Effects of pulsed electric field and metal nanoparticle soaking on germination, growth, and antioxidant properties of rapeseed sprouts

Karolina Sokal; Magdalena Kachel-Górecka; Małgorzata Stryjecka; Marta Krajewska

doi:10.1038/s41598-026-52755-6

. 2026 May 25;16:23907. doi: 10.1038/s41598-026-52755-6

Effects of pulsed electric field and metal nanoparticle soaking on germination, growth, and antioxidant properties of rapeseed sprouts

Karolina Sokal ¹, Magdalena Kachel-Górecka ^1,^✉, Małgorzata Stryjecka ², Marta Krajewska ^3,^✉

PMCID: PMC13434722 PMID: 42185472

Abstract

The aim of this study was to determine the effect of pulsed electric field (PEF) application and soaking (coating) of rapeseeds in metal nanoparticles (AgNP or CuNP) on germination capacity, the stem and root length, the contents of chlorophyll a, chlorophyll b and carotenoids, colour profile and antioxidant properties: antioxidant activity (FRAP), polyphenols, total phenolic compounds (TPC), antioxidant capacity (ABTS) and total antioxidative activity (DPPH) of rapeseed sprouts. It was found that higher PEF energy (5.5 kJ/kg) and soaking seeds in nanoparticle solutions (25 ml/l and 50 ml/l) do not affect seed germination capacity. However, a lower PEF energy (1.0 kJ/kg) and treatment of seeds with the experimental nanoparticle solutions, in most cases, reduced rapeseed germination capacity compared with the control samples. Following the application of a 1.0 kJ/kg PEF, stem lengths were 18.9%-48.4% higher than those of the corresponding control samples. Increased sprout root length was observed for treatments with a PEF level of 5.5 kJ/kg, i.e., the observed values were 3.0%-103.0% higher than those for the corresponding control samples. Copper nanoparticles applied at two experimental concentrations at an energy of 1.0 kJ/kg adversely affected root elongation; the resulting root lengths were 4.3% and 41.0% shorter than those of the respective control samples. The contents of chlorophyll a (chl a), chlorophyll b (chl b) and total carotenoids decreased in sprouts in all experimental variants compared with the control. The chlorophyll a content decreased by 1.89 to 11.11%, while chlorophyll b content decreased by 2.27 to 13.07% compared with the control. The study confirmed an increase in the antioxidant properties of rapeseed sprouts after application of silver or copper nanoparticles with a pulsed electric field. The Antioxidant activity capacity (measured by ferric reducing antioxidant power assay - FRAP), the contents of polyphenols and total phenolic compounds (TPC) ranged from 81.89 to 198.52 mg TE/g DM, 14.23 to 15.97 mg GAE/g DM, and 22.50 to 27.00 mg GAE/g DM, respectively, and showed an upward trend compared with the control samples. The results of the present study suggest that further analysis is required to understand how the methods studied affect sprout development and the increased antioxidant activity.

Supplementary Information

The online version contains supplementary material available at 10.1038/s41598-026-52755-6.

Keywords: Polyphenol, Chlorophylls, Antioxidants, Metal nanoparticles, Rapeseed, PEF, Germination

Subject terms: Biochemistry, Biotechnology, Nanoscience and technology, Plant sciences

Introduction

Rapeseed (Brassica napus L.) is one of the main oil crops grown both in Poland and in Europe, next to sunflower. The plant contains high levels of vitamin E, phytosterols, polyphenols, flavonoids, polypeptides, polysaccharides, and other bioactive compounds¹. The main components of rapeseed are fatty acids, proteins, phenols, and glucosinolates (GLS), which may have health-promoting effects, such as antioxidant and anticancer properties^2,3.

Growing consumer awareness is driving the search for new and richer food products, whether fresh or preserved. Preservation through drying aims to provide humans with the optimal chemical composition of these products. Oilseed sprouts can become one of these products.

Sprouts are grown directly from plant seeds that store nutrients, in either dark or light conditions, and are also referred to as “living vegetables”⁴. Sprouts are plants that can be produced quickly, easily, and economically due to simple equipment and material requirements, as well as a rapid growth process that lasts for several days⁵. Seed germination is a process that begins when a dry and inactive seed absorbs water and ends when the embryonic axis begins to elongate. This process can occur within 24–36 h, depending on the temperature, and is followed by the elongation of the embryonic root and the development of a new plant⁶. Sprouted plant seeds contain high levels of vitamins, minerals, and bioactive substances, making them an extremely valuable dietary supplement. Oilseed sprouts are a source of vitamins (group B, and vitamins E, A and C), minerals (calcium, iron, magnesium, potassium), dietary fibre and polyphenols, as well as bioactive compounds, namely a group of secondary metabolites, also known as phytochemicals⁷. In addition, they have a unique taste and many health benefits, which is why they are preferred by consumers^8,9. As a result of complex biochemical reactions, new compounds, such as vitamins and antioxidants, are formed. Sprouts contain more soluble sugar, chlorophyll and carotenoids than seeds¹⁰.

The sprouting process can support the accumulation of active compounds, including increased levels of amino acids, simple sugars, and other nutrients. Moreover, the sprouting process reduces levels of anti-nutrients and improves the product’s digestibility and sensory characteristics¹. Multiple studies have demonstrated that consuming sprouts can effectively reduce the risk of various diseases, such as intestinal and heart diseases, diabetes, and neoplasms^11,12.

To accelerate germination and improve sprout quality, appropriate support processes can be implemented. These include PEF (pulsed electric fields), nanotechnology and drying. PEF can affect seed germination and, at the same time, serve an important role in the food industry by supporting processes aimed at improving oil and juice extraction efficiency. In addition, it is used to inactivate microorganisms and produce nutrient solutions for hydroponic systems. The action of PEF is based on the phenomena of electrical breakdown and electroporation, which involve the creation of microscopic holes in cell membranes and phospholipid bilayers¹³. This mechanism increases the permeability of plant cells and facilitates and accelerates the release and diffusion through their membranes. The positive effect of PEF is largely dependent on the type of seed coat and the plant species¹⁴.

However, modern agricultural technologies are not limited to methods that improve extraction or germination efficiency. For environmental protection, increasing attention is being paid to nanoparticles as an alternative to traditional pesticides. Their application may prove an effective solution for controlling harmful organisms, i.e., pathogens and pests that threaten plants. Research results show that approximately 90% of traditional plant protection products used in agriculture, depending on the climate and application method, fail to bring the desired results. Consequently, the use of such products often requires repeated applications, causing additional environmental damage¹⁵.

Nanotechnology provides new opportunities for agriculture by offering environmentally friendly solutions for plant protection. Formulations containing nanoparticles, such as silver (AgNPs) or copper (CuNPs), exhibit strong antimicrobial and biocidal properties, making them an effective alternative to traditional plant protection products. Silver nanoparticles, due to their nanometric size, effectively eliminate bacteria and fungi, while copper nanoparticles block the enzymes required for pathogen function, reducing the incidence of plant diseases by approximately 31%¹⁶. Research has shown that CuNPs at 100 µg/ml effectively inhibit the growth of Botrytis cinerea and Sclerotinia sclerotiorum in cucumber cultivation, limiting pathogen development by 94.12% and 92.48%, respectively¹⁷. Not only can the coating of rapeseed with AgNPs or CuNPs accelerate germination, but also stimulate the growth of sprouts by elongating the stem and the root¹⁸. In the context of sustainable agriculture, it is crucial to reduce the use of chemical fertilisers, thereby reducing greenhouse gas emissions and water pollution^19,20. As urban agglomerations develop rapidly, it is becoming necessary to implement modern technologies that support plant growth. According to Arnott et al.²¹, the use of metal nanoparticles supports plant germination and improves the vigour of young sprouts. In addition, numerous scientific sources report comparable or higher germination rates and increased vigour in seeds subjected to nanoconditioning. In contrast, there are physiological indicators of nanoparticle toxicity, including germination energy, root elongation, and leaf biomass. Numerous studies emphasise that some nanoparticles can cause significant adverse effects, such as reduced seed germination, inhibited stem growth, or shortened roots^22,23. According to reports by Mushinskiy and Aminova²⁴, treating potato Solánum tuberósum L. tubers with iron nanoparticles stimulated the growth of sprouts (55.1% and 21.4%) and the roots (34.4% and 12.5%), and increased the chlorophyll a content (57–98%), whereas the application of copper and molybdenum nanoparticles had no significant effect on plant development. To measure the properties of plant sprouts, it is necessary to begin with the drying process. Sublimation drying is a process comprising several stages. It involves freezing the raw material, sublimation of frozen ice, and drying it under vacuum, which allows the unfrozen water contained in the material to be removed. For the sublimation process to be effective, the pressure in the drying chamber and the temperature of the raw material must be below the triple point of water^25,26.

In this study, we decided to focus on rapeseed sprouts primarily due to the significant lack of literature data on their bioactive composition and antioxidant properties compared to other oilseed sprouts. Therefore, we believed that their analysis could significantly contribute to the current state of knowledge and fill an existing research gap.

The lack of literature reports on the effect of the application of the PEF technology and nanoparticles on seeds prompted the authors to conduct a study to determine the effect of the application of metal nanoparticles (AgNPs or CuNPs) in two solutions (25 ml/l and 50 ml/l), and PEF treatment at 1.0 and 5.5 kJ/kg, on rapeseed germination capacity and the antioxidant properties of the sprouts. The obtained rapeseed sprouts were subjected to drying, and the resulting material was tested for antioxidant properties, including the antioxidant activity (FRAP), the total phenolic compounds (TPC), antioxidant capacity (ABTS), total antioxidant activity (DPPH), and the contents of carotenoids as well as chlorophyll a and chlorophyll b. A research hypothesis was also formulated, positing that the application of PEF technology and silver or copper nanoparticles would enhance rapeseed germination and the antioxidant properties of their sprouts.

