-
--
Nayal, 1.S., Thapliyal, R.C., Phartyal, S.S. and Joshi, G. (2002) Seed Sci. & Techno/., 30, 629-639
Germination and storage behaviour of seeds of Grewia optiva (Tiliaceae) - a sub-tropical Himalayan multipurpose evergreen tree J.S. NAYAL, R.C. THAPLlYAL, s.s. PHARTYAL AND G. JOSHI Forest Tree Seed Laboratory, Silviculture India (E-mail:
[email protected])
Division, Forest Research Institute (lCFRE) Debra Dun, Uttaranchal,
(Accepted October 2001)
Summary The Grewia optiva seed has a bony, hard seedcoat that does not appear to restrict the germination of seed and hence does not need any pre-treatment. The temperature around 30°C and BP media had significantly better effect on the germination and other parameter. The seeds stored with high moisture content (17%) in airtight container maintained viability for longer periods as compared to vacuum container. However, the seeds desiccated to 5% mc maintained better storability in vacuum container than the airtight container. Seed mc plays a significant role in storage environment. The reduction in seed mc down to 5% resulted in a significant increase in Pso in airtight as well as in vacuums containers at all temperatures. On the other hand temperature indicates negative logarithmic relation with seed longevity. The data recorded for the relationship between temperature and seed mc during storage was fitted to the basic viability equation and constants were calculated which besides proving the orthodox nature of seed of this species can predict seed longevity in a given condition of seed mc and storage temperature for a particular seed lot.
Introduction
Grewia optiva (Drumn.) is a highly valued multipurpose tree yielding green cattle fodder, furniture, medicine, paper, fiber and dyes. This species has been obliterated from the natural forests and occurs in small patches largely cultivated around homesteads in the outer hills up to 2000m in the central and western Himalayas. The tree is heavily lopped during autumn and following winter at around the same time when the tree is in seeding with the result that there is always acute shortage of good quality seed of this species. There is an immediate need to develop long term preservation of germplasm of this species for sustainable use. The seeds of G. optiva is a single seeded nut, more or less round in shape and of the size smaller than the pea seed. The endosperm is hard and does some restriction to the protrusion of radicle through the seed. Most of the work on the storage of orthodox seeds has been done on soft seed coated species which do not resist the ready movement of moisture in and out of the seed. In G. optiva, the exchange of moisture may be restricted due to the hard endosperm. It was from this point of view that the present investigation on the influence of drying and seed storage temperature on the viability of this species was taken up and the same is reported upon in this paper. 629
J.S. NAYAL, R.C. TIIAPLIYAL,
S.S. PHARTYAL
AND G. JOSHI
Material and methods The fruits were collected at maturity in the month of October, 1996 from Thano Range of Dehra Dun Forest Division (29.98°N latitude and 78.12°E longitude), Uttaranchal and were heaped in the laboratory for a week for softening the pulp. Fruits were depulped by rubbing against a wire mesh and the remaining pulp was removed from the seeds by rubbing with sand followed by thorough washing under running water. Following surface drying for 24 hours under a ceiling fan, moisture content (mc) and initial germination of seed was determined at 103:t 2°C for 17:t 1 hrs (1STA, 1993). Seed pretreatment and germination Detailed investigations were carried out to determine suitable pretreatment and germination test method for the evaluation of seed viability of this species. In the first experiment, seeds were treated for germination with 10 different treatments viz., Soaked in concentrated sulfuric acid for 2,5 and 10 minutes, hot wire scarification, overnight soaking in GA3(100ppm), cold and hot water overnight soaking, mechanical scarificationat the micropyler and cotyledonary end and complete removal of endocarp. A control set was used without any pretreatment. In the second experiment, seeds