JIA-2019-11

2436 Chalisa Chaengsakul et al. Journal of Integrative Agriculture 2019, 18(11): 2435–2445 a complex biological process that is influenced by many factors (Li et al . 2017). The most important factors that accelerate seed deterioration are relative humidity (RH), temperature and oxygen (Groot et al . 2015). Evaluation of seed deterioration in a warehouse is very important in the management of seed inventory. Seed scientists don’t have any method for evaluating seed deterioration before seed quality declines below regulatory requirements. A standard germination test is one of the truly reliable methods, but it is destructive, time-consuming, and highly subjective (Bicanic et al . 2003). Many previous works have emphasized that mitochondrial degradation is the main cause of seed deterioration and leads to the loss of seed vigour (Smagula and Bramlage 1977; Ferguson et al . 1988; Shaban 2013). Reactive oxygen species (ROS) are related to mitochondrial dysfunction and seed deterioration after the start of imbibition. In many cases, aerobic respiration is transformed into anaerobic respiration by ROS (Vartapetian et al . 2003; Li et al . 2017). The major product of anaerobic respiration in the seed is ethanol. Thus, ethanol production during imbibition is considered as a good indicator of seed deterioration (Woodstock and Taylorson 1981; Bicanic et al . 2003; Buckley and Huang 2011; Kodde et al . 2011). Kodde et al . (2011) reported a development of method for evaluating seed deterioration using a modified breath analyser that quantified the headspace ethanol from deteriorated seed, the so-called fast ethanol assay. Although many studies have investigated the ethanol production of stored seed, there has been little work using large seed such as maize (Buckley and Huang 2011; Kodde et al . 2011). Protocol optimization of the fast ethanol assay in maize seed is difficult because maize seed constitutes a low amount of the living embryo per gas chromatography (GC) vial container volume compared to other successfully analysed species, such as cabbage ( Brassica oleracea ), cotton ( Gossypium sp.), and lettuce ( Lactuca sativa ). Storage at an elevated temperature and RH of the atmosphere surrounding the seeds is generally called accelerated ageing (AA). The AA test is a recognised seed vigour test (ISTA2015). Frequently, it is used as an indicator of seed longevity under conventional storage conditions (Baalbaki et al . 2009). Unfortunately, the physiological process of seed deterioration between dried and wet seed is different (Kibinza et al . 2006; Groot et al . 2012). The defence mechanisms, preventing seed from the stress and the activity of enzymes involved in repair, might vary with differences in either temperature or water activity around the seed (Kibinza et al . 2006; Groot et al . 2012). The tests following hot-humid procedures could have limitations in predicting seed shelf-life under rather dry conditions (Groot et al . 2012). Interestingly, the degradation mechanisms of mitochondria in maize seed during this process have never been explained through ethanol assay and mitochondrial- related gene expression, although they may be behind the phenomenon. Currently, model organism databases provide in-depth biological data for the intensively studied deterioration process in seed. The key enzyme producing ethanol is alcohol dehydrogenase that is translated from the ADH gene family (Woodstock and Taylorson 1981; Strommer 2011). In seed, alcohol dehydrogenase plays an important role in ethanol production through the anaerobic respiration pathway (Okimoto et al . 1980; Strommer 2011; Takahashi et al . 2014). Pyruvate decarboxylase is also responsible for the anaerobic production of acetaldehyde and ethanol in higher plants (Kimmerer 1987). Mitochondria generate adenosine triphosphate (ATP) through the mitochondrial respiratory chain complex which consists of four multi- subunit respiration complexes and two mobile electron carriers (cytochrome c and ubiquinone) (Fontanesi et al . 2008). Cytochrome oxidase ( COX ) plays the central role of aerobic metabolism by its participation in respiratory control (Fontanesi et al . 2008). Yin et al . (2016) reported that rice seeds were down-regulated in COX during the transformation of seed viability during ageing from plateau phase to the rapid decreasing phase, the critical node of seed ageing, and then activity of cytochrome oxidase decreased. Before plant cell death, ATP synthase, especially in β -subunits, is the key negative regulator of the process (Chivasa et al . 2011). In Arabidopsis , AOX1a and AOX1d are the significant stress responsive genes amongst hundreds of known genes encoding mitochondrial proteins (Clifton et al . 2006). The AOX gene expression is influenced by stress stimuli, such as cold, oxidative stress, and pathogen attack. This is an important factor constricting oxidative phosphorylation through electron transportation and causes dysfunction in the respiratory metabolism (Vanlerberghe and McIntosh 1997; Clifton et al . 2006). The present paper reports the investigation of commercial maize seed deterioration after natural and artificial ageing in both conventional seed quality and terms of gene expression levels as well as ethanol production using fast ethanol assay to search for an understanding of deterioration and evaluates novel possibilities in detection. 2. Materials and methods 2.1. Seed sources F 1 hybrid maize seeds obtained from Pacific Seeds (Thailand) Co., Ltd., were used in this experiment. Seed samples were produced in 2016 in a commercial seed production field. Afterwards the seeds were stored at