Materials and methods

Study material

Seeds of winter rapeseed (Brassica napus L.) of the “Ricky” variety (uncoated) were sourced from the company Hodowla Roślin Strzelce Sp. z o.o., IHAR Group.

Silver and copper nanoparticles

The study material consisted of two samples of nanoparticles in colloidal form, a type of suspension in which particles suspended in the solvent are less than 100 nm in size. The selected silver and copper nanoparticles, purchased from ITP-system Sp. z o.o. with a concentration of 4000 ppm, are commonly used additives in polar solvent-based products and are adapted for use as plant protection products. To verify the quality of the purchased nanocolloid, nanoparticle samples were analysed using a transmission electron microscope (TEM).

Sample imaging using transmission electron microscopy

Silver and copper samples were ground into a fine powder using an agate mortar. The resulting powder was poured over with 99.8% ethanol (POCH) to form a suspension, and placed in an ultrasonic homogeniser for 10 s. The suspension containing the sample was then collected with a pipette, applied onto copper grids (200 mesh) coated with lacey formvar stabilised with carbon (Ted Pella), and left on blotting paper for the ethanol to evaporate. Subsequently, the samples were placed in a special holder and transferred to an electron microscope. Imaging of the samples was performed using a Titan G2 60–300 kV transmission electron microscope (FEI) equipped with a field emission gun (FEG). Microscopic examination of the samples was performed at an electron beam accelerating voltage of 300 kV. TEM imaging of the samples’ microstructure was performed in bright-field mode, using a CCD camera as the detector. In this mode, imaging is performed using electrons from the zero beam that pass through the sample without being scattered.

Seed preparation for testing

Rapeseeds were placed in a 70% ethanol solution for 15 s to sterilise their surface. They were then transferred to a 1.5% sodium hypochlorite (NaClO) solution for 10 min, and rinsed five times with distilled water. The rinsed rapeseeds were then soaked in tap water (control sample) and in tap water with silver and copper nanoparticles added for 15 min to achieve optimal conductivity. The seeds were weighed on an AS 310.X2 analytical balance (Radwag-Radom, Poland). The weighed material (200 g of seeds) was transferred to a glass vessel filled with 800 ml of water. Two solutions containing 25 ml and 50 ml of nanoparticles in 1 L of tap water were used. The samples prepared in this way were poured into a measuring cuvette and subjected to electric current under the following conditions: voltage of 24 kV, current of 1 kV/cm, distance between electrodes of 24 cm, frequency of 20 Hz and pulse width of 7 µs, in an ELEA Pulsed Electric Field system. The values of the above parameters were identical across all experimental variants. The only variable was the applied energy, i.e. 1.0 and 5.5 kJ/kg. There were three control samples: I – a control sample that contained seeds soaked only in water, II – a control sample that contained seeds soaked in water and seeds subjected to PEF energy of 1.0 kJ/kg, III – a control sample that contained seeds soaked in water and seeds subjected to PEF energy of 5.5 kJ/kg.

The seeds were germinated under laboratory conditions at room temperature (20 ± 2 °C) under natural daylight. The seeds were sown in three replications on Petri dishes with a diameter of 90 cm, lined with three layers of filter paper, 100 seeds per dish, and watered with 1.5 ml of distilled water per dish. Seven days after sowing the seeds, properly developed sprouts were counted to determine the germination capacity and the stem and root lengths were then measured. The calculations were performed according to the formula for calculating germination capacity:

Drying method

The sublimation and vacuum drying process was carried out using an ALPHA 1–4 sublimation dryer from Martin Christ, Osterode am Harz, Germany, which uses a contact heat-transfer method. This instrument consists of a drying chamber, a heating plate power supply system, a water vapour freezing system, and a control and measurement system with an interface. The dryer was equipped with a WPT 5 balance integrated with a computer, enabling continuous monitoring of the dried material’s weight. The sublimation drying process occurred at 40 °C and 52 Pa in the chamber.

Colour measurement

Colour measurement was performed using the reflectance method with an X-Rite 8200 spherical spectrophotometer, with a 12.7 mm measurement aperture. A D₆₅ light source and a standard colorimetric observer with a 10° field of view were used. Before each measurement, the instrument was calibrated using a white standard. Colour measurements were performed in five replications for dried material crushed to a dimensional class of < 100 μm.

The colour coordinates were determined in the CIEL*a*b* system. Colour measurement in this system involves the numerical determination of three coordinates L*, a* and b*, where L* denotes colour lightness and ranges from 0 for a perfectly black body to 100 for a perfectly white body. The a* coordinate determines the colour change from green (− a*) to red (+ a*), while b* determines the colour change from blue (− b*) to yellow (+ b*).

Based on the determined colour coordinates, the colour saturation (c) and colour hue (h) values for the dried material were determined, expressed in cylindrical coordinates²⁷.

Antiradical activity

The ability to neutralise free radicals against ABTS (2,2’-azynobis (3-ethylbenzothiazoline-6-sulphonate) was determined using a method developed by Re et al.²⁸. The ability to neutralise free radicals against DPPH (2,2-diphenyl-1-picrylhydrazyl) was determined according to Brand-Williams et al.²⁹. The decrease in absorbance was measured quantitatively on a spectrophotometer at wavelengths of 734 nm for ABTS, and 517 nm for DPPH. The ability to neutralise ABTS and DPPH free radicals was expressed as the EC₅₀ index, the dry mass concentration (mg/ml) required to reduce the initial concentration of either free radical by 50%.

Total carotenoid and chlorophyll contents

The chlorophyll a and chlorophyll b contents, as well as the total carotenoid content, were determined by a spectrophotometric method using a Hewlett-Packard 8453 Diode Array single-beam absorption spectrophotometer operating in the range of 190–1100 nm. The determination method was based on a method according to Lichtenthaler³⁰. The method involves extracting pigments with an 80% acetone solution, followed by measuring absorbance at wavelengths characteristic of chlorophyll a, chlorophyll b, and carotenoids.

An appropriate portion of the material was weighed, then transferred to a porcelain mortar and ground for two minutes after adding 3 ml of 80% hydrated acetone. The resulting suspension was centrifuged for three minutes at 15,000 x g (g = 9.81 m/s²). The resulting supernatant was collected quantitatively, and its volume was determined. Then, 100 µl of the supernatant was measured with a pipette, and after adding 2 ml of 80% hydrated acetone, the absorption was recorded at wavelengths characteristic of chlorophyll a, chlorophyll b, and carotenoids: 470, 646, 0.8, and 663.2 nm, respectively. The absorption spectra were measured using a quartz cuvette from Sigma-Bandai, Japan.

The contents of chlorophyll a, chlorophyll b, and total carotenoids were calculated using the following formulas:

FRAP analysis

The ferric reducing antioxidant power assay followed a procedure developed by Benzie and Strain³¹ and modified by Thaipong et al.³². For this, 50 µl of the methanolic extract of the analysed sample was added to 1450 µl of a solution prepared from acetate buffer (pH = 3.6), 10 mmol/l of a TPTZ solution, and 20 mmol/l of an FeCl₃⋅6H₂O solution (at a 10:1:1 ratio, v/v/v), and mixed thoroughly. After 8 min, absorbance was measured at λ = 593 nm using a UV 2600i Plus spectrophotometer (Shimadzu, Kyoto, Japan). A blank sample contained 50 µl of redistilled water instead of the extracts. The results are expressed as Trolox equivalents (mg TE g/ DM). The plant material was not subjected to a dedicated nanoparticle removal step prior to extraction. However, methanolic extraction primarily targets soluble antioxidant compounds, and sample preparation steps reduce the likelihood of nanoparticle presence in the analysed extracts. Nevertheless, potential interference from residual nanoparticles cannot be completely excluded.

Polyphenols

To prepare extracts, 10 g portions of the material were weighed and soaked in 100 ml of 99.8% methanol. Extraction was performed in a WL-1 shaker (Biosan, Riga, Latvia), at room temperature, for 60 min. The methanolic extracts obtained were stored in a fridge at approximately 4 °C. The total polyphenol content was determined using the method of Singleton and Rossi³³, with minor modifications. In brief, 0.05 ml of the extract was collected into a 25 ml measuring flask and mixed with subsequently added 2 ml of methanol (p.a., 99.8%), 10 ml of distilled water, and 2 ml of the Folin–Ciocalteu reagent (at a 1:5 ratio). The mixture was left to stand for 3 min. Then, 1 ml of a 10% sodium carbonate (Na₂CO₃) solution was added to the mixture, which was mixed again and left for 30 min. Afterwards, the flasks were filled up with distilled water. Absorbance was measured at a wavelength of 750 nm against the blank sample. The total polyphenol concentration is expressed in gallic acid (GA) equivalents (mg GAE g/ DM).

Statistical analysis

The study results were analysed using Statistica 13 software from StatSoft. Shapiro–Wilk (normality) and Levene (homogeneity of variance) tests were performed, in accordance with the requirements for one-way analysis of variance. One-way analysis of variance (ANOVA) was performed to determine the significance of the impact of the respective factors on the analysed values. The significance of differences between the mean values was assessed using the Tukey test. The adopted significance level was α = 0.05. All the tests and analyses were conducted in three replications.

