Nutritional Composition, Bioactive Components and Antioxidant Activity of Garden Cress (Lepidium sativum L.) Grown Under Deficit Irrigation

Abstract

This study examined how different water restriction levels (T100%, T85%, T75%, and T55%) influence the nutritional and bioactive compounds of Bahar and Dadaş cress (Lepidium sativum L.) cultivars. The highest levels of phenolic compounds found in Dadaş and Bahar cress were quercetin (8.33 ± 0.23–9.32 ± 0.25 µg/L), ferulic acid (8.08 ± 0.18–8.42 ± 0.19 µg/L), catechin (6.83 ± 0.28 µg/L), and caftaric acid (5.40 ± 0.45 µg/L). Mild and moderate drought treatments (85% and 75% humidity) caused notable increases in phenolic compounds. The highest antioxidant enzyme levels were observed as GST, 6GPD, and G6PD in Bahar and Dadaş cress, with enzyme levels rising under drought conditions. Notably, the mild drought treatment roughly doubled peonidin-3-glucoside acetyl levels in the cress cultivars. Sugar contents of Dadaş and Bahar cress cultivars also rose significantly with drought treatment. Riboflavin, the most abundant vitamin in cress cultivars, increased to 40.96 ± 1.24 mg/kg in Dadaş and 30.79 ± 1.60 mg/kg in Bahar cress under drought stress. Amino acids showed the highest increases under severe drought, with asparagine rising by roughly 2.76-fold and leucine increasing by 2.67-fold in Bahar cress. These findings suggest that controlled water restriction can enhance the nutritional and bioactive properties of cress, potentially leading to more nutrient-rich products for the food industry and human health.

1. Introduction

Along with global climate anomalies and ecological imbalance, drought is one of the most significant environmental stressors limiting agricultural production [1]. Physiological water deficit is generally defined as the insufficient amount of water in the soil to meet plant needs, negatively affecting their metabolism [2]. Climate change-related factors such as changes in precipitation patterns, rising atmospheric temperatures, and dry winds are increasing the severity and frequency of drought [3]. This stressor profoundly impacts the morphological, physiological, biochemical, and molecular characteristics of plants, leading to significant yield losses [3,4,5]. Drought stress can negatively affect every stage of plant life, from germination to maturity [2]. Therefore, plants activate a complex array of defense mechanisms to survive [6]. The effects of drought on plant morphology include plant height, leaf area, and leaf yellowing [3]. Physiological effects include stomatal closure, increased oxidative stress, and elevated proline accumulation [3,7]. Biochemical effects include reactive oxygen species (ROS) production, antioxidant defenses, oxidative damage, decreased chlorophyll levels, increased antioxidant enzymes, and carbohydrate accumulation [3,4]. While the effects of drought stress on plant development are generally negative, in some cases, it can offer potential advantages, especially for medicinal and aromatic plants [8]. Stress conditions can increase the production of secondary metabolites, particularly essential oils and phenolic compounds [1,8]. This can enhance the pharmacological or commercial value of these plants [9]. Therefore, controlled application of drought stress may provide benefits in agricultural production. Consequently, it may be possible to develop drought-resistant plant varieties and produce crops with more nutritionally or biochemically rich compounds.

Consumers’ awareness of the positive relationship between food and health is increasing demand for food products rich in functional components [10,11]. Adding these compounds to the human diet can significantly prevent many dangerous diseases such as inflammation and diabetes [10]. Garden cress (Lepidium sativum L.) is a plant belonging to the Brassicaceae family with nutritional and pharmaceutical values [12]. A fast-growing plant, garden cress is versatile, and various parts, including its leaves and roots, can be used for different purposes [13]. Raw leaves can be used to prepare salads and cooked with other vegetables, while dried leaves are used in the treatment of various diseases such as inflammation, bronchitis, rheumatism, and muscle pain, and as a diuretic [14]. Garden cress is considered a member of the “superfood” family due to its nutritional composition and therapeutic uses and is crucial for enhancing the nutritional value of formulated food products [13,14,15]. In addition to its nutritional properties, including proteins, carbohydrates, fiber, minerals, and vitamins, garden cress also contains various phytochemical and phenolic compounds such as saponins, flavonoids, alkaloids, tannins, and terpenes, which are responsible for its potential functional properties [12,14]. This wide nutritional and functional spectrum makes garden cress highly valuable to both the food industry and consumers [15].

