Annals of Botany 63, 601-611, 1989 601

A Comparison of the Low-Moisture-Content Limit to

the Logarithmic Relation Between Seed Moisture and
Longevity in Twelve Species
R. H. ELLIS, T. D. HONG and E. H. ROBERTS
Department of Agriculture, University of Reading, Earley Gate. P.O. Box 236. Reading RG6 2AT, UK

Accepted: 28 November 1988

ABSTRACT
Discontinuities in the relation between seed longevity and seed storage moisture content (%, f.wt basis)
were detected at low moisture contents in the Compositae, Cruciferae, Gramineae and Leguminosac. In all
12 species investigated, relations between seed longevity and moisture content above a critical value were
logarithmic, the value of the viability constant C w (relative sensitivity of longevity to moisture) varying
significantly (P < 0025) amongst the species, from 4-5 in rape (Brassica napus) to 6-3 in mung bean
(Vigna radiata). Ten of the 12 species showed no significant variation in longevity with further reduction
in moisture content below a critical value, but in rice (Oryza saliva) and mung bean significant (P < 0-05),
though only slight, negative and positive logarithmic relations, respectively, were detected between
longevity and moisture contents below the critical value. The critical seed moisture content varied between
2-4 % in niger (Guizotia abyssinica) and 6-2 % in pea (Pisum sativum). Comparison between two cultivars
of timothy (Phleum pratense) showed estimates of C w and the critical moisture content to be invariant
within the species. Variation in critical moisture content amongst species was correlated (P < O01) with
variation in the value of C w . The equilibrium relative humidity, and thus the water potential of the seeds
at their various critical moisture contents, was similar, mean 106% r.h. (s.e. 03). Accordingly, inter-
specific variation in the values of C w and critical moisture content is ascribed to differences in seed
composition. Moreover, in each species the critical moisture content coincided with the inflection point of
the isotherm between seed moisture and equilibrium relative humidity. It follows that variation in longevity
with variation in seed moisture content above the critical value results from variation in the weakly bound
fraction of the water held within the seeds, while it appears that removal of thetightlybound water fraction
does not influence the rate of seed deterioration. Above the critical moisture content we detected no
significant difference (P > 025) in the relative sensitivity of seed longevity to change in equilibrium relative
humidity amongst the 12 species, this being equivalent to a doubling of longevity for each 8-4% reduction
in equilibrium relative humidity, but further work is required to corroborate this tentative conclusion.

Key words: Compositae, Cruciferae, Gramineae, Leguminosae, seed storage, seed longevity, seed moisture
content, improved viability equation, water relations.
the sccd viabilit
TNTunnnrTinw y constant C w amongst species
IN 1 R O D U U ION ^ 19g8)
(( E 1 U s gt
A lower limit to negative logarithmic relations Cw is the slope of the negative logarithmic
between seed longevity and seed moisture content relationship between seed longevity and moisture
has been demonstrated in several species (Ellis, content in the viability equation
Hong and Roberts, 1988; Roberts and Ellis, 1989). _ „ __ /1(V t B -c wto , Ii --c B «-c Q / t m
For seeds of quinoa (Chenopodium quinoa Wil\d.), *• p/ ' ( )

sunflower (Helianthus annuus L.) and linseed
(Linum usitatissimum L.) in hermetic storage at
65 °C, the lower moisture limit, below which
further reduction in seed moisture content did not
influence longevity, was 4 1 , 20 and 2-7% (f.wt
basis), respectively (Ellis el al., 1988). We noted
then the possibility of an association between
variation in this value and variation in the value of
where v is probit percentage viability after p days
storage at m% moisture content (f.wt basis) and
/ °C, A, is a constant specific to the seed lot, and KE,
Cw, CH and CQ are species constants (Ellis and
Roberts, 1980). The power term of Eqn (1) is the
s.d., a-(d), of the distribution of seed deaths in time
(Ellis and Roberts, 1980). It is the time in days for
viability to fall by one probit - from 97-7 % to