Results and discussion

Transmission electron microscopy (TEM) tests

Transmission electron microscopy (TEM) tests (Fig. 1) showed the occurrence of fine spherical Ag nanoparticles with a crystalline structure (AgNPs) in the sample under analysis. Based on the analyses, it was found that the AgNPs have various sizes ranging from a few to several nanometres. In some places, crystals surrounded by an amorphous (stabilising) substance were noted.

Silver crystallites are mainly found individually, less frequently in the form of agglomerates (Fig. 2). The identification of phases in the “Ag” sample, performed on the basis of HRTEM and FFT (fast Fourier transform) imaging, confirmed the presence of Ag crystallites in metallic form, based on characteristic interplanar distances of 2.05Å and 2.36Å, corresponding to the (200) and (111) lattice planes.

Transmission electron microscopy (TEM) analysis (Fig. 3) revealed the presence of small, mainly spherical Cu nanoparticles (CuNPs) with a crystalline structure in the sample under analysis. The CuNP nanoparticles have various sizes ranging from a few to several nanometres. In some places, crystals surrounded by an amorphous (stabilising) substance were noted. Cu crystallites are mainly found individually, and also in the form of agglomerates (Fig. 3).

Assessment of rapeseed germination capacity

The most commonly analysed indicators used to assess the impact of different factors on plant growth include: seed germination rate, the root and stem length, and total and dry root biomass³⁴. Publications describe PEF treatment under various electrical parameters, including electric field, pulse frequency, pulse width, treatment time, and sample conductivity. Researchers have noted that applying a weak, low-frequency electric field can induce changes in biological processes, such as cell metabolism, thereby stimulating their activity³⁵. The biological effects induced by electric fields depend on both the electrical parameters and the physiological state of the treated cells. Dymek et al.⁶ conducted a study in which malting barley seeds were soaked in aerated water for 24 h, and exposed to PEF and variable voltage (0, 110, 160, 240, 320, 400 and 480 V). The seeds were then left to germinate in saturated air. It was demonstrated that, at a pulse width of 1 ms and a field strength of 1200 V/cm, the exposure of germinating barley to PEF had no significant effect on the overall metabolic activity of the seeds and the growth of the roots.

Ahmed et al. (2020) observed that applying PEF at 6 kV/cm and 50 pulses to wheat (Triticum aestivum L.) kernels improved water uptake, seed germination, and sprout growth rates. A significant increase in the total contents of phenols, DPPH, chlorophylls, carotenoids, soluble proteins, minerals and amino acids was observed in the juice of sprouts from kernels that had been exposed to PEF, compared with the juice of sprouts from kernels that had not been exposed to PEF. Based on these data, it can be concluded that PEF effectively supports wheat kernel germination and has a beneficial effect on their metabolism, improving nutrient uptake and strengthening sprouts. The application of PEF to seeds, with appropriate optimisation, can be used as a relatively simple method for producing high-quality, nutritious sprouts without the use of chemicals³⁶. Attri, P. et al.¹³ found that moderate PEF treatment damaged the seed coat, increased water absorption capacity, and enabled embryo development, thereby promoting seed germination. However, seeds exposed to high-intensity PEF exhibit a delay in germination, which can lead to embryo damage and, consequently, fewer emergent sprouts. It is therefore crucial to apply the same electric field strength to different types of seeds under stable atmospheric conditions, to standardise the PEF application procedure and support the germination process.

Research on nanoparticles (NPs) and their interactions with plants has demonstrated that NPs can inhibit germination and plant growth. Other studies indicate that NPs can be absorbed by the root system and then transported to the above-ground parts of plants through conductive tissues. Despite the findings to date, many issues related to the behaviour of nanoparticles in plant systems remain unexplained, including their phytotoxicity, which depends on surface properties, the mechanisms of tissue penetration, and the role of plant cell walls in this process³⁷.

Zainab M. Almutairi and Amjad Alharbi³⁸ analysed the effects of silver nanoparticles (AgNPs) on the germination and sprout development of three crop species: maize, watermelon, and courgette. For maize, a clear increase in germination rate was observed with higher AgNP doses, with the highest rate (6.5 seeds per day) at 1.5 mg/ml. For watermelon, the maximum germination percentage (73.33%) and the highest germination rate (1.59 seeds/day) were achieved at a concentration of 2 mg/ml. For courgettes, the most favourable results were observed at concentrations of 0.5 mg/ml (86.67%) and 2.5 mg/ml (90%), with the corresponding germination rates of 1.68 and 1.66 seeds/day, respectively.

Regulated treatment of plants with AgNPs can improve their overall growth and development. It has been observed that AgNPs reduce the weight of edible plants and vegetables (Cucurbita pepo, Allium cepa, cabbage and lettuce), but in many cases, they also induce seed germination³⁹. Khan et al.⁴⁰ showed that silver nitrate (AgNO₃) significantly (p < 0.05) reduced the root and stem lengths of pearl millet (Pennisetum glaucum) as the concentration increased from 2 mM to 4 mM to 6 mM, while silver nanoparticles (AgNPs) caused only a slight reduction in these parameters compared with the control group. At an AgNO₃ concentration of 2 mM, the root and stem lengths decreased by 41% and 21%, respectively, whereas at the same concentration of AgNPs, the reductions were 28% for the roots and 15% for the stems, suggesting a milder effect of AgNPs at lower doses. At an AgNO₃ concentration of 6 mM, a significant reduction in the root and stem lengths by 68% and 36%, respectively, was observed, compared with AgNPs, which caused a smaller reduction: 61% for the roots and 31% for the stems. Sprouts (P. glaucum L.) exposed to AgNPs exhibited less damage, which was associated with a relatively lower level of oxidative stress compared with plants treated with AgNO₃⁴⁰. Mahakham et al.⁴¹ noted that mature rice seeds treated with phytosynthesised AgNPs at concentrations of 5 and 10 ppm significantly improved germination and sprout vigour compared with the untreated control. According to Hafeez et al.⁴², the germination of “Millat”-2011 variety wheat kernels was not disturbed when concentrations ranging from 0.2 to 0.8 ppm CuNP were applied. Differences were observed at a concentration of 1.0 ppm CuNP, with a significant reduction in seed germination capacity. The application of a CuNP solution at concentrations exceeding 2 ppm was harmful to wheat plants. In contrast, Murashige-Skoog medium combined with low concentrations of Cu-NP (0.2, 0.4, 0.6, 0.8, and 1.0 ppm) significantly increased leaf area, chlorophyll content, fresh and dry weight, and dry root weight compared with control plants. According to Wang et al.⁴³, the application of CuONP had no effect on the germination capacity or germination potential of Chinese cabbage (B. pekinensis), but it had a significant effect on the viability index. The growth of the roots and the stems of B. pekinensis was promoted at a concentration of 10 mg/l CuONP, while being inhibited at a concentration of 1000 mg/l CuONP. Zafar et al.⁴⁴ demonstrated that CuONP nanoparticles affected the germination of mustard seeds and the development of their sprouts. Essa et al.⁴⁵ observed that the effect of biogenic CuNPs on wheat sprout growth depended on the dose.

According to numerous publications, the application of nanoparticles to plant seeds can have both positive and negative effects. Depending on their type and quantity, an excess of nanoparticles can have a phytotoxic effect on seeds. This phenomenon can manifest itself through reduced photosynthetic activity, the production of reactive oxygen species (ROS), DNA damage, blockage of apoplastic pathways, and consequently, impaired nutrient uptake and hydraulic transport. Furthermore, interactions between nanoparticles and plants can modify gene expression and induce genotoxic effects⁴⁶. Furthermore, studies indicate that metal nanoparticles can affect food quality by altering the mineral composition of plants and disrupting metabolism. In the case of CuNP, changes in the content of key nutritional elements, such as Fe and P, have been observed, which may affect the final nutritional value of the resulting plant products⁴⁷. However, the use of metal nanoparticles, such as AgNP and CuNP, in the production of edible plants raises significant concerns regarding food safety, primarily due to their potential uptake, accumulation, and transformation in plant tissues. Bioaccumulation of nanoparticles in plants raises serious concerns regarding their negative impact on plant health and, consequently, human health⁴⁸. Potential health effects resulting from chronic exposure cannot be overlooked either. Literature reviews indicate that both AgNPs and CuNPs may exhibit cytotoxic effects and affect the gastrointestinal tract and gut microbiota in humans after long-term consumption^49,50.

Furthermore, when AgNPs enter the environment, they can biomagnify in the food chain, inhibiting the survival, reproduction, and microbial balance of soil and aquatic organisms. Difficulties in monitoring their morphological transformation pose long-term, uncertain risks. Currently, there is a lack of data on low-dose, long-term field exposure and insufficient research on the morphological forms of AgNPs in humans and animals. Therefore, before incorporating AgNPs into agricultural practices, a systematic assessment of their environmental fate and health risks is required. This is to prevent a “technological solution” from turning into a new pollution problem⁴⁹.

At the same time, more and more studies indicate that the use of appropriate physical methods can modulate the metabolic response of plants. According to Leong et al.⁵¹ PEF treatment of soaked seeds can stimulate metabolic changes in the resulting seedlings, increasing the bioprotective potential of their shoots/sprouts and thus their value as functional foods. In their study, Leong et al.⁵¹ showed that PEF can stimulate the antioxidant metabolism of seeds by increasing glutathione levels and the activity of defense enzymes. According to Attri, P. et al.¹³, it was observed that moderate PEF intensity causes degradation of the seed coat, which increases water absorption and allows the embryo to stretch, which promotes seed germination. On the other hand, seeds exposed to high-intensity PEF treatment cause delayed germination, which may damage the embryo and, as a result, reduced seedling emergence. It was noted that PEF-induced structural changes also alter the physicochemical and functional properties of proteins/enzymes. In general, after moderate PEF treatment, a decrease in the α-helix content and an increase in the β-pleated sheet were observed, while high-intensity PEF treatment caused aggregation. Moreover, enzyme activity decreased after PEF treatment, which was significantly influenced by treatment time, native enzyme structure, PEF intensity, and preheating temperature¹³.

