Effect of treatments on seed dormancy breaking, seedling growth, and seedling antioxidant potential of Agrimonia eupatoria L.

Highlights

• The seed coat removal in Agrimonia eupatoria L. reduced dormancy and improved the germination to 50 %.

• The combination of seed coat removal and application of 0.2 % KNO₃, increased the germination to 93 %.

• The combination of seed coat removal and application of 100 mg L^-1 GA₃, increased the germination to 73 %.

• The combination of seed coat removal and application of 1% KNO₃, increased the seedling length.

• The KNO₃ and GA₃, at low concentration increased the enzyme activity, and antioxidant content in uncoated seeds.

Abstract

Agrimonia eupatoria L. is an important medicinal plant of the Rosaceae family. Antioxidant and volatile components have been identified in this herb. The seed dormancy of A. eupatoria causes time-consuming germination with a low percentage. In this study, the effects of treatments including 4 °C wet cold (24 h and 48 h), seed coat removal, potassium nitrate (0.2 %, 1%, and 3%), and gibberellic acid (100, 200, and 300 mg L⁻¹) were studied on seed dormancy breaking, seed germination indices, seedling growth parameters, the activity of α- amylase, antioxidant potential, as well as catalase and peroxidase activity in the in vitro conditions. The results showed that in coated seeds, seed coat removal was effective treatment on seed dormancy breaking, and increased the germination. The effect of 0.2 % potassium nitrate and 100 mg L⁻¹ gibberellic acid was very low, and 4 °C wet cold (24 h and 48 h) had no effect. The effect of treatments was more significant in uncoated seeds. The most effective treatments in uncoated seed were 0.2 % potassium nitrate and 100 mg L⁻¹ concentration of gibberellic acid, respectively. These treatments had a significant effect on germination indices, growth parameters, antioxidant potential, and α- amylase activity. Further, 1% potassium nitrate was the most suitable treatment for increasing the hypocotyl axis and seedling weight in uncoated seeds. In addition, 4 °C cold (24 h and 48 h) had a very little positive effect on all the mentioned indices. In all measurements, 3% potassium nitrate showed the weakest results.

Introduction

Rosaceae has more than 300 species of trees, shrubs, and herb worldwide, and almost all of the plants in this family are economically and pharmacologically valuable. The genus Agrimonia is native to Europe and Asia and has 15 species (Tsirigotis-Maniecka et al., 2019). Agrimonia eupatoria L. commonly known as agrimony is a valuable medicinal plant of this genus. It is an herbaceous plant with a height of about 30−60 cm and grown in areas with moderate light and moisture on the margins of forests and grasslands (Najafpour and Mirza, 2009). The yellow flowers of this plant have 5 petals, 5 alternate sepals, 15 anthers, and elongated spike inflorescence. The fruit is cone-shaped and has hard hook-like branches (Santos et al., 2017). The seed shape is spherical-angular with a flat basal part, seed color is light brown or dark brown, and its diameter is 7–9 mm. The hard seed coat has hair and hooks on the surface. Agrimony seeds ripen in September (Faghir et al., 2020).

Numerous studies have been conducted on the medicinal properties of A. eupatoria. This plant is a major source of polysaccharides, tannins, flavonoids, catechins, procyanidins, agrimofol, coumarin, silica, malic acid, phytosterols, iron, as well as vitamins B and K (Garcia-Oliveira et al., 2020). In traditional medicine, leaves, flowering branches, and plant seeds have been used to treat diseases such as bladder swelling, intestinal edema, wounds and skin injuries, bleeding gums, eye infections. The anti-inflammatory, antiviral, antibacterial, anti-diabetic, and anti-cancer effects of this plant are associated with its antioxidant compounds (Muruzovic et al., 2016; Tsirigotis-Maniecka et al., 2019). It is also reported to be effective in treating liver disease, kidney disease, gall bladder, rheumatism, laryngitis, and lung disease (Feng et al., 2013). According to Akbar report, fifty-two volatile components have been identified in the leaves and roots of the A. eupatoria (Akbar, 2020).

Seed dormancy in natural conditions causes inactivation and inability of seed germination on the maternal basis, and thus allows for greater dispersal and passage of inappropriate environmental conditions (Sohindji et al., 2020). This can be a factor in the survival of the offspring but when the goal is mass production of economic and medicinal plants, it is usually undesirable. Seed dormancy includes physiological, physical, morphological, morpho-physiological types (Tang et al., 2019). The factors that cause dormancy in the seed include unsuitable environmental conditions, existence of hard seed coat, immature embryos, inactivation of enzymes such as α-amylase (Chahtane et al., 2017).

Seed dormancy breaking plays a key role in beginning of germination and plant growth in the propagation of herbaceous medicinal plants. According to the International Seed Testing Association (ISTA), various methods have been suggested to stimulate seed dormancy breaking, and seed germination such as seed scarification, seed stratification, salinity, chemical solutions and regulatory hormones (ISTA, 2013; Sharififar et al., 2015). Seeds of some plants have double dormancy, so physical and physiological dormancy of the seed should be eliminated using physical treatments such as cold and seed coat removal along with chemical treatments such as potassium nitrate (KNO₃) and plant hormones. KNO₃ and gibberellic acid (GA₃) have been reported as functional chemical compounds in improving seed germination and increasing enzyme activity in many plant species (Mousavi et al., 2019; Sohindji et al., 2020). These two compounds are non-hazardous natural stimulants that enhance the antioxidant potential and production of secondary metabolites by inducing pseudo stress (Caverzan et al., 2016).

Agrimony seeds show dormancy at maturity (ISTA, 2013). Kline and Sørensen (2008) studied the several species of Agrimonia. According to their reports, to overcome of seed dormancy, the seeds must be kept in a humid environment for two months, and then seed germination usually begins 3–5 weeks after soil cultivation. Catana et al. (2020) compared the germination of several varieties of medicinal plants under in vivo and in vitro conditions. According to their reports, the use of mercury chloride to sterilize the agrimony seed made the seed coat more permeable and the germination rate increased up to 50 %. In some species of Rosaceae family such as Sorbus pohuashanensis, Prunus mahaleb, Rubus takesimensis, Rosa multibracteata, seed dormancy has been reported to be of physical and physiological types (Bian et al., 2013; Choi et al., 2016; Zhou et al., 2009). In Prunus dulcis, Prunus persica, and Prunus armeniaca, the removal of seed coat increased the germination percentage (Kanjana et al., 2016; Szymajda et al., 2019).

As A. eupatoria is in the Red list of threatened medicinal plants (Allen et al., 2014), it is necessary to propagate it under in-vitro conditions. Therefore, the present study is conducted to investigate the effect of treatments including 4 °C wet cold (24 and 48 h), seed coat removal, as well as KNO₃ and GA₃ at different concentrations, on 1) reduce seed dormancy and improve germination indices, 2) increase growth parameters such as seedling length, seedling weight, and seedling vigor 3) enhance the antioxidant content 4) increase enzyme activity such as catalase, peroxidase, as well as α-amylase of A. eupatoria.