0305-7364/89/060601 + 11 $03.00/0 © 1989 Annals of Botany Company

24 BOT 63
602 Ellis et al.—Seed Longevity and Moisture

841%, for example-and is thus a measure of and Leguminosae) constant-temperature-oven

seed longevity. At a single constant temperature methods prescribed by the International Seed
relations between this parameter of seed longevity Testing Association (1STA), in which two 4-5 g
and seed storage moisture content can be written samples are dried in a mechanically ventilated
as ovenatl03±2 o Cforl7±l hor 130-133 °Cfor2 h,
respectively (ISTA, 1985 a, b). Rice, pea and mung
0o- = * - C w log l0 m, (2) bean seeds were ground before moisture-content
where 10K is the (extrapolated) value of a at 1 % determinations. Ten to 15 subsamples, each of the
moisture content and the temperature chosen (i.e. size shown for germination tests in Table 1, of each
K=KB-CHt-C t<). lot at each moisture content were then sealed in
Although the value of C w differs amongst species laminated aluminium-foil packets and subse-
(Ellis and Roberts, 1981; Ellis, Osei-Bonsu and quently stored in an incubator maintained at
Roberts, 1982; Ellis, 1988; Ellis et al., 1988), a 65±0-5°C. This high storage temperature was
preliminary, tentative comparison of two species chosen to ensure that complete seed survival curves
between which Cw differed substantially indicated could be obtained at all moisture contents within
that differences in the relative sensitivity of seed a comparatively short period (12 months).
longevity to moisture content could result from Samples were removed from storage at intervals
differences in the hygroscopic equilibria of the varying from 5 min to 35 d, depending on species
seeds, since no difference could be detected in and moisture content for periods up to 350 d.
relations between longevity and equilibrium rela- Before testing these samples for germination, the
tive humidity, and hence water potential (Roberts seeds of all species except niger were humidified
and Ellis, 1989). Here we consider whether limits above water at 20 °C to avoid the possibility of
to negative logarithmic relations between seed imbibition injury during rehydration (Ellis, Hong
longevity and moisture can be detected at low and Roberts, 1985; Ellis, 1987). The durations of
moisture contents in 12 diverse species, six from these treatments, and the subsequent seed moisture
the Gramineae and two species each from the contents immediately before germination tests, are
Compositae, Cruciferae and Leguminosae, and shown in Table 1. No humidification treatment
whether any differences detected amongst the was given to seeds from this lot of niger since
species can be related to water potential. preliminary tests gave no indication of imbibition
injury at the lowest moisture contents investigated.
All samples, together with controls (not stored
MATERIALS AND METHODS at 65 °C) for each moisture content, were then
Seeds of two species of Compositae, niger (Guizotia tested for germination in the regimes shown in
abyssinica (L.f.) Cass.) and lettuce (Lactuca saliva Table 1. Seeds were either tested on top of two
L.), two species of Cruciferae, mustard (Brassica filter papers (Whatman 181) moistened with 4-5 ml
juncea (L.) Czern. & Coss.) and oilseed rape of deionized water in 90 mm Petri dishes (TP in
(Brassica napus L.), six species of Gramineae, Table 1), or between rolled paper towels (Bowater
finger millet {Eleusine coracana (L.) Gaertn.), teff T22) moistened with deionized water (BP in
(Eragrostis tef(Zucc.) Trotter), rice (Oryza sativa Table 1). The testas of pea and mung bean seeds
L. subsp. indica Kato), pearl millet (Pennisetum which had not imbibed after 7 d were chipped and
americanum Auth.), timothy (Phleum pratense L.) returned to tests. Most germination-test tempera-
and foxtail millet {Setaria italica (L.) Beauv.), and ture regimes (Table 1) were those prescribed by
two species of Leguminosae, pea (Pisum sativum the ISTA (19856) with the following exceptions:
L.) and mung bean (Vigna radiala (L.) Wilczek), niger, where no prescriptions were available;
were selected for investigation. Cultivar details are lettuce, where a slightly lower temperature was
provided in Table 1. All seed lots had been stored applied for a longer duration with light applied to
at — 20 ± 1 °C with the moisture contents shown in avoid potential problems from thermodormancy;
Table 1 between production or receipt and the and rice, where the temperature regime recom-
beginning of this investigation. mended from our earlier studies in this crop (Ellis,
Seed moisture contents were adjusted from their Hong and Roberts, 1983) was applied. In general,
initial values (Table 1) before experimental storage the durations of the germination tests (Table 1)
by drying over regularly regenerated silica gel at were slightly longer than those prescribed by the
20 °C, or by humidification above water in a closed ISTA, in order to allow sufficient time for the
container at 20 °C. After equilibrating for 1-3 d at development of seedlings from stored seeds (par-
3-5 °C in sealed containers, seed moisture contents ticularly where hardseededness was encountered).
(f.wt basis) were then determined by the low All seedlings were evaluated according to the
(Compositae and Cruciferae) or high (Gramineae criterion of normal germination (ISTA, 1985 a, b).
T A B L E 1. Information on cultivars, initial seed moisture contents and the regimes selected to humidify and germinate seeds of the 12 species studied