After the application of CuNPs at 0.06 mg/ml, growth was accelerated by 172.78 ± 23.11% and 215.94 ± 37.76% for wheat roots and stems, respectively. However, the lowest relative growth rates, 81.94 ± 11.70% and 72.46 ± 18.78% for wheat roots and stems, respectively, were observed after CuNPs were applied at 0.43 mg/ml. The results of the assessment of rapeseed germination capacity, carried out in this study, are presented in Table 1. The germination process was carried out under laboratory conditions. The results for seed germination capacity in the control object were 95% for 1 kJ/kg and 94.3% for 5.5 kJ/kg. The germination capacity results obtained after the application of AgNPs in a 25 ml/l solution and energy of 1.0 kJ/kg amounted to 88.3%. When CuNPs were applied under the same conditions, the obtained value was 96%. Compared with the control object with the same energy supplied, the germination capacity was 1% higher after CuNP application, but 6.7% lower after AgNP application. After the application of AgNPs at a concentration of 25 ml/l and an energy of 5.5 kJ/kg, the germination capacity was the same as in the control and amounted to 94.3%.

Table 1.

Effect of the solution of metal nanoparticles and pulsed electric field treatment on germination capacity of rapeseeds and on the stem and root lengths of rapeseed sprouts.

Sample	PEF energy unit	Germination capacity	Stem length	Root length
Sample	PEF energy unit	%	mm
Control	kJ/kg	94.00 ± 1.00^a	40.00 ± 1.15^a	53.00 ± 1.15^a
Control-1.0		95.00 ± 1.00^ab	40.70 ± 1.15^aa	53.67 ± 1.15^aa
Control-5.5		94.30 ± 1.15^aa	35.00 ± 1.00^ab	44.00 ± 1.00^ab
Cu25-1.0		96.00 ± 1.00^bb	60.30 ± 1.53^bb	31.67 ± 1.53^bc
Cu50-1.0		93.30 ± 1.53^bb	54.70 ± 0.58^bb	51.33 ± 1.53^bb
Cu25-5.5		95.70 ± 1.15^bb	47.70 ± 0.58^bb	75.67 ± 1.53^bc
Cu50-5.5		94.30 ± 1.53^aa	26.70 ± 1.53^bb	45.33 ± 0.58^bb
Ag25-1.0		88.30 ± 1.53^bc	55.70 ± 0.58^bb	67.67 ± 0.58^bb
Ag50-1.0		93.30 ± 0.58^ca.	48.30 ± 1.15^bc	86.00 ± 1.00^bb
Ag25-5.5		94.30 ± 0.58^aa	51.30 ± 0.58^bc	75.33 ± 1.53^bc
Ag50-5.5		94.30 ± 1.15^aa	50.30 ± 0.58^bc	89.33 ± 0.58^bc

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a, b, c – the first letter in each column indicates a comparison with the corresponding control after PEF treatment, while the second letter refers to a comparison with the initial crude control (p < 0.05); identical letters to those in controls denote no statistically significant differences.

The application of CuNPs resulted in a 95.7% increase in germination capacity, which is 1.4% higher than the control sample with the corresponding energy. Other studies showed that soaking seeds in AgNP or CuNP solutions at 50 ml/l and the application of 1.0 kJ/kg of energy resulted in a germination capacity of 93.3%, while the control sample (1.0 kJ/kg) showed a 1.7% lower germination rate. It is concluded that the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution has an adverse effect on the growth of rapeseed sprouts. As for the application of CuNPs or AgNPs in a 50 ml/l solution and energy of 5.5 kJ/kg, no changes in plant germination capacity were observed in relation to the control object, and the values obtained in both cases amounted to 94.3%.

Rapeseed sprout stem and root lengths

Table 1 provides the results concerning the length of rapeseed sprouts’ stems on the seventh day of the experiment aimed at determining seed germination capacity. In the objects in which seeds were soaked in AgNP solutions at concentrations of 25 and 50 ml/l, and with applied energies of 1.0 kJ/kg and 5.5 kJ/kg, 36.9%, 46.7%, 18.9% and 43.8% increases were observed, respectively, for the stem length compared with sprouts from the control object. In tests in which seeds were soaked in a CuNP solution at 25 ml/l, with applied energies of 1.0 kJ/kg and 5.5 kJ/kg, increases in stem length of 48.4% and 36.2%, respectively, were observed compared with the control.

After applying a CuNP solution at 50 ml/l and an energy of 1.0 kJ/kg, a 34.4% increase in stem length was observed compared with the control sample treated with the same energy level. When an energy of 5.5 kJ/kg was applied, a 23.8% reduction in stem length was observed compared with the control sample. For the AgNP solution, the greatest increase in length was observed in sprouts soaked in a 25 ml/l solution and energy of 5.5 kJ/kg. The result was 46.7% higher than that for the control. For samples soaked in CuNP solutions, the greatest increase in the length was noted for sprouts soaked in a 25 ml/l solution and an energy level of 1.0 kJ/kg. This increase was 48.4% higher compared with the control.

The same table (Table 1) shows the lengths of rapeseed sprouts’ roots on the seventh day of the experiment (Fig. 1) aimed at determining seed germination capacity. In the objects in which seeds were soaked in AgNP solutions at 25 and 50 ml/l, with applied energies of 1.0 kJ/kg and 5.5 kJ/kg, a significant increase in root length was observed relative to the control objects, by 26.1, 71.2, 60.2, and 103.0%, respectively.

After soaking the seeds in 25 ml/l CuNP solutions, a 41.0% reduction in root length was observed at 1.0 kJ/kg, and a significant 72% increase in root length at 5.5 kJ/kg compared with the control was noted. After soaking the seeds in a 50 ml/l CuNP solution, a 4.3% reduction in root length was observed at 1.0 kJ/kg, whereas an increase in root length was observed at 5.5 kJ/kg. The increase in length amounted to 3.0% compared with the control object. After soaking the seeds in 25 ml/l CuNP solutions, a 41.0% reduction in root length was observed at 1.0 kJ/kg, and a significant 72% increase in root length at 5.5 kJ/kg compared with the control was noted. After soaking the seeds in a 50 ml/l CuNP solution, a 4.3% reduction in root length was observed at 1.0 kJ/kg, whereas an increase in root length was observed at 5.5 kJ/kg. The increase in length amounted to 3.0% compared with the control object.

The contrasting effects observed between CuNPs and AgNPs may be attributed to their different modes of action in plant systems. Copper nanoparticles are known to release Cu²⁺ ions, which at higher intracellular concentrations can induce oxidative stress and inhibit cell division and elongation, particularly in root tissues. This effect may have been intensified by the application of PEF, which increases membrane permeability and facilitates the uptake of nanoparticles and metal ions. In contrast, silver nanoparticles at moderate concentrations may stimulate plant growth by enhancing water uptake and triggering adaptive metabolic responses. Therefore, the inhibitory effect observed for CuNPs (25 ml/l, 1.0 kJ/kg) and the stimulatory effect of AgNPs under similar conditions likely reflect differences in nanoparticle reactivity, ion release, and plant physiological response.

The colour of sprouts

The colour of rapeseed sprouts can be an important indicator of quality, particularly for assessing their health, vitality, and the impact of environmental conditions or treatments applied. The chlorophyll concentration in leaves is an important parameter that is regularly measured as an indicator of chloroplast content, photosynthesis, and plant metabolism. Chlorophyll is an antioxidant that is present and stored in the chloroplasts of green-leaved plants, and is primarily found in the green parts of the leaves, stems, flowers, and roots^52,53.

Chlorophyll a and chlorophyll b are essential pigments of plant photosystems⁵⁴. In addition, chlorophyll a is the main photosynthetic pigment in plants, which helps produce energy. However, the concentration of chlorophyll a in plants is 2–3 times higher than that of chlorophyll b⁵³. According to Pandey et al.⁵⁵, the application of AgNP or AgNO₃ at concentrations of 100, 500 and 1000 mg/l and 100 and 500 mg/l, respectively, on mustard seeds (Brassica juncea L.) showed that AgNPs contributed to an increase in the chlorophyll content at all concentrations under analysis, compared with the control group. Shahraki et al.⁵⁶ found that, at concentrations of 40 and 80 ppm, both silver and silver nitrate nanoparticles resulted in a reduction in the weight and the length of basil (Ocimum basilicum L.) roots and stems, as well as in the contents of chlorophyll a and chlorophyll b. Essa et al.⁴⁵ observed that after the foliar application of fertilisation at 0.06 mg/ml CuNPs to wheat leaves, the chlorophyll a and b, and total chlorophyll contents increased after 21 days of application. At the same time, a high concentration of CuNPs (0.43 mg/ml) proved to be most effective in reducing the chlorophyll (a, b and total) contents in the leaves after the same period of application. Nguyen et al.⁵⁷ observed that the application of CuNPs to maize plants increased the levels of anthocyanins, chlorophyll, and carotenoids compared with water-treated plants under drought stress.

Table 2 presents the results for the colour of rapeseed sprouts after drying. The colour parameters were determined using the CIEL*a*b* system, where the L parameter denotes the relative lightness of the sample. The results for the control rapeseed sprout colour samples showed they were relatively light, with values ranging from 53.45 to 55.90. The lowest value of 53.45 was obtained for the control sample subjected to 1.0 kJ/kg, and the highest value of 55.90 was obtained for the sample that had not been exposed to energy.

