. The seeds were exposed to X-ray inspection and classified
according to their morphological profiles of embryonic and endospermic tissues as revealed by the radiography.
The radiation exposure condition of 20 Kv for 1.5 minutes generated more precise seed images. Four anatomic
classes were selected, potentially related to germination (full seed, empty seed, seed with reduced embryo and
embryo cavity proportion and seed with endosperm and non-visible embryo). X-ray tested seeds were submitted
to the germination test to detect possible associations between seed anatomy and its corresponding seedling, and
non-germinated seed. The results of this work showed an association between the morphological information
provided by the X-ray test and the standard germination test, indicating its potential to be applied on papaya
seed quality assay.
Experimental and discussion
Papaya (Carica papaya L.) is one of the most important tropical fruits that is cultivated
and consumed in tropical and subtropical countries (Chen et al., 1991). Brazil is the
largest papaya producer worldwide and the Brazilian papaya production corresponds to
24% of the world production. Regarding to export, Brazil is in the 3rd position, only
behind Mexico and Malasya (Agrianual, 2006).
Seedlings developed from seeds are the major propagation type used for commercial
purposes. However, despite the technologcal advances, current studies about papaya seeds
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X-RAY TEST IN PAPAYA SEED
germination are not sufficient to elucidate the causes of their non-uniformity and their
long-time periods required to germinate. Researches to elucidate the germination behavior
of papaya seeds have reported contradictory results. Some of them attributed the slow
and irregular germination of papaya seeds to the presence of inhibitor compounds in the
sarcotesta (Chow and Lin, 1991).
Thus, the poor information about physiological, morph-anatomical and genetic
characteristics of papaya seeds restricts the propagation of this fruit crop (Foster, 1943;
Santos 2009). This scenario limits the expansion of any program that requires high-quality
seeds for the species propagation, its conservation and a wide utilization. As a result, it
is necessary to develop evaluation methods of seed quality, emphasizing those methods
which may produce comparable results with standard procedures.
The X-ray test has been routinely used to analyze seeds from different plant species.
The test consists of a radiographic analysis of internal seed structures to detect any
damage or abnormality that would restrict the seed germination. This technique does not
require previous seed treatment and the low radiation level that is absorbed by the seed
neither induces genetic mutations nor affects the germination performance. Besides, it
is a precise, quick, easy to perform and nondestructive method, generating additional
information about seed viability (Simak and Gustafsson, 1953; Chavagnat and Le Lezec,
1985; Bino et al., 1993).
The objective of the inclusion of the X-ray test in the Rules of Seed Analysis (Brasil,
1992; ISTA, 1993) was to complement the information given by the germination test.
The seed radiography allows the visualization of mechanical injuries, damage caused by
insects, cracks or fractures caused by pre- and post-harvest handling (ISTA, 1993; Poulsen
et al., 1998). Moreover, it allows the detection of embryo abnormalities, as well as the
determination of their development stage (Simak and Gustafsson, 1953)
Although the use of X-ray test is increasing and it useful at different stages of the seed
production and utilization, including genetic breeding, no reports about its application on
papaya seeds were found in the literature. The objective of the present work was to verify
the potential of X-ray test in evaluating the quality of papaya seeds and in establishing a
suitable procedure for this species.
Seeds from UENF/CALIMAN 01 Brazilian’ hybrid were analyzed. One hundred seeds
were distributed on a Plexiglas plate with isolated cells. During the radiation exposure, the
plate was placed on a radiographic film and positioned 35 cm from the radiation emitting
source, using the X-ray Faxitron HP, adjusted at 20 kV and with an exposition time
of 1.5 minutes. The exposure conditions were determined in previous assays (data not
shown). The radiographic films were developed and the radiographic images were used
for seed evaluation. Seeds were classified according to their morphological profiles from
embryonic and endospermic tissues shown in the radiography. The terminology used for
seed classification was based on those described by Simak (1989). After the X-ray test,
seeds from each category were submitted to germination test and any possible association
between the seed anatomy and respective seed structures was verified.