Humidification Germination test details

No of
Cultivar Initial Final No of repli-
name or moisture moisture seeds cates
Family and accession content Duration content per per Temperature Duration
species no. (%, f.wt) (d) (%, f.wt) test test Medium' (°C) (d)

Compositae
G. abyssinica Ginchi 6-9 — — 200 5 TP 20/30 (16 h/8 h) 10
L. saliva Trocadero Improved 5-5 1 20-25 420 6 TP 15 14
Cruciferae
B.juncea White Agriculture 6-1 1 16-20 240 4 TP 20/30 (16 h/8 h) 10
B. napus Jet Neuf 4-3 1 16-20 240 4 TP 20/30 (16 h/8 h) 10

Gramineae
E. coracana PR 202 140 0-25 13-15 400 6 TP 20/30 (16 h/8 h) 14
E. tef DZ 01-787 11-3 1 25-30 450 6 TP 20/30 (16 h/8 h) 14
0. saliva
P. amcricanum
BPI 76NS
IKMV 8201
11-0
7-0
1
1
15-17
14-16
230
400
6
6
TP
TP
34/11
20/30
(16 h/8 h)
(16 h/8 h)
14
14 I
P. pratense S325 13-7 2 25-30 440 6 TP 20/30 (16 h/8 h) 14
Erecta 12-5 2 25-30 440 6 TP 20/30 (16 h/8 h) 14
S. ilalica I.Se 702 12-9 2 18-20 400 6 TP 20/30 (16 h/8 h) 14

Leguminosae
P. satiuum Feltham First 13-6 4-6 18-20 320 5 BP 20 28
V. radiata — 10-8 2-3 12-15 330 5 BP 20/30 (16 h/8 h) 14

* Described in text.
604 Ellis et al.—Seed Longevity and Moisture

Further subsamples of seeds at the various of the 13 lots, i.e. at higher moisture contents
moisture contents were taken to determine the longevity declined significantly with increase in
relation between seed moisture content and equi- moisture. The appropriate values of the constants
librium relative humidity for each lot. Equilibrium K and Cw of Eqn (2) derived from these analyses,
relative humidities were determined at 20 ±0-5 °C which describe the fitted lines of negative slope at
using a Novasina Humidat IC 1, previously cali- the higher moisture contents in Figs 1 and 2, are
brated at 12, 55 and 94% relative humidity usin<; given in Table 2. The open symbols around 15%
appropriate standard salts. moisture content for niger and lettuce (Fig. 1A
and D) indicate observations excluded from
analysis. These were omitted in view of the
RESULTS AND DISCUSSION possibility that these estimates were at moisture
The curves showing the decline in percentage contents above the upper moisture content limit to
normal germination against period of storage were the improved viability equation, which is generally
sigmoidal for each seed lot at each moisture low in oily seeds; for example, the value is about
content, as is normal in such studies (Ellis and 15% moisture content in lettuce (Roberts and
Roberts, 1981). Equation (1) describes such seed Ellis, 1989).
survival curves by negative cumulative normal In all but two species there was no evidence that
distributions with a common origin (which indi- longevity was altered by change in moisture
cates the value of the seed lot constant, KJ at zero content at the low moisture contents. In niger,
time. Accordingly, negative cumulative normal mustard and oilseed rape only two observations
distributions constrained to a common origin for were available in each case at moisture contents
each seed lot were fitted to the seed survival data below the critical value. Nevertheless it is clear
at all moisture contents by probit analysis. The that each pair of estimates of <r did not differ (Fig.
resultant estimates of K, for each seed lot are 1 A, B, E). In the remaining seed-lots more than
shown in Table 2. two observations were available below the critical
moisture content. In all but those of rice and mung
bean no significant response of longevity to change
Relations between longevity and moisture content in moisture within this range was detected (P > 0-1
The estimates of the value of <x at each seed for lettuce and pea, P > 0-25 for finger millet, teff,
storage moisture content provided by probit pearl millet, both timothy lots and foxtail millet).
analysis for species which are members of the Accordingly horizontal lines are shown in Figs 1
Compositae, Cruciferae and Leguminosae are and 2 for these 10 species to describe relations
shown in Fig. 1, while those for the Gramineae are between seed longevity and moisture content below
shown in Fig. 2. Clearly over much of the range of the critical value. These results at moisture contents
moisture contents investigated the results conform below the critical value conform with the pattern
to Eqn (2), i.e. a negative logarithmic relation detected previously (Ellis et al., 1988). In contrast,
between seed longevity and percentage moisture for mung bean and rice significant negative
content. But in each case there is a break in the (P < 0-05) and positive (P < 0-05) relations bet-
relationship at a relatively low moisture content, ween longevity and moisture were observed below
the value of which varies amongst species. Esti- the critical value. Thus lines of negative and
mates of the lower moisture content limit to the positive slope are shown in Figs IF and 2C,
relations described by Eqn (2) were determined for respectively. We are not able to explain why, in
each lot as follows. contrast to the other species examined to date,
The results of longevity at all moisture contents significant responses were detected at low moisture
were first analysed within a single set for each lot contents in these two species or whether significant
in accordance with Eqn (2) by linear regression responses will be detected in other lots of each
analysis. Data were subsequently analysed in two species, but clearly, even in these lots, there is very
separate sets for each lot, data being apportioned little effect of moisture below the critical moisture
between the two sets (l° w c r and higher moisture content; i.e. the fitted slopes (Table 2) are shallow
contents) at arbitrary junctions until the overall in comparison with those at greater moisture
residual deviance had been minimized. Dividing contents. Note that although preliminary obser-
the data in this way provided a significant decrease vations of incomplete data in lettuce gave rise to
in the residual deviance for each seed lot (P < the possibility of a slight increase in longevity with
O005, except P < 0-025 for rice, and P < 0-01 for further reduction in moisture below the critical
foxtail millet). As expected, the response of seed value (Roberts and Ellis, 1989), analysis of the full
longevity to change in moisture content at the results shows that lettuce seed longevity was not
higher values was significant (P < 0-005) for each influenced significantly by reduction in moisture
 Ellis et al.—Seed Longevity and Moisture 605