Table 2.

Analysis of rapeseed sprout colour.

Sample	PEF energy unit	L	a	B	C	h
Control	kJ/kg	55.9 ± 0.68^a	-2.76 ± 0.5^a	28.98 ± 0.67^a	29.11 ± 0.68^a	84.57 ± 0.97^a
Control-1.0		53.45 ± 0.43^ab	-4.24 ± 0.57^ad	29.33 ± 0.23^ab	29.64 ± 0.25^ad	81.77 ± 1.09^ac
Control 5.5		54.17 ± 0.51^aa	-4.21 ± 0.27^ad	33.43 ± 0.31^ab	33.7 ± 0.30^ab	82.82 ± 0.48^ab
Cu25-1.0		58.97 ± 0.19^cb	-5.55 ± 0.17^bc	27.51 ± 0.43^ba	28.07 ± 0.43^ab	78.60 ± 0.35^bf
Cu50-1.0		59.41 ± 0.39^be	-3.84 ± 0.25^bd	31.59 ± 0.55^f	31.82 ± 0.57^bb	83.07 ± 0.35^bg
Cu25-5.5		63.92 ± 0.56^bc	-4.87 ± 0.46^ab	29.84 ± 0.4^ab	30.23 ± 0.,35^de	80.72 ± 0.95^cd
Cu50-5.5		60.06 ± 0.37^ec	-4.87 ± 0.64^ab	28.1 ± 0.35^cd	28.53 ± 0.44^ab	80.19 ± 1.17^de
AG25-1.0		61.04 ± 0.31^bc	-4.96 ± 0.29^ab	32.95 ± 0.56^bc	33.32 ± 0.57^bb	81.44 ± 0.48^ac
Ag50-1.0		57.52 ± 0.31^db	-4.39 ± 0.4^ad	30.96 ± 0.28^ef	31.27 ± 0.32^bb	81.93 ± 0.67^ab
Ag25-5.5		58.43 ± 0.23^ad	-5.4 ± 0.34^bc	30.17 ± 0.28^be	30.65 ± 0.31^ef	79.86 ± 0.56^df
Ag50-5.5		58.59 ± 0.48^ab	-4.59 ± 0.14^ac	34.2 ± 0.36^ab	34.51 ± 0.35^ba	82.35 ± 0.27^ab

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a, b, c, d, e, f – the first letter in each column represents a comparison with the corresponding control after PEF application, while the second letter refers to a comparison with the initial crude control (p < 0.05); identical letters to those in the control indicate no statistically significant differences.

Upon soaking rapeseeds in AgNPs, the L parameter ranged from 57.52 (for the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution) to 61.04 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 14.20% and 7.61%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. The application of higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in increases in the parameter value of 7.86% and 8.16%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

After soaking rapeseeds in CuNPs, the L parameter value ranged from 58.97 (for the application of energy of 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 63.92 (for the application of energy of 5.5 kJ/kg and a 25 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the L parameter by 10.33% and 11.15%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l copper nanoparticle solutions increased the L parameter by 18.00% and 10.87%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy. When comparing the analysed samples to the control where sprouts have not been exposed to energy, the parameter in question (L) shows an increase of 5.49% (1 kJ/kg Cu25); 6.28% (1.0 kJ/kg Cu50); 14.35% (5.5 kJ/kg Cu25); 7.44(5.5 kJ/kg Cu50); 9.19% (1.0 kJ/kg Ag25); 2.90%(1.0 kJ/kg Ag50); 4.53%(5.5 kJ/kg Ag25) and 4.81% (5.5 kJ/kg Ag50), respectively. This parameter is relative and refers to the mixing of the base colour with black (low L) or white (high L). For control objects treated with 1.0 kJ/kg and 5.5 kJ/kg, a decrease in L values was observed compared with the control not subjected to the electric field.

Parameters a and b determine the colour of the sample. Negative values of a indicate the proportion of the green colour in the overall colour, while positive values indicate the proportion of the red colour. Negative values of parameter a in the range from − 5.55 for the 1.0 kJ/kg Cu25 object to -2.76 for the control sample without the application of energy indicate the proportion of the green colour in all samples in the overall colour of rapeseed sprouts. In all objects, an increase was observed in the parameter value by 101.09% (1.0 kJ/kg Cu25); 39.13% (1.0 kJ/kg Cu50); 76.45% (5.5 kJ/kg Cu25); 76.45% (5.5 kJ/kg Cu50); 79.71% (1.0 kJ/kg Ag25); 59.06% (1.0 kJ/kg Ag50); 95.65% (5.5 kJ/kg Ag25) and 66.30% (5.5 kJ/kg Ag50), respectively, compared with the control object without the application of energy. Lower values of the parameter a indicate a greater proportion of the green colour in the overall colour.

After soaking rapeseeds in AgNPs, the value of parameter a ranged from − 5.4 (for the application of energy of 5.5 kJ/kg and a 25 ml/l nanoparticle solution) to -4.39 (for the application of energy of 1.0 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 16.98% and 3.54%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. Moreover, the application of higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 28.27% and 9.03%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy. Following rapeseed soaking in CuNPs, the value of parameter a ranged from − 5.55 (for the application of energy of 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to -3.84 (for the application of energy of 1.0 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 30.90% and 9.43%, respectively, compared with the control sample subjected to 1 kJ/kg energy. The application of higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 15.68% and 15.68%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy. In the case of parameter b, negative values indicate the proportion of the blue colour, while positive values indicate the proportion of the yellow colour. Analysis of this parameter b showed the proportion of the yellow colour in all samples in the overall colour of rapeseed sprouts. Analysis of the results obtained revealed a decrease in the parameter concerned of 5.07% (1.0 kJ/kg Cu25) and 3.04% (5.5 kJ/kg Cu50), and an increase of 9.01% (1.0 kJ/kg Cu50); 2.97% (5.5 kJ/kg Cu25), 13.70% (1.0 kJ/kg Ag25); 6.83% (1.0 kJ/kg Ag50); 4.11% (5.5 kJ/kg Ag25); 18.01% (5.5 kJ/kg Ag50), as compared with the control object not exposed to energy.

After soaking rapeseeds in AgNPs, the value of parameter b ranged from 30.17 (for energy applied at 5.5 kJ/kg and a 25 ml/l nanoparticle solution) to 34.2 (for energy applied at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions was characterised by increases in the parameter in question of 12.34% and 5.56%, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. Moreover, the application of higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions contributed to a decrease in the value of parameter b of 9.75% and an increase of 2.30%, respectively, compared with the control sample subjected to energy at 5.5 kJ/kg. Following rapeseed soaking in CuNPs, the value of parameter b ranged from 27.51 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 31.59 (for the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in a decrease of 6.21% and an increase of 7.71%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. However, the application of higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions showed a decrease in the parameter value of 10.74% and 15.94%, respectively, compared with the control sample subjected to energy at 5.5 kJ/kg.

Parameter c indicates colour saturation. Considering the control object without the energy applied, decreases in the parameter value of 3.57% for 1.0 kJ/kg Cu25, and of 1.99% for 5.5 kJ/kg Cu50 and increases of 9.31% (1.0 kJ/kg Cu50); 3.85% (5.5 kJ/kg Cu25); 14.46% (1.0 kJ/kg Ag25); 7.42% (1 kJ/kg Ag50); 5.29% (5.5 kJ/kg Ag25); 18.55% (5.5 kJ/kg Ag50) were observed, respectively.

After soaking rapeseeds in AgNPs, the value of parameter c ranged from 30.65 (for the application of energy at 5.5 kJ/kg and a 25 ml/l of nanoparticle solution) to 34.51 (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) and nanoparticle solutions at 25 and 50 ml/l increased the parameter value by 12.42% and 5.50%, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases of 9.05% and 2.40%, respectively, in the parameter value, compared with the control sample subjected to energy at 5.5 kJ/kg.

After soaking rapeseeds in CuNPs, the value of parameter c ranged from 28.07 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 31.82 (for the application of energy at 1 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases of 5.30% and 7.35%, respectively, in the parameter value, compared with the control sample subjected to energy at 1.0 kJ/kg.

Moreover, higher energy values (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions resulted in decreases in the parameter value of 10.30% and 15.34%, respectively, compared with the control sample subjected to energy at 5.5 kJ/kg.

Parameter h denotes the colour hue, which is determined after converting colours in the L*a*b* system into the Cartesian system. This parameter’s value indicates the shade of yellow. In all objects, a decrease in this parameter was observed, respectively, of 7.06% (1.0 kJ/kg Cu25); 1.77% (1.0 kJ/kg Cu50); 4.55% (5.5 kJ/kg Cu25); 5.18% (5.5 kJ/kg Cu50); 3.70% (1 kJ/kg Ag25); 3.12% (1.0 kJ/kg Ag50); 5.57% (5.5 kJ/kg Ag25); 2.63% (5.5 kJ/kg Ag50), as compared with the control object not exposed to energy.

After soaking rapeseeds in AgNPs, the value of parameter h ranged from 79.86 (for the application of energy at 5.5 kJ/kg and a 25 ml/l of nanoparticle solution) to 82.35 (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by a minimal decrease in the value of the parameter concerned of 0.4% for 25 ml/l, and a slight increase of 0.2% for 50 ml/l, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases in the parameter value of 3.57% and 0.57%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy. Following rapeseed soaking in CuNPs, the value of parameter h ranged from 78.60 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 83.07 (for the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 3.88% and increased it by 1.59%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 2.54% and 3.18%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy. For control objects of 1.0 kJ/kg and 5.5 kJ/kg, a decrease in the value of the analysed parameter was observed in relation to the control not subjected to an electric field.