were subjected to germination tests with 21 different combinations of temperatures (15, 20, 25, 30, 35, 40 and 20/30°C) and media (top of paper, between paper and sand). Seeds were placed on top of paper, in petridishes, or between two layers of paper towels or in sterilized white quartz sand filled in trays in which seeds were sown 1 cm deep. Three replications of 100 seed for the first and of 50 seed for the second experiment were used. Seeds were germinated in cabinets with about 1000 Lux illumination provided during high temperature period by means of cool white fluorescent tubes. Germination was recorded daily and seed with radicle about 1 cm was considered germinated. The last count was made on day 35, after which germination test was terminated. The Mean germination time (MGT) was calculated according to Piotto (1995). Desiccation and storage of seeds Seed with 17% initial mc was dried in a drying cabinet at 30°C and 15%RH to 10 and 5% mc. Seeds with above 3 moisture levels and 5 storage temperatures viz., -5,5, 15, 25°C and ambient room temperature (RT) were stored in air tight plastic and vacuum containers. In all seeds were stored in 31 different treatment conditions including one control set in an open container at ambient room conditions. Moisture content and germination test Seed mc was determined on wet basis at 103 :t 2°C for 17 :t I hours. Seeds were drawn from each storage condition at an interval of about two-month for germination test on top of towel paper in glass petridishes at 30°C. The viability of seed was determined by using three replications of 25 seeds each. Statistical analysis The data on seed germination and MGT of first two experiments and germination during storage was subjected to ANOVA. The rate of deterioration (d-l)was calculated from the
-
GERMINATION AND STORAGE OF SEEDS OF GREWIA OPTNA
survival curves as described earlier (Nayal et ai., 2000). Following probit analysis, half viability period (Pso)was calculated (Roberts, 1973). Results The pretreatment experiment showed no significant effect on germination over and above the control (table I). Soaking seeds in sulfuric acid for 5 minute, in ordinary tap water and in GA3 (lOOppm)had only a slight effect on the germination percentage. The treatments however had significant effect on the time taken to complete germination, Le., the MGT (p=0.05). Treatment TI, T5, T6 and T9 were comparatively more effective in early and faster rate of germination and took minimum time ranging from 13.9 to 15.0 days as compared to treatments (T8) which took as long as 19.48 days, followed by control (TI1). Tables 2 and 3 show that the mean germination per cent and MGT differed significantly between temperature, media and their interaction (p=O.05).The mean value of germination was highest at 30 and 35°e with 23.4 and 21.3% respectively. Other temperatures resulted in significantly poor germination. The temperature 200e had minimum 3.42% germination while at 15°e practically no germination was recorded irrespective of media used. Among media, between paper was the best with a maximum mean of 18.86% germination, closely
Table I. Effect of pretreatment on germination and MGT of G. optiva seed.
--
Treatments
Germination
MGT
TI.
Soaking in sulfuric acid for 2 rnnt.
19.00
15.85
T2.
Soaking in sulfuric acid for 5 rnnt.
37.75
14.83
TI.
Soaking in sulfuric acid for 10 rnnt.
32.25
15.91
T4.
Hol wire scarification
22.75
18.06
T5.
Soaking in GAJ (IOOppm)for 24 hrs.
38.00
13.99
T6.
Soaking in tap water for 24 hrs.
37.25
14.34
T7.
Soaking in hot water for 24 hrs.
33.00
17.81
T8.
Nicking at micropyler end
15.75
19.48
T9.
Nicking at opposite to micropyler end
35.75
15.00
TlO.
Kernel
27.25
16.10
TII.
Without treatment (Control)
36.00
19.32
S.Em.
1.59
0.78
CD 5%
3.24
1.60
CD 1%
4.37
2.15 1'\1
J.S. NAYAL.R.C. THAPLlYAL. S.S. PHARTYAL AND G. JOSHI
Table 2. Effect of temperature and media on germination of G. optiva seed. TP
Temperatures (0C)
BP
SAND
Mean of temperature
15
0.0
0.0
0.0
0.0
20
4.00
5.00
1.25
3.42
25
24.75
23.00
9.75
19.20
30
25.75
30.00
14.50
23.40
35
30.50
27.25
6.00
21.30
40
23.00
23.50
5.00
17.20
20/30
17.25
23.25
10.30
16.90
Mean of media
17.89
18.86
6.68
14.50
S.Em.
