In early development seed moisture content is high during a period of cell division and differentiation (histo-differentiation), this is followed by reserve transfer from the parent plant when moisture content decreases slightly. Once the seed reaches mass maturity, the point at which maximum dry weight is achieved (Ellis and Pieta Filho, 1992), the abscission layer forms, the seed is no longer attached to the parent plant and moisture level decreases. The rate at which moisture decreases is dependent on properties of the seed and seed covering structures as well as on environmental conditions. Seeds are hygroscopic – once detached from the parent plant they will absorb or desorb moisture depending on the relative humidity of their environment and will continue to do so until they have reached equilibrium with environmental conditions. The moisture status of seeds within fleshy fruits will remain high, whereas for seeds in dry fruits moisture content is much more dependent on environmental conditions. For species with dry fruits moisture content is therefore an indicator of seed maturity, where seeds with a higher moisture content are likely to be less mature.

Most agricultural species are desiccation tolerant (orthodox) and have seeds that survive drying to low moisture contents. There are also seeds which show desiccation sensitive (recalcitrant) seed storage behaviour and do not tolerate drying to low moisture levels. Seeds which only tolerate partial drying are described as intermediate. Seeds were originally classified as being orthodox or recalcitrant by Roberts (1973). It is important to understand seed storage behaviour before making any decisions on how to treat a seed collection. Information on seed storage behaviour is available from several sources (Hong et al., 1998, Royal Botanic Gardens Kew, 2020).  

If seeds are being stored long term, to avoid fluctuations in moisture it is important that either the environment in which they are being stored is controlled, or that seeds are dried to a suitable moisture content and then stored within moisture-proof containers. 

Moisture content is the most common method of measuring the amount of water in seeds, but there are alternatives - relative humidity (or water activity) and water potential can also be used.  

Equilibrium relative humidity is measured with a hygrometer, and there are various types of these meters available. To test the equilibrium relative humidity (eRH) or water activity of a seed sample, seeds are enclosed within a sealed chamber with seeds filling as much of the chamber as possible. Seeds equilibrate with the air in the chamber, and then the relative humidity of that air is measured. Provided the seeds have equilibrated the air will have the same relative humidity as the eRH of the seeds. The use of eRH as a measure of moisture status in seeds is summarised well by Probert et al. (2003).  

For some seed collections eRH may be preferable to a moisture test as it is a non-destructive test, and also because different components of seeds may be at different moisture contents (dependent on properties of the different parts of the seed, seed coat, etc) whereas the eRH of all components should be the same. 

There is a reverse sigmoidal relationship between seed moisture content and eRH, described by an isotherm. The shape of the isotherm will differ for seeds of different species and is dependent on their oil content. The shape of the isotherm also depends on whether seeds are absorbing or desorbing moisture when moisture content is measured, an effect known as hysteresis (Sun, 2002). eRH cannot directly predict moisture content but can be an equally useful measure of seed moisture status.  

In orthodox seeds the two factors with the greatest influence on how well seed quality is maintained during storage are temperature and moisture content. Reductions in moisture content and temperature are known to improve seed longevity (Roberts and Ellis, 1989). A general rule of thumb (Harrington, 1972) is that seed storage life will double for every 1% decrease in seed moisture content. This rule applies down to moisture contents that are in equilibrium with approximately 15% RH. Agricultural seeds, which are stored from one year till the next, tend to be stored at moisture levels of 5-10%, but seeds stored in gene banks for long-term conservation may be stored at lower moisture levels to ensure long-term survival.  

A knowledge of seed moisture content can inform decisions on how freshly harvested seeds should be handled (Hay and Probert, 2003; Butler et al., 2009). If seeds are harvested before fully mature their moisture content will be high, and they may benefit from delayed or slow drying. If moisture content is low, then it is likely that immediate storage at a low moisture content is the best course of action. Variable maturity within seed collections is likely to be more of an issue for wild species as they have not been bred to all mature at the same time. A post-harvest treatment, such as holding seeds at an elevated RH for a period before drying can reduce variation and produce a more uniform population of seeds. 

In crop species moisture content is determined either gravimetrically using an oven or using a moisture meter. For wild species it would not be practicable to test for moisture using a moisture meter. Most moisture meters require a relatively large sample of seeds and need to be calibrated individually for each species for which they are used, even for species that are relatively like one another. Calibration of moisture meters is a lengthy process and is therefore not recommended. Determination of moisture content using an oven is the most accurate method of determining moisture content.  

There are various temperatures and durations specific to different species for moisture testing given in both The International Rules for Seed Testing (ISTA, 2020) and the Association of Official Seed Analysts Rules for Testing Seeds (AOSA, 2020) The AOSA Rules tend to be more species specific, and the ISTA Rule more general, therefore the ISTA Rules are described below, but it may be worthwhile consulting both sets of Rules and accompanying handbooks. ISTA has two oven methods for moisture content determination – the low constant temperature method and the high constant temperature method. The high temperature method (130°C for one or two hours) allows a faster determination of moisture content. The low temperature method (103°C for 17 hours) is the reference method and the method that should be used for any species not listed in the ISTA Rules.  