Chlorophyll and carotenoid contents

Analysis of chlorophyll a content (Table 3) in rapeseed sprouts showed that it ranged from 37.6 mg/100 g DM in the sample treated with 5.5 kJ/kg energy and 50 ml/l of copper nanoparticles to 42.3 mg/100 g DM in the control sample not exposed to energy. In all samples, a decrease in chlorophyll a levels was noted in relation to the control object not exposed to energy, of 4.49% (1.0 kJ/kg Cu25); 1.89% (1.0 kJ/kg Cu50); 6.38% (5.5 kJ/kg Cu25); 11.11% (5.5 kJ/kg Cu50); 8.75% (1.0 kJ/kg Ag25); 7.80% (1 kJ/kg Ag50); 6.62% (5.5 kJ/kg Ag25) and 10.40% (5.5 kJ/kg Ag50), respectively. The largest decrease was observed when applying energy at 5.5 kJ/kg and copper nanoparticles in a 50 ml/l solution (11.11%).

Table 3.

Contents of chlorophylls and carotenoids in rapeseed sprouts.

Sample	PEF energy, unit	Chlorophyll a	Chlorophyll b	∑ Carotenoids
Sample	PEF energy, unit	mg/100 g DM
Control	kJ/kg	42.3 ± 0.39^a	17.6 ± 0.04^a	28.5 ± 0.11^a
Control-1.0		40.1 ± 0.45^ac	17.9 ± 0.11^aa	27.5 ± 0.22^ab
Control-5.5		39.4 ± 0.49^ab	17.2 ± 0.15^aa	28.7 ± 0.09^ab
Cu25-1.0		40.4 ± 0.30^aa	16.8 ± 0.15^bb	27.2 ± 0.19^ab
Cu50-1.0		41.5 ± 0.29^bb	16.1 ± 0.17^ac	26.6 ± 0.19^bc
Cu25-5.5		39.6 ± 0.23^ab	15,3 ± 0,11^eb	26.3 ± 0.23^bc
Cu50-5.5		37.6 ± 0.18^bb	15,8 ± 0,18^cb	25.6 ± 0.27^ab
AG25-1.0		38.6 ± 0.49^bc	16,9 ± 0,10^ba	26.8 ± 0.08^cd
Ag50-1.0		39.0 ± 0.57^ad	16.7 ± 0.18^bc	26.4 ± 0.13^bb
Ag25-5.5		39.5 ± 0.34^ab	16.0 ± 0.11^ac	25.4 ± 0.26^ab
Ag50-5.5		37.9 ± 0.58^be	16.2 ± 0.04^ab	25.6 ± 0.17^ab

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a, b, c, d—the first letter in each column indicates a comparison with the corresponding control after PEF treatment, while the second letter refers to a comparison with the initial crude control (p < 0.05); identical letters to those in the control denote no significant differences.

After soaking rapeseeds in AgNPs, the value of chlorophyll a ranged from 37.9 (for the application of energy at 5.5 kJ/kg and a 50 ml/l of nanoparticle solution) to 39.5 mg/100 g DM (for the application of energy at 5.5 kJ/kg and a 25 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by a decrease in the value of the parameter concerned of 3.74% for 25 ml/l and 2.74% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions resulted in minimal increases of 0.25% and 3.81%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

After soaking rapeseeds in CuNPs, the chlorophyll a content ranged from 27.6 (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution) to 41.5 mg/100 g DM (for the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions increased the parameter value by 0.75% and 3.49%, respectively, compared with the control sample subjected to 1.0 kJ/kg. However, a higher energy value (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions resulted in a minimal increase in the parameter value of 0.51%, and a decrease of 4.57%, respectively, compared with the control sample subjected to energy at 5.5 kJ/kg.

Analysis of the chlorophyll b content in rapeseed sprouts showed the values to fall within the range from 15.5 in the sample when applying energy at 5.5 kJ/kg and 25 ml/l of copper nanoparticles, to 17.9 mg/100 g DM in the control sample when applying a pulsed electric field of 1 kJ/kg. In the control sample, when applying a PEF of 1 kJ/kg, a minimal increase in chlorophyll b levels of 1.70% was observed in relation to the control object without energy exposure, whereas a decrease of 2.27% was observed when applying a pulsed electric field of 5.5 kJ/kg. In subsequent tests, a decrease in chlorophyll b levels was observed in relation to the control object without the application of energy, of 4.55% (1.0 kJ/kg Cu25); 8.52% (1.0 kJ/kg Cu50); 13.07% (5.5 kJ/kg Cu25); 10.23% (5.5 kJ/kg Cu50); 3.98% (1.0 kJ/kg Ag25); 5.11% (1.0 kJ/kg Ag50); 9.09% (5.5 kJ/kg Ag25) and 7.95% (5.5 kJ/kg Ag50), respectively.

After soaking rapeseeds in AgNPs, the chlorophyll b ranged from 16.0 (for the application of energy at 5.5 kJ/kg and a 25 ml/l of nanoparticle solution) to 16.9 mg/100 g DM (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by a decrease in the value of the parameter concerned of 5.59% for 25 ml/l and 6.70% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. The application of higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases in the parameter value of 6.98% and 5.81%, respectively, compared with the control sample subjected to 5.5 kJ/kg.

After soaking rapeseeds in CuNPs, the chlorophyll b content ranged from 15.3 (for the application of energy at 5.5 kJ/kg and a 25 ml/l nanoparticle solution) to 16.8 mg/100 g DM (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases of 6.15% and 10.06%, respectively, in the parameter value compared with the control sample subjected to 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 11.05% and 8.14%, respectively, compared with the control sample subjected to 5.5 kJ/kg.

Analysis of the carotenoid content in rapeseed sprouts showed values ranging from 25.4 mg/100 g DM in the sample with energy at 1.0 kJ/kg and 25 ml/l of silver nanoparticles to 28.5 mg/100 g DM in the control sample not exposed to energy. In the control sample, when applying a PEF of 5.5 kJ/kg, a minimal increase of 0.70% in carotenoid levels was observed compared with the control object without energy exposure. In subsequent tests, a decrease in carotenoid levels was observed in relation to the control object without the application of energy, of 4.56% (1.0 kJ/kg Cu25); 6.67% (1.0 kJ/kg Cu50); 7.72% (5.5 kJ/kg Cu25); 10.18% (5.5 kJ/kg Cu50); 5.96% (1.0 kJ/kg Ag25); 7.37% (1.0 kJ/kg Ag50); 10.88% (5.5 kJ/kg Ag25) and 10.18% (5.5 kJ/kg Ag50), respectively.

After soaking rapeseeds in AgNPs, the carotenoid content ranged from 25.4 (for the application of energy at 5.5 kJ/kg and a 25 ml/l nanoparticle solution) to 26.8 mg/100 g DM (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by a decrease in the value of the parameter concerned of 2.25% for 25 ml/l and 4.0% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 11.50% and 10.80%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

After soaking rapeseeds in CuNPs, the carotenoid content ranged from 25.6 (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution) to 27.2 mg/100 g DM (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases in the parameter value of 1.09% and 3.27%, respectively, compared with the control sample subjected to 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) and 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 8.36% and 10.80%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

Antioxidants

Table 4 presents the results of determinations of total oxidative activity, polyphenol content, phenolic compound content, and antioxidant activity against ABTS and DPPH radicals in rapeseed sprouts.

Table 4.

Antioxidant capacity of rapeseed sprouts.

Name	Unit	FRAP	Polyphenols	TPC	ABTS	DPPH
Name	Unit	[mg TE/g DM]	[mg GAE/g DW]		[µmol trolox/g DW]
Control	kJ/kg	81.89 ± 0.20^a	14.23 ± 0.02^a	24.50 ± 0.25^a	93.80 ± 0.32^a	58.80 ± 0.37^a
Control-1.0		82.46 ± 0.09^ab	14.36 ± 0.03^aa	25.40 ± 0.13^ab	95.50 ± 0.29^ab	59.20 ± 0.5^aa
Control-5.5		83.83 ± 0.26^ab	14.43 ± 0.02^aa	25.60 ± 0.26^aa	94.90 ± 0.29^ab	58.60 ± 0.51^aa
Cu25-1.0		95.93 ± 0.27^bc	14.95 ± 0.02^aa	24.70 ± 0.11^bd	87.30 ± 0.51^bc	58.70 ± 0.28^aa
Cu50-1.0		196.74 ± 0.60^b	14.53 ± 0.02^aa	25.00 ± 0.27^aa	96.20 ± 0.70^ab	60.10 ± 0.29^bb
Cu25-5.5		97.75 ± 0.24^bc	14.94 ± 0.03^aa	25.20 ± 0.19^aa	96.20 ± 0.53^bc	60.70 ± 0.28^bb
Cu50-5.5		198.52 ± 0.32^bc	14.63 ± 0.02^aa	25.60 ± 0.24^ab	96.20 ± 0.79^bc	60.10 ± 0.37^bb
AG25-1.0		89.61 ± 0.13^bc	15.91 ± 0.03^bb	25.60 ± 0.19^ab	98.30 ± 0.43^bc	59.90 ± 0.31^ac
Ag50-1.0		186.29 ± 0.33^bc	15.47 ± 0.02^bb	26.30 ± 0.16^bc	98.20 ± 0.67^bc	63.20 ± 0.33^ab
Ag25-5.5		91.13 ± 0.18^bc	15.97 ± 0.02^bb	26.50 ± 0.19^bb	99.30 ± 0.52^cd	63.10 ± 0.38^bb
Ag50-5.5		188.94 ± 0.72^bc	15.55 ± 0.03^bb	27.00 ± 0.21^bc	98.80 ± 0.60^bc	64.50 ± 0.29^bc

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a, b, c, d—the first letter in each column denotes a comparison with the corresponding control after PEF treatment, whereas the second letter refers to a comparison with the initial crude control (p < 0.05); identical letters to those in the control indicate no significant differences.