CD 5%
CD 1%
-
Media
.92
1.83
2.44
Temp. M*T
1.39
2.79
3.72
2.42
4.85
6.45
Table 3. Effect of temperature and media on mean germination time (MGT) of G. optiva seed. TP
Temperatures (0C)
BP
SAND
Mean of
15
36.00
36.00
36.0
temperature 36.0
20
16.90
16.60
17.6
17.1
25
11.50
12.40
15.6
13.1
30
9.65
9.19
13.9
10.9
35
12.00
9.41
16.5
12.7
40
15.20
14.90
16.7
15.6
20/30
14.10
14.20
17.1
15.1
Mean of media
16.50
16.10
19.1
17.2
S.Em.
CD 5%
CD 1%
Media
0.28
0.56
0.74
Temp.
0.43
0.85
1.13
M*T
0.74
1.48
1.96
;:;':\')
--./
GERMINATION
AND STORAGE
OF SEEDS OF GREWIA
OPTIVA
followed by top of paper (17.89%) while the sand media with minimum 6.68% was the worst irrespective of the temperature used. The interacting effect of temperature and media revealed that 35°C on top of paper had the highest 30.5% germination. The temperature at 20°C in sand media recorded least mean germination of 1.25% which was significantlydifferent from other combinations. The MGT was least 10.9 at 30°C and highest 36.0 at 15°C and significantly different to MGT at other temperatures.At 25, 35,40 and 20/30°C similar and non-significant values of MGT were shown. The MGT in BP and TP took significantlyminimum 16.1to 16.5 days as compared to sand media which took 19.1 days. The interacting effect of temperature and media showed that 30°C in BP had minimum value of MGT Le., 9.19 days, which was closely followed by 9.41 and 9.65 at 35°C in BP and 30°C on TP respectively.The maximum MGT was observed at 15°C irrespective of the media used. Effect of storage temperature and seed mc on longevity The germination data recorded during storage of seed and analysed for variance revealed significant differences among temperature, moisture, container and their interactions. The seed stored in vacuum container recorded mean value of germination (13.8%) somewhat similar to that recorded for airtight container (13.3%). The germination was least in seeds with original moisture content, significantly different to desiccated seeds. Desiccation of seeds to 5% mc had highly significant effect on viability irrespective of the storage temperatures. Seeds desiccated to 10% closely followed this. Among storage temperatures, -5°C had the maximum value of germination significantly different to other temperatures irrespective of seed mc. The storage of seed at 25°C and ambient room conditions had deleterious effect on viability. The interacting effect of mc, temperature and containers revealed that the seed stored in open container at ambient temperature had lost complete viability after around one year. Seeds with original mc stored at 25°C and RT both in airtight and vacuum containers, lost complete viability after 499 days. At 5% mc and -5°C storage temperature in both airtight and vacuum containers, the viability was significantly longer, Le., up to 1107 days with 18.7 and 20.0% germination respectively as compared to other storage conditions. Rate of deterioration (d-I) The survival curves for seeds stored at different combination of moisture, temperature and containers show that the rate of deterioration was less in vacuum container than in airtight container (figure I and 2a,b,c). The rate of deterioration at 17, 10 and 5% mc ranged from 0.0016 to 0.0041, 0.0010 to 0.0023 and 0.0006 to 0.0017 respectively for airtight container whereas the range was 0.0022 to 0.0045, 0.0009 to 0.0018 and 0.0005 to 0.0011 for vacuum container. The rate of deterioration was minimum at 5% mc in airtight container at all storage temperatures except at 25°C which had somewhat deleterious effect on the rate of loss of viability. Interaction of temperature, moisture and container show that -5 and 5°C storage temperature had minimum value of slope at all moisture levels and containers while at 15°C and RT the values of slopes were larger and almost similar irrespective of the seed mc (table 4).
J.S. NAYAL, R.C. THAPUYAL,
45
___25°C
,.~."
35
'\~'\""'" \\ ':,
30
\
25 20
'\ '\.