TABLE 2. Relations between longevity (<r, d) and moisture (m,% f.wt) above and below the critical
moisture content {m^, and the value of the seed lot viability constant (AT,) of Eqn (l)for 13 seed lots from
12 contrasting species

Relations between longevity {or) and moisture (m)

Below• m c Equilibrium
m
c lcutuvc
Above m
Family and Intercept mc
Slopef (%,f.wt) humidity
species (s.e.) (s.e.) (s.e.) (±s.e.) at m^'/o)

Compositae
G. abyssinica 2-22 1 62 n.s. 2-38 9-8 3-42 4-78
(0-03) (002) (2-24-2-53) (014) (017)
L. saliva 2-22 1-64 n.s. 2-56 9-8 3-78 5-25
(002) (005) (2-45-2-67) (009) (Oil)
Cruci ferae
B. juncea 2-57 1-56 n.s. 2-82 102 361 4-56
«M>3) (001) (2-58-3O8) (018) (O20)
B. napus 1-98 1-50 n.s. 2-85 120 3-56 4-54
(0-02) (001) (2-70-3-01) (Oil) (013)
Graminae
E. coracana 2-26 1-76 n.s. 415 9-8 4-90 508
(002) (001) (3-70-4-65) (027) (028)
E.tef 1-89 201 n.s. 4-51 9-9 5-94 6-01
(001) (003) (3-90-5-22) (O40) (042)
O. saliva 2-63 1-70 O70 4-43 1O2 4-51 503
(003) (007) (016) (407^1-83) (014) (015)
P. americanum 1 21 1-60 n.s. 408 11-5 4-57 4-86
(0O1) (005) (3-81^4-36) (014) (015)
P. pralense 1-20 1-33 n.s. 3-62 9-8 3-98 4-75
cv. S325 (001) (003) (3-30-3-96) (O20) (020)
cv. Erecta 1-76 1-57 n.s. 3-61 9-8 4-22 4-75
(001) (002) (3-53-3-68) (008) (020)
S. ilalica 1-70 1-54 n.s. 4O0 106 4-52 4-95
(Ml) (0O1) (317-5O6) (054) (0-58)
Leguminosae
P. sal arum 1-50 1-43 n.s. 6-20 110 5-70 5-39
(002) (002) (4-84-7-95) (063) (061)
V. radiala 1 86 103 -099 6-04 130 6-70 6-27
(002) (O12) (021) (5-28-6-92) (041) (041)

* Eqn (1).
t n.s. indicates no significant response of longevity to moisture below mc.
t Eqn (2).