The FRAP method measures a sample’s ability to reduce Fe³⁺ in the Fe³⁺-TPTZ complex to Fe²⁺. The total oxidative activity potential (FRAP) (Table 4) in the tested material ranged from 81.89 in the control sample to 198.52 mg TE/g DM when applying CuNPs in a 50 ml/l solution, and applying energy at 1.0 kJ/kg. In all variants of the experiment, the application of PEF and seed soaking in AgNPs and CuNPs increased total oxidative activity in the analysed rapeseed sprouts. The application of PEF (1.0 kJ/kg and 5.5 kJ/kg) in the control samples resulted in an increase in the analysed parameter in both variants of the experiment by 0.70% (control − 1 kJ/kg) and 2.37% (control − 5.5 kJ/kg), respectively, in relation to the untreated control sample. The highest increase in total oxidative activity was recorded for the sample subjected to 1.0 kJ/kg and seed soaking in a CuNP solution at 50 ml/l (140.25%) compared with the untreated control sample. When energy of 5.5 kJ/kg and CuNPs (25 and 50 ml/l) were applied, the increase in oxidative activity was 97.75 TE/g DM (16.61%) and 198.5 TE/g DM (136.81%), as compared with the control sample subjected to energy at 5.5 kJ/kg. The application of 1.0 kJ/kg of energy to rapeseeds and 50 ml/l AgNP resulted in a 125.92% increase in total activity compared with the corresponding control sample. In addition, the applied value of 5.5 kJ/kg PEF in both AgNP solutions resulted in values higher than those of the control samples by 8.71% (Ag25) and 125.38% (Ag50), respectively.

The relatively high FRAP values observed in some treatments, particularly those involving CuNPs, should be interpreted with caution. Although the assay was performed on methanolic extracts of plant material, the presence of residual nanoparticles cannot be entirely excluded. Metallic nanoparticles, especially copper-based ones, may exhibit redox activity and potentially contribute to the reduction of the Fe³⁺–TPTZ complex used in the FRAP assay. Therefore, part of the observed increase in antioxidant capacity may be associated with analytical interference. Future studies should include nanoparticle-only controls and complementary methods to better distinguish between true biological effects and potential assay artefacts.

It should be noted that the relatively high FRAP values observed in some treatments (e.g., Cu50) were carefully verified and were consistent across all replicates. The FRAP assay was performed on methanolic extracts of freeze-dried plant material rather than on nanoparticle suspensions, which significantly limits the possibility of direct optical interference from nanoparticles. Moreover, similar increasing trends were observed for other antioxidant-related parameters (total polyphenols and TPC), supporting a biological origin of the effect. Nevertheless, potential interactions between residual nanoparticles and the FRAP reagent cannot be completely excluded, and this aspect should be considered in future studies using additional controls or complementary analytical methods.

The polyphenol content of rapeseeds and sprouts is determined by the environmental conditions and genetic background. The available literature is dominated by studies on the adaptation of rapeseeds to drought stress, which is becoming increasingly common worldwide, and is the main constraint on crop productivity in many arid and semi-arid regions^58,59. According to Ayyaz et al.⁶⁰, the presence of polyphenols contributes to ROS scavenging, mitigates the adverse effects of drought, and improves rapeseed drought tolerance. In the current study, the polyphenol content of rapeseed sprouts (Table 4) ranged from 14.23 mg GAE/g DW for the control sample without PEF interference in the seeds to 15.97 mg GAE/g DW for the sample after the application of CuNPs at 25 ml/l and energy at 5.5 kJ/kg. In all variants of the experiment, an increase in polyphenol content was observed compared with the control samples. In the current study, the largest increase in polyphenols in sprouts was recorded for samples subjected to energy at 1.0 kJ/kg and 5.5 kJ/kg, when applying CuNPs in both solutions (25 and 50 ml/l). The results obtained were higher than those for the corresponding control samples by 4.11% (1.0 kJ/kg Cu25), 1.18% (1.0 kJ/kg Cu50), 3.53% (5.5 kJ/kg Cu25), and 1.39% (5.5 kJ/kg Cu50), respectively.

Phenolic compounds (TPC) in plants are one of the main groups of compounds that act as basic antioxidants or free radical scavengers. They are among the most important lipid components associated with rapeseed plants. They have gained increased attention due to their numerous bioactive and health benefits. These include antioxidant, anti-inflammatory, antineoplastic, antimicrobial, antidiabetic, hypolipidemic, and anti-obesity effects^61,62. Knowledge of the phenolic profile of both rapeseed and its sprouts can provide insight into its physiological activity, help identify adulterated rapeseed products, and contribute to the discovery of new natural antioxidant compounds⁶². Free and bound phenolic compounds have different bioavailability and different health effects. The effect of germination on the phenolic content and antioxidant activity has been studied in many species, including Brassicaceae species, such as radish and broccoli, and the visible differences are closely related to the species, but also to the stage of sprout growth (i.e. from three days to two weeks after the start of incubation)^2,63. Currently, the available literature on the phenolic content and antioxidant activity of its sprouts is insufficient⁶⁴, whereas the germination of this species has been thoroughly studied. In the current study, the overall phenolic compound content was higher than that reported by Falcinelli et al.⁶⁵ and Benincasa et al.⁷, in which rapeseed sprouts were tested for resistance to salinisation of the growth environment. However, it is difficult to discuss the total phenolic content in the current study’s sprout samples after PEF treatment and seed soaking in nanoparticles because there is no similar treatment reported in the literature on rapeseed sprout cultivation.

An analysis of the TPC content in rapeseed sprouts in the current study showed that it ranged from 24.5 mg GAE/g DW in the control sample without energy application to 27.00 mg GAE/g DW when applying energy at 5.5 kJ/kg and AgNPs in a 50 ml/l solution. In all samples, an increase in the phenolic compound content was observed in relation to the untreated control object by 0.82% (1.0 kJ/kg Cu25); 2.04% (1.0 kJ/kg Cu50); 2.86% (5.5 kJ/kg Cu25); 4.49% (5.5 kJ/kg Cu50); 4.49% (1.0 kJ/kg Ag25); 7.35% (1.0 kJ/kg Ag50); 8.16% (5.5 kJ/kg Ag25) and 10.20% (5.5 kJ/kg Ag50), respectively.

After soaking rapeseeds in AgNPs, the phenolic compound content ranged from 25.6 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 27.0 mg GAE/g DM (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by an increase in the value of the parameter concerned by 0.79% for 25 ml/l and 3.54% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions increased the parameter value by 3.52% and 5.47%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

Following rapeseed soaking in CuNPs, the phenolic compound content ranged from 24.7 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 25.6 mg GAE/g DM (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in decreases in the parameter value of 2.73% and 1.57%, respectively, compared with the control sample subjected to 1.0 kJ/kg. However, a higher energy value (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions caused a decrease in the value of the parameter concerned of 1.56% for 25 ml/l, whereas for 50 ml/l, it remained at the same level in relation to the control sample subjected to 5.5 kJ/kg energy.

Analysis of the antioxidant capacity of ABTS in rapeseed sprouts showed that values ranged from 87.3 µmol Trolox/g DM when applying energy at 1.0 kJ/kg and copper nanoparticles at 25 ml/l, to 99.3 µmol Trolox/g DM when applying energy at 5.5 kJ/kg and silver nanoparticles at 25 ml/l. In subsequent tests, an increase in the level of the analysed parameter was observed in relation to the control object not exposed to energy, of 2.56% (1.0 kJ/kg Cu50), 2.56% (5.5 kJ/kg Cu25), 2.56% (5.5 kJ/kg Cu50) 4.80% (1.0 kJ/kg Ag25); 4.69% (1.0 kJ/kg Ag50); 5.86% (5.5 kJ/kg Ag25) and 5.33% (5.5 kJ/kg Ag50), respectively, and a decrease of 6.93% (1 kJ/kg Cu25).

After soaking rapeseeds in AgNPs, the antioxidant capacity ranged from 98.2 (for the application of energy at 1.0 kJ/kg and a 50 ml/l nanoparticle solution) to 99.8 µmol trolox/g DM (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by an increase in the value of the parameter concerned by 2.93% for 25 ml/l and 2.83% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, the application of higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions increased the parameter value by 4.64% and 4.11%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

Upon soaking rapeseeds in CuNPs, the antioxidant capacity ranged from 87.3 (for the application of energy at 1.0 kJ/kg and a 25 ml/l nanoparticle solution) to 96.2 µmol trolox/g DM (for the application of energy at 1.0 and 5.5 kJ/kg as well as 25 and 50 ml/l nanoparticle solutions). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions decreased the parameter value by 8.59% and increased it by 0.73%, respectively, compared with the control sample subjected to 1.0 kJ/kg energy. However, higher energy values (5.5 kJ/kg) and nanoparticle solutions at 25 and 50 ml/l increased the parameter value by 1.37% and 1.37%, respectively, compared with the control sample subjected to energy at 5.5 kJ/kg.