AND G. JOSHI
_RT
(a)
40
S.S. PHARTYAL
- -- - - - 15°C
5°C 5°C
',>-.
15
"
',:,-, "':'"
'..
\
10
..... ..'....................
'\" ""
5
""'
o o
200
:-..:
'------..-400
'......
-"-..
600
800
45 (b)
40 35 30 c: .Q 1ii c: 'E CD
(!)
25 20 15 10 5 0 200
0
400
600
800
1000
1200
45 (e)
40
".
"" '- -'...
35
c:
0 as c: 'E Qj (!)
, " ...........-. '....-..'"" ," -...........-... ..... "" ---". ,, ,, ,, , '.... '....-....
30 25 20 15 10
-- ----
5 0 0
200
400 600 800 Storage period (days)
1000
1200
Figure I. Survival curve of G.optiva seeds stored with original1? (a), reduced 10 (b) and 5% me (c) in vacuum containers. Curve derived from probit analysis of seed stored at 5 different temperatures.
GERMINATION AND STORAGE OF SEEDS OF GREW/A OPT/VA
45
-RT (a)
40
-__25°C
35
15°C
30
5°C
0
25
5°C
:;c:
20
c:
'E CD
(!)
15 10 5 0 0
200
400
600
800
1000
1200
800
1000
1200
400 600 800 Storage period (days)
1000
1200
45
(b)
40 35 30 25 20 15 10 5
o o
200
45
400
600 (e)
40 35 30 25 20 15 10 5
o o
200
Figure 2. Survival curve of G.optiva seeds stored with original 17 (a), reduced 10 (b) and 5% me (c) in airtight container. Curve derived from probit analysis of seed stored at 5 different temperatures.
J.S.NAYAL,R.C.THAPUYAL.S.S.PHARn'ALANDG. JOSHI
Table 4. Effect of temperature, mc and containers on the rate of loss of viability (d.l) of G. optiva seed. Temperatures
Vacuum containers
Airtight containers
CC)
17%
10%
5%
17%
10%
5%
RT
0.0031
0.0023
0.0010
0.0045
0.0016
0.0011
25
0.0041
0.0016
0.0017
0.0039
0.0016
0.0016
15
0.0029
0.0018
0.0011
0.0039
0.0018
0.0011
5
0.0017
0.0023
0.0011
0.0019
0.0012
0.0011
-5
0.0016
0.0010
0.0006
0.0022
0.0009
0.0005
Half viability period (Pso) The vacuum container had a higher half viability period as compared to airtight container, which had some deleterious effect on the Pso(table 5). At 17, 10 and 5% seed mc the Pso ranged from 144 to 368, 256 to 590 and 347 to 983 days respectively in airtight container whereas varied from 131 to 310, 328 to 666 and 368 to 1180 days respectively in vacuum container. Considering the temperature of storage, at -5°C the Psohad the maximum value iuespective of mc and containers as compared to other storage temperatures. Seeds desiccated to 5% mc have a higher value of Psoas compared to original 17% mc. Table 5. Effect of temperature,
mc and containers on the half viability period (Pso) of G. optiva seed.
Vacuum containers
Airtight containers
Temperatures (0C)
17%
10%
5%
17%
10%
5%
RT
190
257
590
131
369
536
25
144
368
347
151
368
368
15
203
328
536
151
328
536
5
347
256
536
310
492
536
-5
368
590
983
269
666
1180
Relationship between moisture, temperature and seed longevity The relationship between moisture, temperature and longevity in orthodox seeds has been described numerically through a viability equation (Roberts, 1973) which has facilitated the prediction of longevity of above category of seed in storage such aslog Pso= Kv - Clm - C2t
(i)
(where, m=mc (%), t.= temperature (0C)and Ch C2and Kv are constants) relative time taken (in days) for the viability to fall to 50% (Pso)under constant moisture content and temperature conditions was applied and approximate values of constants were calculated and applied to equation (i) as follows:
GERMINATION AND STORAGE OF SEEDS OF GREWIA OPT/VA
Temperature (0C) a 25 b
15
c
25
mc (%) 17 17 5
Pso 144 203 347
Using data from (a) and (c) C, = (log c Pso-log a pso) / mc (a-c) C, = (log 347 -log 144) / (17 - 5) C,=O.0318 Using data from (a) and (b) C2 =(log b Pso - log a Pso)/ temperature (a-b) C2= (log 203 - log 144) / (25 - 15) C2= 0.0149 From equation (i) Kv = log Pso+ C,m + C2t Substituting values for C, and C2 under these conditions the Kv
= 3.0711
Thus the values of viability constants for G. optiva was, CI= 0.0318, C2=0.0149 and Kv = 3.0711.