To our knowledge, although some studies have applied water restriction to cress, these studies have only examined its effect on cress’s growth factors [12,16,17,18]. No research has been done to show how water restriction affects the nutritional and functional compounds of different cress cultivars. Therefore, this study aimed to investigate the effects of varying levels of water restriction (T 100%, T 85%, T 75%, and T 55%) on the nutritional and functional compounds of two cress cultivars, Dadaş and Bahar. Specifically, the study assessed how water restriction levels impacted the phenolic compound profile, antioxidant enzymes, anthocyanin levels, sugar content, vitamin levels, and amino acids in these cress cultivars.

2. Materials and Methods

This study was conducted in greenhouses at the Atatürk University Plant Production Application and Research Center and in the laboratory of the Department of Horticulture, Faculty of Agriculture (Erzurum, Turkey). Natural daylight was used in the greenhouses, and the temperature was set at approximately 25 °C with an 18/6 (day/night) cycle for the photoperiod.

2.1. Plant Material

Garden cress (Lepidium sativum L.) Bahar and Dadaş cultivars served as the plant material for the study. Cress seeds were sown in 26-L pots filled with a mixture of sand, soil, and peat in a 2:1:1 (v:v:y) ratio. The seeds were sown in three rows, each 1 cm deep, with 50 seeds per row. For each cress cultivar, the four irrigation treatments were applied to three pots (replicates), resulting in a total of 12 pots per cultivar. After sowing, the pots were randomly placed on benches in the greenhouse. Plant growth began three days after sowing.

2.2. Water Restriction Treatments

The irrigation amount and moisture content in the pots were determined using a portable moisture meter (HH2 Moisture Meter, WET Sensor, Delta-T Devices Ltd., 128, Low Road Burwell, Cambridge, UK) based on volume. For the irrigation treatments, the moisture meter was calibrated for the soil used in the experiment, and the volumetric moisture content at field capacity was measured. After identifying the field capacity of the pots, water restriction treatments were applied as follows: T 100% (control group), T 85% (mild drought stress), T 75% (moderate drought stress), and T 55% (severe drought stress). Irrigation was performed approximately every 2–3 days, with regular adjustments based on the average greenhouse temperature (minimum 15.2 °C, maximum 26.2 °C). Water restriction treatments started at sowing, and 14 treatments were conducted. Irrigation continued until the plants were harvested 36 days later.

2.3. Determination of Phenolic Compound Amounts of Garden Cress Cultivars

To analyze the phenolic compounds in garden cress samples, an RP-HPLC system (Shimadzu Scientific Instruments, Tokyo, Japan) was used, following the method specified by Sagdic et al. [19]. The system included an LC-10ADvp pump, a DAD detector, a CTO-10Avp column heater, an SCL-10Avp system controller, a DGU14A degasser, and a SIL-10ADvp auto-sampler. Separations were carried out at 30 °C on an Agilent Eclipse XDB C-18 reversed-phase column (Agilent Technologies, Santa Clara, CA, USA). A mobile phase of 2.0% acetic acid in distilled water (A) and methanol (B) was employed. Samples were introduced into the column at a flow rate of 0.8 mL/min. Twenty-five milligrams of dried garden cress extract was dissolved in 1 mL of methanol, and 10 µL of this solution was injected. Results are expressed as µg/L of per dry weight.

2.4. Determination of Antioxidant Enzymes of Garden Cress Cultivars

Garden cress samples were rinsed three times with 50 mM Tris-HCl + 0.1 M Na₂SO₄ (pH 8.0) and homogenized with liquid nitrogen. Then, samples were mixed with buffer (10 mM NaN₃ + 100 mM PVP + 0.1 M Na₂SO₄ (pH 8.0) + 50 mM Tris-HCl). Finally, the centrifugation was carried out at 15,000 rpm for 60 min at 4 °C. Enzyme extracts were added to initiate all enzymatic reactions, and enzymatic activity was measured using a spectrophotometer (Shimadzu 1208 UV, Kyoto, Japan) at 25 °C [20,21].

