Seeds of 5 cultivars of lettuce (Lactuca sativa L.) were adjusted to moisture contents between 5 and 22% and subjected to temperatures of 5, -18, -70, and -196°C (liquid nitrogen) for 7 days. Seeds with moisture contents up to 18% were not damaged by either -18 or -196°C; however, the -70°C treatment resulted in loss of germination even with seeds containing as little as 16% moisture. Seeds held at 5°C showed no loss in viability. The loss in germination at -70°C appeared to be related to cooling rate. High moisture (18 to 20%) seeds were cooled to -196°C at different rates from 1 to 200°C/min. Germination was reduced in seeds cooled at rates slower than 25°C/min. At 200°C/min cooling rate and 18 to 20% seed moisture both germination and root growth occurred although at reduced levels compared with control seeds held at 5°C. Prolonged storage (33 days) in liquid nitrogen of rapidly frozen high moisture lettuce seeds did not result in further loss of germination after the initial freezing to -196°C.
Dehydration effects on freezing characteristics and survival in liquid nitrogen were studied in 11 species of tropical seeds and in silver maple (Acer saccharinum L.) seeds. Differential thermal analysis was used to determine the threshold moisture level below which seed tissue water was in an unfreezable state. Desiccation-sensitive seeds, areca palm [Chrysalidocarpus lutescens (Bory) Wendl.] and silver maple, did not survive dehydration below the threshold moisture level and did not survive exposure to liquid nitrogen. Nine of 10 desiccation-tolerant seeds [strawberry guava, Psidium cattleianum Sabine; passion fruit, Passiflora edulis f. flavicarpa Deg.; Ceara rubber, Manihot glaziovii Mull. Arg.; dwarf schefflera, Schefflera arboricola (Hayata) Merrill; common guava, Psidium guajava L.; papaya, Carica papaya L., apple of sodom, Solarium sodomeum L.; prickly poppy, Argemone glauca Pope; and seamberry, Sabalparviflora Becc.] survived dehydration to as low as 2% to 12% moisture content (below the threshold moisture levels determined) and in the dehydrated state survived exposure to liquid nitrogen. Coffee (Coffea arabica L. var. Bourbon) seeds tolerated dehydration to as low as 8% moisture content but did not survive exposure to liquid nitrogen. These results demonstrate the feasibility of cryopreserving seed germplasm of several tropical species.
Previous research showed a relationship between viability and electrolyte leakage; here, we develop an index of viability based on electrolyte leakage from sample populations of seeds soaked in deionized distilled water. Conductivity of leachates from individual seeds was determined for 10 lots of lettuce (Lactuca sativa L.), each germinating at 99%. Conductivity data for two lots of soybean (Glycine max L.) seeds germinating at 100% and 74%, respectively, were obtained from literature. Cumulative frequency distributions (CFD) with a class interval of one µA, were fitted with a natural logarithmic form of the Richards function, which requires no arbitrary starting values. The procedure provided an effective estimation of slopes [(dCF/dµA)MAX] of hypothetical lines tangent to inflection points of the respective sigmoidal CFD curves. We suggest that this maximum slope, or internal slope can be used as a seed viability index. The index is unaffected by outlier µA readings and reflects the shape of the CFD. It is also a measure of seed-to-seed variability in leachate conductivity. The se of the 10 internal slopes derived from the 10 lettuce seed lots was 3.8. The viability index is sensitive, since nearly a four-fold difference in internal slope was found for the two soybean seed lots. The greater the internal slope, the less the variation among individual seed conductivities and the higher the seed quality.