The germination test was conducted according to the Rules for Seed Analysis (Brasil,
1992). The results were expressed in number of germinated seeds, recorded as normal
seedlings or radicle protrusion, seeds with broken tegument and non-germinated seeds.
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S.A. DOS SANTOS, R.F. DA SILVA, M.G. PEREIRA, J.C. MACHADO, C.F. MACHADO, F.M. BORÉM, V.M. GOMES AND O.A.O. TONETTI
Based on the morphology of the embryonic and endospermic tissues observed in the
radiography (figure 1a-d), papaya seeds were classified as full seeds, empty seeds, seeds
with reduced proportion between embryo and embryo cavity, and seeds with endosperm
and non-visible embryo. The frequency of the different categories identified in the
radiographic analysis is shown in table 1.
Figure 1. Radiographic images of Carica papaya L. seeds classified as: (a) full seed, (b) empty seed, (c) seeds
with reduced embryo and embryo cavity ratio and (d) seed with visible endosperm and non-visible embryo. em,
embryo; end, endosperm; teg, tegument; ec, embryo cavity. Germination response of Carica papaya L. seeds:
(e) normal seedling, (f) radicle protrusion (considered as germinated seed), (g) seed with broken tegument and,
(h) non-geminated seed. Legends: pr, primary root; co, cotyledons; hp, hypocotyls.
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X-RAY TEST IN PAPAYA SEED
Table 1. Carica papaya L. seeds classified by radiographic analysis as full seeds (FS), empty seeds (ES), seeds
with reduced embryo and embryo cavity ratio (E/EC-R) and seeds with non-visible embryo (NVE), and their
respective response to germination test.
Radiographic classes
Germination categories
Total
FS
ES
E/EC-R
NVE
Germinated seeds (normal seedlings)
10
00
01
02
13
Germinated seeds (radicle protrusion)
40
00
00
01
41
Seeds with broken tegument
03
00
20
00
23
Non-germinated seeds
07
01
10
05
23
Total
60
01
31
08
100
Following X-ray analysis and classification papaya seeds were submitted to the
germination test. The categories used to evaluate the germination test were: germinated
seeds, seeds with broken tegument and non-germinated seeds (figure 1e-h). The germinated
seeds were grouped in two classes, according to their germination stage: radicle protrusion
and normal seedling, in which the essential structures are completely developed and
healthy (table 1).
The results of the germination test allowed establishing relationships between the
seeds germination categories and their respective anatomic profiles as determined in
the radiographic test (table 1). Thus, 83.3% of the seeds with morphologically normal
embryonic and endospermic tissue germinated and generated 18.8% normal seedlings
(that is 10 normal seedlings germinated from 53 full seeds).
The results presented in the present work illustrate the potential of the X-ray test to
analyze papaya seeds quality. We found a satisfactory association between the radiographic
analysis and the germination test, indicating that this technique can be applied together
with other methods already used to evaluate papaya seed quality, giving additional
information about seed viability. The penetration of the X-ray is determined by the device
voltage. Both the exposure time and the device amperage regulate the radiographic density
or the darkness degree. The combinations of exposure time and amperage may differ
between species, the X-ray device and the sensitivity of the photographic film (ISTA,
1993). An association between the seed anatomy established with the radiographic test
and its respective germination has already been verified in the following species: tomato
(van der Burg et al., 1994), maize (Cicero et al., 1998; Carvalho et al., 1999), Cupressus
sempervirens (Battisti et al., 2000), Lithraea molleoides (Machado and Cicero, 2003).
For papaya seeds reports about the application of X-ray inspection were not found in the
literature. In this study the radiation exposure at 20 Kv for 1.5 minutes generated well
resolved images for papaya seeds on the photographic film and can be considered a good
set of experimental conditions.
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S.A. DOS SANTOS, R.F. DA SILVA, M.G. PEREIRA, J.C. MACHADO, C.F. MACHADO, F.M. BORÉM, V.M. GOMES AND O.A.O. TONETTI
Acknowledgements
The authors thank to UENF, Fundação de Amparo à Pesquisa do Estado do Rio de
Janeiro/Brazil (FAPERJ), and UFLA.
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