2.5. Determination of Anthocyanin Content of Garden Cress Cultivars

To determine the anthocyanin content of garden cress, 2 g of lyophilized ground powder was extracted with methanol containing 0.1% HCl. A mobile phase consisting of 0.3% phosphoric acid (A) and acetonitrile (B) was passed through an HPLC 1120 system (Agilent Technologies, Palo Alto, CA, USA) equipped with a C18 column at a flow rate of 1 mL/min. 10 µL of garden cress extract were injected, and anthocyanins were identified at 520 nm. The resulting peaks were compared to anthocyanin standard peaks [22]. For identification and quantification of cress cultivars’ anthocyanins, following anthocyanin standards were used: delphinidin-3-glucoside, cyanidin-3-glucoside, petunidin-3-glucoside, peonidin-3-glucoside, malvidin-3-glucoside, peonidin-3-glucoside acetyl, malvidin-3-glucoside acetyl, and malvidin-3-glucoside-p-coumaroyl.

2.6. Determination of Sugar Composition of Garden Cress Cultivars

Two grams of garden cress were extracted with 20 mL of 80% (v/v) ethanol solution overnight in a shaking water bath, followed by 150 min of sonication. 10 μL of the extract was passed through an HPLC (Agilent Technologies, 1100 series, Santa Clara, CA, USA) system equipped with an RID (1260 series) and a Purospher STAR NH₂ (250 × 4.6 mm, 5 μm, Merck, Darmstadt, Germany) column at a flow rate of 1 mL/min and an oven temperature of 40 °C. Sugar standards were supplied by Sigma-Aldrich (Shanghai, China), and the sugar contents of the cress were calculated based on calibration charts [23].

2.7. Determination of Vitamin Content of Garden Cress Cultivars

Extraction and HPLC procedures reported by Keskin et al. (2022) were used to determine the retinol (vitamin A), thiamine (B1), riboflavin (B2), pyridoxine (B6), and ascorbic acid (vitamin C) contents of garden cress cultivars [21]. 20 µL of the filtered extract was passed through an HPLC system equipped with a PDA detector (at 270 nm) and a C-18 column at 40 °C and a flow rate of 0.8 mL/min. 100 mM NaH₂PO₄ buffer (pH 2.2) containing 0.8 mM C₈H₁₇NaO₃S and C₂H₃N (9:1, v/v) was used as the mobile phase.

2.8. Amino Acid Composition of Garden Cress Cultivars

The HPLC system was equipped with a DAD detector, a Zorbax Eclipse AAA analytical column, and an autosampler for analyzing the amino acid composition of garden cress cultivars. FMOC and OPA were used for in-line derivatization before injection into the columns. FMOC- and OPA-derived amino acids were monitored at 262 nm and 338 nm, respectively. Sarcosine and norvaline served as internal standards for FMOC and OPA derivatives. Amino acid concentrations are reported in nmol/µL per dry weight [20].

2.9. Statistical Analysis

Results are given as the mean standard deviation of three replicates. Statistical analysis was performed using JMP 14.0, and differences between samples were determined using the Tukey test.

3. Results

3.1. Phenolic Compounds of Garden Cresses

Phenolic compounds are secondary metabolites produced by plants and are part of their defense mechanisms. Plants produce phenolic compounds as part of their antioxidant defenses to protect against various stresses or harmful effects. This study shows the phenolic content of Bahar and Dadaş cress cultivars subjected to different levels of water restriction in

Table 1. Phenolic compound amounts of garden cress cultivars subjected to different levels of water restriction by HPLC (µg/L per dry weight).