content below the critical value. It would seem
prudent, therefore, to maintain an open mind for
the present as to the form of the relations between
seed longevity and moisture below the critical
values in rice and mung bean, particularly as in
each case the relations shown were only just
significant at P = 0-05.
Earlier studies above these critical values have
shown that, although Cw is invariant amongst lots
within a species, estimates of its value can differ
significantly even amongst related species (Ellis,
Osei-Bonsu and Roberts, 1982), and that the
variation amongst contrasting species can be
considerable (Ellis, 1988). The results from this
study support these conclusions. The value of Cw
did not differ significantly (P > 0-10) between the
single lots of the two timothy cultivars. Thus Fig.
2E and Table 2 show common slopes for both
cultivars. Comparison of Cw amongst the 12
diverse species in this study showed that its value
606 Ellis et al.—Seed Longevity and Moisture

3 4 6 8 10 16
Moisture content (%, f. wt)

FIG. 1. Relations between seed moisture content (% f.wt, logarithmic scale) and the s.d. of the frequency
distribution of seed deaths in time, <r (days, logarithmic scale), for niger (A), mustard (B), pea (C), lettuce (D), oilseed
rape (E) and mung bean (F), in hermetic storage at 65 °C (•). Table 2 quantifies the pair of solid lines shown for
each species. The critical moisture content is that value where the lines intersect (shown by broken lines). The open
symbols at 14-16% moisture content in A and D were not included in regression analyses, for reasons described in
the text. The dotted line in D indicates predictions based on Eqn (1) and the values of the species viability constants
K%, Cw, CH, and CQ provided by Kraak and Vos (1987).

did differ significantly (P < 0025), i.e. the relative
response of longevity to variation in seed moisture
content differed amongst the species. Seed lon-
gevity in the two crucifers was least sensitive to
differences in moisture, while mung bean was most
sensitive (Table 2).
Although C w did not differ between the two
timothy lots, estimates of the constant K, Eqn (2),
and for the value of <r below the critical moisture
content did (P < 0-025 and P < 0-05, respectively).
This does not agree with the observation upon
which the improved viability equation was based:
i.e. that, for a given seed storage environment, the
value of <r does not vary amongst seed-lots and
genotypes within a species (Ellis and Roberts,
1980). However, seed lots of forage grasses arc
rarely homogeneous, often both immature and
overripe seeds may be present, and so this aspect
of the viability equation may not be appropriate
for such species.
The estimate of 5-25 for Cw for lettuce (Table 2)
is slightly greater than that of 4-80 determined by
Kraak and Vos (1987), and substantially greater
than the 3-84 reported by Rao, Roberts and Ellis
(1988). Rao et al. (1988) acknowledged that Kraak
and Vos's estimate was likely to be the more
accurate since it has been determined from many
more observations; and so it has now proved.
Moreover, and as indicated in a preliminary
analysis on data from incomplete survival curves
(Roberts and Ellis, 1989), the values of the four
species viability constants of Eqn (1) (i.e. KB, Cw,
CH and CQ) determined by Kraak and Vos (1987)
on two different cultivars provide an extremely
robust prediction of the relations between lon-
gevity and moisture contents above the critical
value reported here for a seed lot of yet another
cultivar (Fig. 1D). Similarly, the estimate of 5-39
for Cw for pea (Table 2) is in line with earlier,
tentative, estimates of between 4-8 and 50 (Ellis
and Roberts, 1982), while that of 4-75 for timothy
is similar to an estimate of 5-27 provided by R. D.
Smith (personal communication).
The lower moisture limits to the standard 'air
dry' negative logarithmic relations between seed
longevity and moisture content were then esti-
mated for each seed-lot by determining the point
of intersection of each pair of regression lines
describing relations at lower and higher moisture
contents. These estimates of the critical moisture
content vary considerably; from 2-38% in niger
to 6-2% in pea (Table 2). Examination of the
estimates for the two timothy cultivars shows
precise agreement at 3-6 % moisture content (Table
2), supporting the assumption that the lower limits
to logarithmic relations between longevity and
moisture are characteristic of a species. R. D.
Smith (personal communication) found that at
 Ellis et al.—Seed Longevity and Moisture 607

«B
100 - • — • - •

10
I
O'l
o-oi ,

100

10

0-1
0-01
1
> J - / * - •' <*

3 4 6 8 10 16 I 2 34 6 8 10 16
Moisture content (%, f. wt)

FIG. 2. Relations between seed moisture content (% f.wt, logarithmic scale) and the s.d. of the frequency distribution
of seed deaths in time, er (days, logarithmic scale), for finger millet (A), teff (B),rice(C), pearl millet (D), timothy (E)
and foxtail millet (F), in hermetic storage at 65 °C (#). In E results are presented for two timothy cultivars, S325
( • ) and Erecta ( • ) . Table 2 quantifies the pair of solid lines shown for each seed-lot. The critical moisture
content is that value where the hnes intersect (shown by broken lines).