According to Guo et al.⁶⁶, during the germination phase, DPPH radical scavenging activity increases, probably due to the increased total contents of phenols and melatonin, and total isoflavone content. According to Pająk et al.⁶³, the antioxidant activity of other seeds usually increases during germination. As for mung bean sprouts, the antioxidant activity value increases almost tenfold, while for sunflower seeds, it increases twofold. Broccoli and radish sprouts exhibited approximately 20% and 40% (respectively) higher reducing capacity than their seeds. Analysis of the overall DPPH antioxidant activity in rapeseed sprouts showed values ranging from 58.6 µmol Trolox/g DM in the control sample to 64.5 µmol Trolox/g DM when applying energy at 5.5 kJ/kg and silver nanoparticles at 50 ml/l. In the control sample, when applying PEF of 1.0 kJ/kg, a minimal increase in the parameter of 0.68% was observed, whereas for 5.5 kJ/kg, a minimal decrease in the overall antioxidant activity of 0.34% was observed in relation to the control object without energy application. In subsequent tests, an increase in the level of the analysed parameter was observed in relation to the control object not subjected to energy, respectively of 2.21% (1.0 kJ/kg Cu50); 3.23%(5.5 kJ/kg Cu25); 2.21% (5.5 kJ/kg Cu50); 1.87% (1.0 kJ/kg Ag25); 7.48% (1.0 kJ/kg Ag50); 7.31% (5.5 kJ/kg Ag25) and 9.69% (5.5 kJ/kg Ag50), while the minimum decrease of 0.17% was observed for 1.0 kJ/kg Cu25.

Following rapeseed soaking in AgNPs, the overall antioxidant capacity (DPPH) ranged from 59.9 (for the application of energy at 1.0 kJ/kg and a 50 ml/l of nanoparticle solution) to 64.5 µmol trolox/g DM (for the application of energy at 5.5 kJ/kg and a 50 ml/l nanoparticle solution). The application of a lower energy value (1.0 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions was characterised by an increase in the value of the parameter concerned of 1.18% for 25 ml/l and 6.76% for 50 ml/l, respectively, compared with the control sample subjected to energy at 1.0 kJ/kg. However, higher energy (5.5 kJ/kg) as well as 25 and 50 ml/l nanoparticle solutions increased the parameter value by 7.68% and 10.07%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

After soaking rapeseeds in CuNPs, the overall antioxidant activity (DPPH) ranged from 58.7 (for the application of energy at 1.0 kJ/kg and a 25 ml/l of nanoparticle solution) to 60.7 µmol trolox/g DM (for the application of energy at 5.5 kJ/kg and a 25 ml/l nanoparticle solution). The application of lower energy (1.0 kJ/kg) and 25 and 50 ml/l nanoparticle solutions resulted in minimal decreases of 0.84% and 1.52%, respectively, in the parameter value compared with the control sample subjected to 1.0 kJ/kg energy. In contrast, a higher energy value (5.5 kJ/kg), as well as 25 and 50 ml/l nanoparticle solutions, increased the parameter value by 3.58% and 2.56%, respectively, compared with the control sample subjected to 5.5 kJ/kg energy.

Study limitations

This study has certain limitations that should be considered when interpreting the obtained results.

First, the lack of a dedicated control group containing only nanoparticle solutions prevents the complete exclusion of potential interference effects. In particular, it cannot be ruled out that the presence of metal nanoparticles, especially CuNP, could have influenced the results of antioxidant property assays (e.g., FRAP), leading to potential overestimation. We also emphasize that future studies should include dedicated control samples containing only nanoparticles and employ complementary analytical methods, which will enable clear differentiation between actual biological effects and potential analytical artifacts related to the presence of nanoparticles.

Second, the lack of a detailed physicochemical characterization of the nanoparticles in the working solution used, including, among other things, their toxicity. DLS (Dynamic Light Scattering), NTA (Nanoparticle Tracking Analysis), and quantitative TEM microscopy data limit the ability to clearly assess the nanoparticles’ hydrodynamic diameter, stability, and potential aggregation under experimental conditions. These elements can significantly impact their absorption and biological activity. The lack of this data also hinders the comparison of the obtained results with other studies and limits the ability to fully interpret the observed physiological effects.

Furthermore, this study did not directly assess the accumulation of nanoparticles or ionic species in plant tissues, preventing a clear determination of the relationship between the applied dose and actual biological exposure. This may be important when interpreting stimulating and potentially toxic effects.

Taking these limitations into account is essential for a more objective interpretation of the results. They also point to the direction of future research, which includes the introduction of appropriate control samples and expanded characterization of metal nanoparticles, as well as analysis of metal accumulation in plant tissues.

Conclusion

The study demonstrated that the application of PEF and metal nanoparticles (AgNPs and CuNPs) on rapeseeds significantly affected germination capacity, sprout growth and antioxidant properties. The most important conclusions are as follows:

The application of the PEF method alone contributed to an increase in the germination capacity of 1.0% and 0.3% for energies of 1.0 and 5.5 kJ/kg, respectively, compared with the control with no PEF applied.
CuNPs in a 25 ml/l solution had a positive effect on germination capacity for both energies applied, compared with the control objects; for 1.0 kJ/kg, a 1% increase, and for 5.5 kJ/kg, a 1.4% increase.
CuNPs and AgNPs in a 50 ml/l solution at 5.5 kJ/kg had no effect on stimulating germination capacity, compared with the control with the application of the appropriate energy.
In the case of stem length, the application of the PEF method alone contributed to an increase of 1.67% and a decrease of 12.50% for 1.0 and 5.5 kJ/kg, respectively, compared with the control without treatment.
The root length increased by 1.26% and decreased by 16.98% after the application of 1.0 and 5.5 kJ/kg. Similar results were obtained for the stem length.
AgNPs had a positive effect on root elongation in rapeseed sprouts, with the greatest elongation for the 50 ml/l solution at 5.5 kJ/kg, i.e., a 103.0% increase compared with the control object, and a 68.55% increase compared with the untreated control.
The application of both methods resulted in a decrease in chlorophyll a and b and carotenoid contents in rapeseed sprouts.

The application of PEF and seed coating in CuNPs or AgNPs had a positive effect on the antioxidant activity (FRAP and polyphenols) in the analysed sprouts.

8.
The most favourable energy level and nanometal solution in the FRAP test was 5.5 kJ/kg and CuNPs at 50 ml/l, which resulted in the largest increase in FRAP values of 136.81% compared with the control variant, and of 142.42% compared with the control sample with no PEF applied.
9.
The highest increase in the polyphenol content was observed after the application of AgNPs in a 25 ml/l solution at 1.0 and 5.5 kJ/kg (increases of approximately 10–12%, compared with the control).

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary Material 1^{(15.7KB, docx)}

Supplementary Material 2^{(15.2KB, docx)}

Supplementary Material 3^{(16.1KB, docx)}

Supplementary Material 4^{(15.3KB, docx)}

Author contributions

Methodology: M.K-G., M.S., M.K., K.S.; Investigation: M.K-G., M.S., K.S.; Supervision: M.K-G., M.K., K.S.; Writing – original draft: M.K-G., K.S.; Writing – review & editing: M.K., M.K-G., K.S.; Review and editing: M.K-G., K.S., M.S.

Data availability

The data that support the findings of this study are available on request from the corresponding author: Marta Krajewska ( [marta.krajewska@up.edu.pl](mailto: marta.krajewska@up.edu.pl) ) and Magdalena Kachel-Górecka (magdalena.kachel@up.edu.pl).

Declarations

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Contributor Information

Magdalena Kachel-Górecka, Email: magdalena.kachel@up.edu.pl.

Marta Krajewska, Email: marta.krajewska@up.edu.pl.

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Associated Data

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Supplementary Materials

Supplementary Material 1^{(15.7KB, docx)}

Supplementary Material 2^{(15.2KB, docx)}

Supplementary Material 3^{(16.1KB, docx)}

Supplementary Material 4^{(15.3KB, docx)}

Data Availability Statement

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[CR63] 63.Pająk, P., Socha, R., Gałkowska, D., Rożnowski, J. & Fortuna, T. Phenolic profile and antioxidant activity in selected seeds and sprouts. Food Chem.143, 300–306 (2014). [DOI] [PubMed] [Google Scholar]

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[CR65] 65.Falcinelli, B. et al. Germination under moderate salinity increases phenolic content and antioxidant activity in rapeseed (Brassica napus var oleifera Del.) sprouts. Molecules22, 1–13 (2017). [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR66] 66.Guo, S., Ge, Y. & Na Jom, K. A review of phytochemistry, metabolite changes, and medicinal uses of the common sunflower seed and sprouts (Helianthus annuus L). Chem. Cent. J.11, 1–10 (2017). [DOI] [PMC free article] [PubMed] [Google Scholar]

PERMALINK

Effects of pulsed electric field and metal nanoparticle soaking on germination, growth, and antioxidant properties of rapeseed sprouts

Karolina Sokal

Magdalena Kachel-Górecka

Małgorzata Stryjecka

Marta Krajewska

Abstract

Supplementary Information

Introduction

Materials and methods

Study material

Silver and copper nanoparticles

Sample imaging using transmission electron microscopy

Seed preparation for testing

Drying method

Colour measurement

Antiradical activity

Total carotenoid and chlorophyll contents

FRAP analysis

Polyphenols

Statistical analysis

Results and discussion

Transmission electron microscopy (TEM) tests

Fig. 1.

Fig. 2.

Fig. 3.

Assessment of rapeseed germination capacity

Table 1.

Rapeseed sprout stem and root lengths

The colour of sprouts

Table 2.

Chlorophyll and carotenoid contents

Table 3.

Antioxidants

Table 4.

Study limitations

Conclusion

Supplementary Information

Author contributions

Data availability

Declarations

Competing interests

Footnotes

Contributor Information

References

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

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