GCT	GA	VA	TCA	T-p-CA	FA	CA	C	EC	Q	R	M	T
B0	3.73 ± 0.20 f	4.94 ± 0.11 f	4.14 ± 0.20 e	4.88 ± 0.17 e,f	8.42 ± 0.19 c	5.40 ± 0.45 d	5.11 ± 0.03 e	2.82 ± 0.19 c	9.32 ± 0.25 b,c	3.56 ± 0.16 e	1.14 ± 0.02 d	5.02 ± 0.21 e
B1	6.35 ± 0.07 c	7.44 ± 0.38 c	6.15 ± 0.08 c	6.27 ± 0.18 b,c	12.67 ± 0.44 a	8.65 ± 0.28 b	9.24 ± 0.18 b	3.93 ± 0.09 b	12.84 ± 0.26 a	5.42 ± 0.30 b	2.51 ± 0.25 c	8.64 ± 0.51 c
B2	6.16 ± 0.20 c	6.71 ± 0.21 d	5.90 ± 0.23 c	5.96 ± 0.45 c d	10.55 ± 0.78 b	8.34 ± 0.26 b	8.14 ± 0.32 b,c	3.64 ± 0.40 b	10.26 ± 0.44 b	5.29 ± 0.13 b	2.93 ± 0.31 b,c	8.56 ± 0.27 c
B3	4.10 ± 0.13 f	5.21 ± 0.19 f	4.23 ± 0.10 e	4.76 ± 0.16 f	7.87 ± 0.51 c	5.71 ± 0.29 d	5.47 ± 0.16 d,e	2.92 ± 0.09 c	8.36 ± 0.56 c	3.92 ± 0.24 d,e	1.49 ± 0.04 d	5.43 ± 0.23 d,e
D0	4.68 ± 0.26 e	5.62 ± 0.08 e,f	4.82 ± 0.24 d	5.25 ± 0.26 e f	8.08 ± 0.18 c	6.33 ± 0.10 c d	6.83 ± 0.28 c d	3.48 ± 0.24 b,c	8.33 ± 0.23 c	4.40 ± 0.19 c d	1.74 ± 0.03 d	6.16 ± 0.59 d
D1	7.82 ± 0.12 a	9.58 ± 0.18 a	8.07 ± 0.19 a	8.24 ± 0.24 a	12.04 ± 0.42 a	10.58 ± 0.34 a	11.70 ± 0.87 a	5.11 ± 0.37 a	13.50 ± 0.29 a	6.44 ± 0.09 a	3.71 ± 0.27 a	11.56 ± 0.34 a
D2	7.30 ± 0.05 b	8.14 ± 0.38 b	7.41 ± 0.26 b	6.90 ± 0.15 b	9.87 ± 0.72 b	9.84 ± 0.64 a	11.27 ± 1.10 a	4.99 ± 0.13 a	9.83 ± 0.50 b	6.29 ± 0.28 a	3.49 ± 0.38 a,b	10.33 ± 0.19 b
D3	5.23 ± 0.15 d	6.01 ± 0.21 e	4.94 ± 0.12 d	5.49 ± 0.19 d,e	7.29 ± 0.47 c	6.81 ± 0.35 c	6.34 ± 0.19 d,e	3.63 ± 0.12 b	7.27 ± 0.17 d	4.64 ± 0.29 c	1.78 ± 0.05 d	6.26 ± 0.27 d

GCT: Garden cress type, GA: Gallic acid, VA: Vanillic acid, TCA: Trans-caffeic acid, T-p-CA: Trans-p-coumaric acid, FA: Ferulic acid, CA: Caftaric acid, C: Catechin, EC: Epicatechin, Q: Quercetin, R: Rutin, M: Myricetin, T: Tyrosol. Levels not connected by same letter are significantly different (p ≤ 0.05). B: Bahar garden cress cultivar, D: Dadaş garden cress cultivar, B0–D0: T 100% (without drought treatment, control group), B1–D1: T 85% (mild drought stress), B2–D2: T 75% (moderate drought stress), and B3–D3: T 55% (severe drought stress).

. Additionally, the HPLC chromatograms of phenolic compounds are included in the Supplementary file (Figure S1). Among the cultivars not subjected to water restriction (B0 and D0), Dadaş cress had higher levels of phenolic compounds, except for quercetin (p ≤ 0.05). The three phenolic compounds detected at the highest levels for Bahar cress were quercetin (9.32 ± 0.25, µg/L), ferulic acid (8.42 ± 0.19, µg/L) and caftaric acid (5.40 ± 0.45, µg/L), while for Dadaş cress they were quercetin (8.33 ± 0.23, µg/L), ferulic acid (8.08 ± 0.18, µg/L), and catechin (6.83 ± 0.28, µg/L). Conversely, applying water restriction to the cress cultivars significantly increased the levels of phenolic compounds (p ≤ 0.05). The three phenolic compounds detected at the highest levels in Bahar and Dadaş cress with 85% humidity level under water restriction were quercetin, caftaric acid and catechin. Notably, there were significant increases in phenolic compounds, especially with mild and moderate water restriction (85% and 75%). Although phenolic levels in cress exposed to severe water restriction (55%) were slightly higher than in unwater-restricted plants, they decreased compared to mild and moderate stress. The results indicate that water restriction significantly influences phenolic production in cress, with mild and moderate stress promoting synthesis, while severe drought reduces these compounds. An additional key finding was the variation in phenolic profiles among cress cultivars, with Dadaş cress exhibiting particularly high phenolic content.