62 °C the logarithmic relation between timothy
seed longevity and moisture content extended
down to 3 % , the lowest moisture content he
investigated. Although this is slightly lower than
the critical value estimated here, we believe these
differences are reconcilable, since several estimates
of longevity at moisture contents well below the
critical value are required in order to provide a
reliable estimate of the critical value.
The lowest estimates of critical moisture content
were obtained in niger and lettuce (Table 2). There
is good evidence to support our conclusion that
lettuce seed longevity can be greatly extended by
desiccation to comparatively low seed moisture
contents. Lettuce seeds can withstand desiccation
to 1% or, even, 0 4 % moisture content without
injury to germination (Nutile, 1964), and des-
iccation to such low moisture contents is not
deleterious to longevity (Kosar and Thompson,
1957; Nutile, 1964). One problem with such studies
has been that the maximum storage periods (4 or
5 years) at the ambient or cool temperatures
employed (10-30 °C) have not been sufficient to
distinguish between longevity at different low
moisture contents (Kosar and Thompson, 1957;
Nutile, 1964). Nakamura (1975), however, stored
lettuce seeds for 20 years at ambient temperatures
and this period was sufficient to demonstrate that
longevity at 2-5% moisture content was greater
than that at 6-3 % moisture content. His results for
pea and timothy stored at ambient temperatures
for 9 and 13 years, respectively, are less supportive
of this study's conclusion that very low moisture
contents (those in equilibrium with around 10%
r.h.) are not damaging to longevity but, as he
acknowledged, his germination test procedures
took no account of the possibility of Imbibition
injury (Nakamura, 1975).
The estimates of a at the critical moisture
content, as well as at lower moisture contents for
all species but rice and mung bean, fall within the
range 20-100 d (Figs 1 and 2). Although the
variation in this value within the Gramineae is
considerable, that within each of the Compositae
608 Ellis et al.—Seed Longevity and Moisture
FIG. 3. Comparison of the estimates of the critical
moisture content (%, f. wt) and C w , the relative sensitivity
of seed longevity to moisture above the critical value,
amongst contrasting species. The open symbols show
estimates derived from an earlier study (Ellis etaJ., 1988).
The regions A, B, C and D bounded by broken lines
identify estimates for species from Compositae, Cruci-
ferae, Gramineae, and Leguminosae, respectively. The
symbols E and F are for species from Linaceae and
Chenopodiaceae, respectively.
and Cruciferae is negligible (42-44 d and 32-36 d,
respectively). These estimates are similar to those
obtained for linseed, quinoa and sunflower (Ellis
et al., 1988). We noted then that those differences
were comparatively minor, However, given the
variation within the Gramineae, these similarities
in longevity at the critical moisture content are
probably of little practical consequence.
We also speculated that the critical moisture
content may be positively associated with the
value of Cr (Ellis et al., 1988); i.e. the greater the
sensitivity in the variation of the seed longevity of
a species to variation in moisture, the greater the
critical moisture content. Analysis of the results
from both studies, i.e. comparing 16 seed lots of 15
species, does show a significant correlation between
the critical value and C^ (r = 0727, for 14 d.f.,
P < 0-01). This conclusion is also valid if the
analysis is limited to the results from this study
alone (r = 0-698 for 11 d.f., P < 0-01).
To some extent the correlation is associated
with taxonomic differences. The two species from
Leguminosae show the highest critical moisture
and their longevity is the most sensitive to
differences in moisture content, whereas the species
from Compositae and Cruciferae show much lower
critical moisture contents and their longevity is
considerably less sensitive to moisture (Fig. 3).
Indeed it is possible to extend taxonomic associa-
tions of the differences amongst species further by
considering the variation in critical moisture
content shown in Fig. 3 and Table 2 within the
Gramineae: the values for finger millet and teff,
both within the tribe Eragrostideae, are very
similar; similarly the values for pearl and foxtail
millets, both within the tribe Paniceae, are very
similar, but slightly lower than those for the
former. However, it may not be possible to extend
these associations too far. For example, timothy
and teff are both within the subfamily Pooideae
and yet these species showed the greatest variation
in critical moisture content within the Gramineae.
Moreover, the fact that seed composition affects
its water potential at a given seed moisture content
provides a more direct explanation of differences
in critical moisture content (Roberts and Ellis,
1989), although these characteristics will, of course,
tend to be correlated with taxonomic differences.
Relations between longevity and equilibrium relative
humidity
Variation in the value of the critical moisture
content amongst species may also be negatively
associated with seed oil content (Ellis et al., 1988).
Oil content is a major factor influencing relations
between seed moisture content and equilibrium
relative humidity (e.g. Cromarty, Ellis and
Roberts, 1982). These relations, often described as
isotherms, are shown in Figs 4 and 5 for each seed
lot over the same moisture content ranges as those
employed in the longevity study. For practical
reasons these isotherms were determined at 20 °C,
rather than the temperature of 65 °C chosen for
the storage longevity investigation. Each curve
represents a combination of absorption and de-
sorption isotherms; at moisture contents above
the initial value shown in Table 1 they represent
absorption isotherms; whilst at moisture contents
below the initial value they represent desorption
isotherms. In this way the form of the equilibria
shown in Figs 4 and 5 should correspond to the
conditions which applied in the longevity experi-
ments, although the height of the curve would
have been different in the latter (see below).
Isotherms of this kind are typically sigmoidal,
comprising three distinct regions: one at low seed
moisture contents where moisture content rises
steeply for a small increase in relative humidity; a
second at intermediate moisture contents where
the gradient is shallow; and a third region at high
moisture contents where seed moisture content
again increases more steeply for a small increase in
relative humidity. (This latter region is missing
 Ellis et al.—Seed Longevity and Moisture 609