Discussion
Results of pretreatment of G. optiva seeds did not show any significant effect on germination over and above those without treatments, thus indicating that the hard coat of G. optiva did not restrict the protrusion of radicle or impeded the absorption of moisture from the substratum. The low germination percentage of seed of this species was due to empty seeds as seen by the cutting test of ungerminated seeds. The pretreatments, however, affected the MOT as seeds scarified with sulfuric acid for 5 minute, OA3 or in ordinary tap water progressively reduced the MOT from 19 to around 14 days. Shrestha and Oautam (1989) observed that the seeds soaked in cold and hot water for 24 hours germinated maximum as compared to seed scarified with acid. Sneh Lata et at. (1993) and Singh et at. (1997) have reported more or less similar results. Effect of temperature indicated that the seed germinated well over a wide range of temperatures, from 25 to 40°C but 30°C was the optimal temperature that significantly affects the germination as well as MOT. The lower temperature, 15 and 20°C adversely affected the germination and MOT. Alternating 20/30°C temperature also does not seem to yield good results though, ISTA (1993) recommended alternating temperatures as the best for several tropical species. Among media, BP had significant effect on germination and MOT as compared to sand media. The result of interaction of temperature and media shows that 30°C temperature in BP media is the best for all the parameters.
I.S. NAYAL, R.C. THAPUYAL,
S.S. PHARTYAL
AND G. JOSHI
The longevity of seed is affected by storage temperature, seed mc and the type of container. The effectiveness of a container for storage is dependent on its ability to maintain the original moisture content. The viability of seed was maintained for longer period in vacuum than the airtight container. This shows that the complete removal of oxygen may lead to slow build up of carbon dioxide in vacuum containers. Airtight container with C02 cause early death of the seeds in acorns (Holmes and Buszenwicz, 1956). Storage in airtight container results in a high eOi02 ratio, which has been regarded as favourable for seed longevity in orthodox seed (Goldabach, 1979, quoted in Willan, 1985). For long term storage of orthodox seeds, the most effective method is a combination of moisture proof containers with controlled low temperatures provided by refrigeration. In all types of containers, viability loss was directly proportional to increase in temperature. Similar trend is reported in Hoiopteia integrifolia, where fall in viability was faster at 30°C compared to 5°C (Maithani et ai., 1987). The reduction in mc from original 17 down to 5% leads to significant increase in time taken to decline to 50% viability (Pso)in both type of containers. There was an increase from 144 and 131 to 983 and 1180 days in airtight and vacuum container respectively. The rate of deterioration shows similar trend and exhibits positive relation with mc. As the mc declines from its original, the rate of deterioration decreases simultaneously from 0.0041 and 0.0045 to 0.0006 and 0.0005 in airtight and vacuum container respectively.The trend shows increase in the longevity of seed, which seems to follow the rule of thumb proposed by Harrington (1963, 1970) according to which the life of the seed is doubled for every 1% decrease in mc. In the present case a negative logarithmic relationship of temperature with seed longevity is indicated. As the temperature decreased from 25°C to 5°C and -5°C the time taken to decline the viability to 50% (Pso)increased while the rate of deterioration decreased simultaneously in both the airtight and vacuum containers irrespective of moisture levels used. In addition, temperature affected the rate of deterioration significantly at all seed moisture levels and storage temperature doubled the life of seed. The seed storage data obtained from this study leads to the conclusion that seed falls well within the