The ISTA Rules recommend testing two replicates of a sample size of 4-5g. For some species, or for small seed collections, this may be impracticable. Research (e.g. Hay et al. 2010) has demonstrated that it is possible to determine moisture content accurately for individual seeds weighing just one or two milligrams, so it is feasible that a much smaller sample of seeds than 4-5g could be used to give a good estimate of moisture content. If this were to be done, then a balance with an appropriate degree of accuracy should be used – it should be possible to record the weight of the sample to at least three significant figures. It would also be preferable to use smaller containers if testing small moisture samples. Ideally the weight of the container should not be more than 10 times greater than the weight of the seeds being tested. 

There are certain species listed in both the ISTA and AOSA Rules that require to be cut or ground prior to testing for moisture. These species tend to have impermeable or thick seed coats or other covering structures that impede the loss of moisture during a moisture content test. Grinding or cutting therefore permits accurate measurement of moisture content. Information on whether cutting or grinding is required may not be available for many species, but it should be kept in mind that this may be necessary. Any grinding or cutting should be done as quickly as possible to minimise exposure of the seed to ambient conditions and therefore minimise any change in moisture prior to the test being carried out.  

The requirements for all moisture tests will be applicable to wild species and should be followed if possible. These include:  

• Prior to testing any seed sample intended for moisture content testing should be kept in a moisture-proof container.
• Throughout the test seeds should be exposed to ambient conditions for as short a time as possible. 
• The sample should be mixed well before testing. This can be done by simply stirring the sample with a spoon or similar implement.
• After drying in the oven seeds should be cooled in containers with the lids on in a desiccator to avoid any re-uptake of moisture.

Further detailed information on moisture testing can be found in both the ISTA Handbook on Moisture Determination and the AOSA Seed Moisture Testing Handbook.  

Association of Official Seed Analyst (AOSA). 2020 AOSA Rules for Testing Seeds, Volume 1. Published by the Association of Official Seed Analysts. 

Butler, L. H., Hay. F. R., Ellis, R. H. and Smith, R. D. 2009. Post-abscission, pre-dispersal seeds of Digitalis purpurea remain in a developmental state that is not terminated by desiccation ex planta. Annals of Botany, 103, 785-794.  

Ellis, R. H. and Pieta Filho, C. 1992. The development of seed quality in spring and winter cultivars of barley and wheat. Seed Science Research, 2, 9-15. 

Harrington, J. F. 1972. Seed storage and longevity. p. 145-250. In T.I. Kozlowski (ed.) Seed biology. Ill. Insects and seed collection, storage and seed testing. Academic Press, New York.  

Hay, F. R., Smith, R. D., Ellis, R. H. and Butler, L. H. 2010. Developmental changes in the germinability, desiccation tolerance, longevity and hard seededness of individual seeds of Trifolium ambiguum. Annals of Botany, 105, 1035-1052. 

Hay, F. R. and Smith, R. D. 2003. Seed Maturity: when to collect seeds from wild plants, pp. 97-133. In: Seed Conservation turning science into practice (eds R. D. Smith, J. B. Dickie, S. H. Linington, H. W. Pritchard and R. J. Probert. Royal Botanic Gardens, Kew, UK.  

Hong, T. D., Linington, S. H. and Ellis, R. H. 1998 Compendium of information on seed storage behaviour. Royal Botanic Gardens, Kew, UK.  

International Seed Testing Association (ISTA). 2020. International Rules for Seed Testing. The International Seed Testing Association, Bassersdorf, Switzerland. 

Probert, R. J., Manger, K. R. and Adams, J. 2003. Non-destructive measurement of seed moisture, pp. 369-387. In: Seed Conservation turning science into practice (eds R. D. Smith, J. B. Dickie, S. H. Linington, H. W. Pritchard and R. J. Probert. Royal Botanic Gardens, Kew, UK. 

Roberts, E. H. 1973. Predicting the storage life of seeds. Seed Science and Technology, 1, 499-514. 

Roberts, E. H. and Ellis, R. H. 1989. Water and seed survival. Annals of Botany, 63, 39-52. 

Royal Botanic Gardens Kew. 2020. Seed Information Database (SID). Version 7.1. Available from: http://data.kew.org/sid/ (April 2020) 

Sun, W. Q. 2002. Methods for the study of water relations under desiccation stress, pp. 47-92. In: Desiccation and Survival in Plants: Drying without Dying (eds M. Black and H. W. Pritchard). CABI Publishing, UK.