3.2. Antioxidant Enzyme Levels of Garden Cresses

Upon exposure to oxidative stress, plants induce the activity of antioxidant enzymes such as peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) to counteract detrimental effects, including damage to cellular membranes, organelles such as mitochondria and chloroplasts, and the subsequent degradation of cellular structures and components [24]. In this study, the levels of eight antioxidant enzymes in the Bahar and Dadaş cress cultivars subjected to different degrees of drought stress—glutathione reductase (GR), glutathione S-transferase (GST), glucose-6-phosphate dehydrogenase (G6PD), 6-phosphogluconate dehydrogenase (6GPD), CAT, POD, superoxide dismutase (SOD), and APX—are presented in Figure 1. The highest antioxidant enzyme levels were GST, 6GPD, and G6PD in both cultivars. In Bahar cress, GST was 231.28 ± 19.93 EU/g, 6GPD was 221.98 ± 5.63 EU/g, and G6PD was 85.12 ± 3.77 EU/g. In Dadaş cress, these values were 213.17 ± 18.38, 186.23 ± 6.06, and 73.48 ± 3.26 EU/g, respectively. The antioxidant enzyme levels of the Bahar cress cultivar were higher than those of the Dadaş cress cultivar (p ≤ 0.05). A significant increase in antioxidant enzyme levels was observed in both cress cultivars under drought treatment. In particular, antioxidant enzyme levels increased significantly as a result of mild and moderate drought treatments (p ≤ 0.05). Although the highest antioxidant enzyme levels were recorded after mild drought treatment, the highest amounts of SOD and APX enzymes were detected after severe drought treatment. However, a limited increase in the levels of GR, G6PD, 6GPD, and CAT enzymes, and a limited decrease in the levels of GST enzymes, were observed following severe drought treatment. Conversely, a significant increase in POD and APX enzyme levels was detected in both the Bahar and Dadaş cress cultivars after severe drought treatment (p ≤ 0.05). Drought stress restricts the uptake of carbon dioxide (CO₂), which is essential for photosynthesis in plants. This limitation causes the excess energy from photosynthesis to be converted into harmful molecules known as ROS. Plants possess natural defense mechanisms, such as enzymatic antioxidants, to counteract these harmful effects, thereby increasing enzyme levels to mitigate ROS-induced damage [25,26].

3.3. Anthocyanin Content of Garden Cresses

In this study, the effects of drought stress on the levels of delphinidin-3-glucoside, cyanidin-3-glucoside, petunidin-3-glucoside, peonidin-3-glucoside, malvidin-3-glucoside, peonidin-3-glucoside acetyl, malvidin-3-glucoside acetyl, and malvidin-3-glucoside-p- coumaroyl anthocyanins in cress cultivars were analyzed using HPLC (

Table 2. Anthocyanin contents of garden cress cultivars subjected to water restriction by HPLC (g/100 g per dry weight).

Garden Cress Types	D3G	C3G	P3G	PEO3G	M3G	PEO3GA	M3GA	M3GpC
B0	4.11 ± 0.44 d	3.64 ± 0.05 d	7.27 ± 0.17 d	91.50 ± 2.14 c,d	51.12 ± 0.73 c	54.56 ± 2.19 e	19.17 ± 0.88 e	2.57 ± 0.13 d
B1	6.69 ± 0.35 b	5.35 ± 0.18 b	10.09 ± 1.04 b	125.56 ± 9.96 a	96.61 ± 5.22 b	91.36 ± 3.14 c	19.48 ± 2.02 d	3.20 ± 0.18 c,d
B2	6.99 ± 0.07 b	4.77 ± 0.18 c	10.04 ± 0.43 b	98.82 ± 1.68 b,c	96.04 ± 8.19 b	86.79 ± 4.77 c	29.83 ± 1.87 c,d	4.03 ± 0.26 b
B3	4.34 ± 0.26 d	3.86 ± 0.19 d	7.79 ± 0.23 c,d	85.46 ± 3.18 c,d	54.75 ± 2.63 c	58.73 ± 4.90 d,e	29.63 ± 1.32 d	3.39 ± 0.23 b,c
D0	4.89 ± 0.30 c,d	4.50 ± 0.07 c	8.57 ± 0.22 b,c,d	87.92 ± 1.82 c,d	64.52 ± 0.93 c	63.26 ± 2.82 d,e	30.97 ± 2.55 c,d	3.06 ± 0.24 c,d
D1	7.84 ± 0.24 a	6.48 ± 0.29 a	13.56 ± 0.52 a	119.48 ± 9.38 a,b	116.61 ± 10.81 a	119.92 ± 4.12 a	40.55 ± 1.17 a	5.06 ± 0.28 a
D2	7.28 ± 0.19 a,b	5.73 ± 0.13 b	12.81 ± 1.10 a	102.17 ± 10.89 b,c	116.59 ± 10.08 a	103.68 ± 5.70 b	38.00 ± 0.70 a,b	4.03 ± 0.27 b
D3	5.19 ± 0.30 c	4.60 ± 0.23 c	9.12 ± 0.27 b,c	79.39 ± 3.19 d	66.69 ± 3.21 c	67.62 ± 5.64 d	34.16 ± 1.34 b,c	3.02 ± 0.20 c,d