Equilibrium relative humidity (%)

FK3. 4. Relations between seed moisture content (%, f.wt) and the equilibrium relative humidity (%) of seeds of niger
(A), mustard (B), pea (C), lettuce (D), oilseed rape (E) and mung bean (F), at 20 °C (O)- The curves shown are
absorption and desorption isotherms above and below, respectively, initial seed moisture contents (Table 1). The
broken horizontal lines indicate the critical moisture contents determined in the seed longevity study.

16
12
8
4
0

16
12
8
4
0

16
12
8
4
0
20 40 60 80 0 20 40 60
Equilibrium relative humidity (%)

FIG. 5. Relations between seed moisture content (%, f.wt) and the equilibrium relative humidity (%) of seeds of finger
millet (A), teff (B), rice (C), pearl millet (D), timothy (E), and foxtail millet (F), at 20 °C (O)- In E results are
presented for two timothy cultivars, S 325 (O) and Erecta ( • ) . Further details as Fig. 4.
610 Ellis et al.—Seed Longevity and Moisture
from the isotherms shown in Figs 4 and 5 as very
high seed moisture contents were not included in
this study.) The major influence of temperature on
these isotherms is on the height of the second
region; this is reduced the warmer the temperature
(e.g. Cromarty et al., 1982). Similarly, the major
difference amongst seed lots within a species is also
the height of the central region - as Fig. 5E shows
for the two timothy cultivars.
These three regions are also largely indicative of
the three categories of bound water that occur in
tissues: water bound at strong sites, water bound
at weak sites, and water bound to other water
molecules already absorbed by multimolecular
sorption (D'Arcy and Watt, 1970). Over the first
region the water within seeds is dominated by the
strongly bound fraction, whilst in the third region
the increase in water within the seed is dominated
by multimolecular water (Vertucci and Leopold,
1987).
In contrast to the variation detected amongst
species in critical moisture content, the equilibrium
relative humidities of these values - interpolated
from the isotherms shown in Figs 4 and 5 - show
little variation amongst species (Table 2); the
mean estimate for all 12 species is 10-6% r.h.
(s.e. 0-3), or roughly — 350 MPa. Moreover, not
only does this value hardly vary amongst the
species, but it is also quite clear that the critical
moisture content coincides in all 13 seed lots with
the point of inflection on the isotherms (Figs 4 and
5), and which represents the distinction between
the first and second types of bound water. Clearly
then, the discontinuities in the logarithmic relations
between longevity and moisture at low moisture
contents shown here occur because of a change in
the category of bound water being removed by
desiccation; subsequent longevity in storage only
increases with further desiccation until all of the
water bound at weak sites is removed. This then
appears to implicate only the water bound at weak
sites as the cause of variation in longevity at
different moisture contents in 'air-dry' storage.
If the differences in the value of the critical
moisture content amongst these 12 species can be
explained in terms of differences in the hygroscopic
equilibria of the seeds, then consideration should
be given as to whether the variation in Cw amongst
the 12 species (Table 2) can be similarly explained.
A recent comparison of barley (Hordeum vulgare
L.) and lettuce, which have contrasting isotherms
and contrasting sensitivities of seed longevity to
percentage moisture content, showed the relative
effect of changes in equilibrium relative humidity
on seed longevity to be more or less identical
(Roberts and Ellis, 1989). The equation used in
that analysis incorporated the temperature terms
shown in Eqn (1), but here, since all observations
are at a single temperature, we can modify Eqn (2)
as follows:
Iog19<r = K^-C^rh, (3)
where rh is equilibrium relative humidity, C^ is the
sensitivity of longevity to change in equilibrium
relative humidity, and K^ is a constant (the
extrapolated estimate of log10 a at 65 °C and 0 %
relative humidity). When Eqn (3) was applied it