definition of 'orthodox' (sensu Roberts 1973) with respect to the relationship between mc and temperature and longevity for seeds under constant conditions. Present experiment clearly show that seed viability was maintained for longer periods when mc was reduced from its original 17 to minimum 5% and storage temperature either 5 or -5°e. These conditions were the best for seed storage of this species. The slope of survival curve and rate of deterioration (d-') exhibit similar trend and indicates that rate of loss in viability decreases with decrease in moisture as well as in temperature. This is in accordance with the fact that the longevity of orthodox seeds increases in a specific and predictable way over a wide range of environmental conditions by decreasing storage temperature and mc (Roberts, 1973). The value of viability constant are, e.=0.0318, e2=0.0149 and Kv=3.0711. Since the data indicates orthodox storage physiology and even though the range of storage temperature and seed moisture levels were not large enough, the available data was used to calculate the viability constants to predict the longevity of seed in the given conditions of storage. The tentative information can be helpful for further work on the development of improved viability equation for predicting the viability of seed of this species. Ellis and Roberts (1980) used the same equation for tentative estimation of potential longevity under various storage
GERMINATION AND STORAGE OF SEEDS OF GREWlA OPTlVA
conditions. The equation when applied to other conditions of temperature and moisture was able to predict approximately. Ellis and Roberts (1980) emphasized that if the equation is applied, they should be used as a rough guide to the expected behaviour of high quality seed lots.
Acknowledgement The present paper forms a part of the Indian Council of Forestry Research and Education project on "Storage of Forest Tree Seeds" funded by the World Bank and the same is acknowledged.
References Ellis, RH. and Roberts, E.H. (1980). Improved equations for the prediction of seed longevity. Annals of Bot., 45, 13-30. Goldbach, H. (1979). In germination 399-402.
and storage
of Bixa orellana
seeds. Seed Science
and Technology,
7,
Harrington, J.P. (1963). Packaging seed for storage and shipment. Seed Science and Technology, 1,701-709. Harrington, J.P. (1970). Seed and pollen storage for conservation of plant gene resource. In: Genetic Resources in Plants, their exploration and conservation, Handbook. No. 11, International Biological Programme, London. Holmes, G.D. and Buszewicz, G. (1956). Longevity of acorns with several storage methods. Report. For. Res., For. Com. London 1954/55,88-94.
"'-'"
ISTA (1993). International Rules for Seed Testing. Seed Science and Technology, 21. Maithani, G.P., Bahuguna, V.K., Sood, O.P. and Rawat, M.M.S. (1987). Effect of temperature and container on Horoptera integrifolia plant seeds for minimum retention of viability and vigour. Ind. For., 113, 466-470. Nayal, 1.S., Thapliyal, RC., Rawat, M.M.S. and Phartyal, S.S. (2000). Desiccation tolerance and storage behaviour of neem (Awdirachta indica A. Juss.) seeds. Seed Science and Technology, 28, 761-767. Piotto, B. (1995). Influence of scarification and prechilling on the germination of seeds of Pistacia lentiscus. Seed Science and Technology, 23, 659--663. Roberts, E.H. (1973). Predicting the storage life of seeds. Seed Science and Technology, 1,499-514. Shrestha, RK. and Gautam. M.K. (1989). Pretreatment on Grewia optiva seeds. Technical Paper No. 2/89 Lumle Agriculture Centre. Pokhra, Nepal. Singh, C., Kumar, V., Sharma, S.K., Singh, C. and Kumar, V. (1997). Germination behaviour of Grewia optiva seeds under different presowing treatment. Van Vigyan., 35, 3-4, 132-136. Sneh Lata, Verma, K.R. and Lata, S. (1993). Presowing treatment ofbhimal (Grewia optiva Drumond) seeds. Ind. For., 119, 135-138 Willan, RL. (1985). A guide to forest seed handling with special reference to the tropics. FAO Forestry Paper 20/2. FAO, Rome.