D3G: Delphinidin-3-glucoside, C3G: Cyanidin-3-glucoside, P3G: Petunidin-3-glucoside, PEO3G: Peonidin-3-glucoside, M3G: Malvidin-3-glucoside, PEO3GA: Peonidin-3-glucoside acetyl, M3GA: Malvidin-3-glucoside acetyl, M3GpC: Malvidin-3-glucoside-p-coumaroyl. Levels not connected by same letter are significantly different (p ≤ 0.05). B: Bahar garden cress cultivar, D: Dadaş garden cress cultivar, B0–D0: T 100% (without drought treatment, control group), B1–D1: T 85% (mild drought stress), B2–D2: T 75% (moderate drought stress), and B3–D3: T 55% (severe drought stress).

and Figure S2). The most abundant anthocyanins in Dadaş and Bahar cress were peonidin-3-glucoside (87.92 ± 1.82–91.50 ± 2.14 g/100 g). Following this, malvidin-3-glucoside (64.52 ± 0.93–51.12 ± 0.73 g/100 g) and peonidin-3-glucoside acetyl (63.26 ± 2.82–54.56 ± 2.19 g/100 g) were predominant. Aside from peonidin-3-glucoside, Dadaş cress had higher concentrations of anthocyanins than Bahar cress (p ≤ 0.05). An increase in anthocyanin levels was observed in cress cultivars due to drought stress. Notably, a significant rise in anthocyanin levels occurred under mild and moderate drought conditions (p ≤ 0.05). For example, in Dadaş cress subjected to mild drought, peonidin-3-glucoside increased from 87.92 ± 1.82 to 119.48 ± 9.38, malvidin-3-glucoside from 64.52 ± 0.93 to 116.61 ± 10.81, and peonidin-3-glucoside acetyl from 63.26 ± 2.82 to 119.92 ± 4.12, g/100 g. Similar results were seen in Bahar cress following mild drought treatment. A significant increase was especially observed in peonidin-3-glucoside acetyl levels, which nearly doubled with mild drought application to the cress cultivars. Under severe drought, total anthocyanin levels decreased compared to mild and moderate drought conditions. However, malvidin-3-glucoside acetyl levels remained significantly higher than in the control (p ≤ 0.05).

3.4. Sugar Composition of Garden Cresses

The amounts of sucrose, glucose, fructose, rhamnose, galactose, xylose, and arabinose in Bahar and Dadaş cress cultivars were measured using HPLC (Figure 2 and Figure S3). The highest sugar levels found in Dadaş and Bahar cress were fructose, glucose, and sucrose. For the Dadaş cultivar, the highest levels were 15.85 ± 0.76 g/100 g for fructose, 13.32 ± 0.62 g/100 g for glucose, and 3.59 ± 0.17 g/100 g for sucrose. For the Bahar cress cultivar, these amounts changed to 10.18 ± 0.30 g/100 g, 8.19 ± 0.15 g/100 g, and 2.50 ± 0.26 g/100 g, respectively. Rhamnose, galactose, xylose, and arabinose appeared in much lower amounts in both cress types. Dadaş cress contained higher sugar levels than Bahar cress (p ≤ 0.05). A significant increase in sugar content was observed in both cress cultivars after drought treatment (p ≤ 0.05). Especially, mild and moderate drought conditions caused notable increases in sugar levels. The highest sugar level was observed in Dadaş cress under mild drought, with 20.40 ± 0.56 g/100 g for fructose. Similarly, fructose also had the highest level in Bahar cress under drought conditions. Sugar levels in cress cultivars exposed to severe drought were slightly higher than in the control groups (drought-free) (p > 0.05). However, sugar levels in cultivars exposed to mild and moderate drought were significantly higher than in the control groups (p ≤ 0.05).