was found that C rt (i.e. the relative response of
longevity to a 1 % change in equilibrium relative
humidity) did not differ significantly (P > 0-25)
amongst the 12 species. Moreover, the common
estimate of C rt derived for the 12 species from this
analysis was 0-0358 (s.e. 0-0011), similar to that of
0-0376 determined in the earlier comparison
between barley and lettuce (Roberts and Ellis,
1989). The value of the coefficient C^ determined
here is roughly equivalent to a doubling of
longevity for each 10% reduction in equilibrium
relative humidity (the actual factor for a 10%
reduction in r.h. is 2-28, or a doubling of longevity
for an 8-4% decrease in r.h.). However, although
the concept of an identical response of seed
longevity to change in equilibrium relative hu-
midity, and thus to water potential (Roberts and
Ellis, 1989) amongst contrasting species is ap-
pealing, we do not suggest the evidence is
conclusive. The reason for our caution is as follows.
In contrast to our earlier study where relations
between the logarithm of longevity and equilibrium
relative humidity were essentially linear, in this
study (which included lower equilibrium relative
humidities) there was a tendency for these relations
to deviate from linearity (towards sigmoids). As a
consequence the residual variance following the
fitting of Eqn (3) was much greater than that for
Eqn (2), where no consistent departures from
linearity were detected (Figs 1 and 2), the pro-
portion of the variance of log10 a attributed to its
regression on rh or log10 m being reduced to 0-945
from 0-985 (variant slopes in both cases). And so
the conclusion that the relative sensitivity of
longevity to equilibrium relative humidity does not
differ significantly amongst species, can, at present,
only be tentative since it is based on a com-
paratively high error mean square.
One practical objective of our studies is to
determine to what extent seed storage at very low
moisture contents could be employed for genetic
resources conservation by long-term seed storage,
and in particular whether further desiccation below
about 5 or 6% moisture content could reduce or
avoid the requirement for refrigeration (IBPGR,
1985; Ellis, Hong and Roberts, 1986). Although
several questions need to be resolved concerning
 Ellis et al.—Seed Longevity and Moisture
the genetic stability of very dry seeds in storage
(Rao et al., 1987, 1988), these results and others
(Ellis et al., 1986, 1988) indicate that this 'ultra-
dry ' storage could be feasible for species with oily
seeds which tend to have comparatively poor
longevity, but less so for species with starchy seeds
(where longevity is in any case normally fairly
good). Note that ultra-dry storage is unlikely to be
hazardous since desiccation below the critical value
is unlikely to be deleterious (Figs 1 and 2) - and
even in mung bean only slightly so. Moreover,
since the critical moisture content coincides with
the inflection of point of the desorption isotherm
(cf. Figs 1 and 2 with 4 and 5, respectively), over-
drying is unlikely to be a practical problem. Whilst
we cannot yet say whether the critical moisture
content varies with storage temperature, the fact
that it coincides with this inflection point — which
is comparatively unaffected by temperature -
suggests that any variation with storage tem-
perature is likely to be minor. Clearly, however,
further work is required to verify this assumption.
ACKNOWLEDGEMENTS
We thank the International Board for Plant
Genetic Resources (IBPGR) forfinancialsupport;
Dr T. T. Chang, International Rice Research
Institute (IRRI), Los Banos, The Philippines, for
providing'rice seeds; Dr L. Fussell, Sub-Sahelian
Center of the International Crops Research In-
stitute in the Semi-Arid Tropics (ICRJSAT),
Niamey, Niger, for providing pearl millet seeds;
Dr K. L. Mehra, National Bureau of Plant Genetic
Resources, New Demi, India, for providing seeds
of finger and foxtail millets; and Dr M. Worede,
Plant Genetic Resources Centre, Addis Ababa,
Ethiopia, for providing seeds of niger and teff.
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