3.5. Vitamin Content of Garden Cresses

Both Dadaş and Bahar cress were rich in Riboflavin, containing 29.47 ± 1.30 and 14.03 ± 1.62 mg/kg, respectively (Figure 3 and Figure S4). Pyridoxine, thiamine, retinol, and ascorbic acid were the most abundant in both cress cultivars, respectively. A significant increase in the vitamin content of the cress cultivars was observed with drought treatment. The highest vitamin levels in Dadaş cress were seen after mild drought, while the highest levels in Bahar cress appeared after moderate drought. This indicates that different drought levels can significantly affect vitamin content in various cress cultivars. Conversely, although the vitamin contents of cress exposed to severe drought were higher than those of non-drought cress, they were still lower than levels under mild and moderate drought conditions.

3.6. Amino Acid Composition of Garden Cresses

In this study, changes in the amounts of essential amino acids (phenylalanine, valine, tryptophan, isoleucine, methionine, histidine, leucine, and lysine) and asparagine and tyrosine in cress cultivars under drought conditions were analyzed using HPLC (

Table 3. Amino acid composition of garden cress cultivars subjected to water restriction by HPLC (nmol/µL per dry weight).

GCT	Phenylalanine	Valine	Tryptophan	Isoleucine	Methionine	Histidine	Leucine	Lysine	Asparagine	Tyrosine	Proline
B0	54.09 ± 2.25 f	31.89 ± 1.32 e	46.32 ± 1.93 d	46.65 ± 1.45 d	40.96 ± 1.70 c	41.24 ± 1.71 e	47.38 ± 1.97 e	32.55 ± 1.36 e	32.67 ± 1.36 d	32.55 ± 1.35 f	35.70 ± 1.48 f
B1	131.01 ± 4.93 b	77.25 ± 2.91 b	113.18 ± 2.54 b	112.19 ± 4.22 b	102.52 ± 2.19 a	99.89 ± 3.76 a,b	114.74 ± 4.32 b,c	79.83 ± 1.30 b	79.11 ± 2.98 b	78.83 ± 2.97 a,b	86.48 ± 3.25 b,c
B2	102.76 ± 2.57 e	59.99 ± 1.98 d	89.95 ± 0.77 c	88.95 ± 2.50 c	79.41 ± 3.11 b	79.93 ± 3.07 d	90.82 ± 2.44 d	61.15 ± 2.10 d	71.36 ± 2.12 c	61.15 ± 2.10 e	66.76 ± 2.72 e
B3	140.54 ± 7.14 a,b	80.90 ± 2.62 a,b	114.15 ± 1.80 b	124.16 ± 9.98 a,b	107.21 ± 9.13 a	101.28 ± 6.89 a,b	126.83 ± 10.16 a,b	79.22 ± 4.39 b	90.19 ± 3.42 a	75.89 ± 3.28 b,c	87.23 ± 5.29 b
D0	115.27 ± 3.73 c,d	65.40 ± 2.28 c,d	96.47 ± 0.84 c	96.80 ± 0.50 c	85.36 ± 0.66 b	83.59 ± 4.63 c,d	98.86 ± 0.62 d	65.33 ± 2.08 c,d	66.23 ± 1.99 c	66.67 ± 2.44 d,e	72.81 ± 3.24 d,e
D1	117.20 ± 0.99 c	68.50 ± 1.22 c	96.64 ± 3.23 c	99.64 ± 1.54 c	90.34 ± 2.84 b	90.94 ± 2.79 b,c	103.47 ± 1.61 c,d	69.83 ± 1.36 c	70.08 ± 1.38 c	69.83 ± 1.35 c,d	76.96 ± 0.98 c,d
D2	104.83 ± 3.76 d,e	61.81 ± 2.21 c,d	97.76 ± 0.38 c	96.43 ± 2.58 c	86.03 ± 2.95 b	79.93 ± 2.86 d	91.81 ± 3.29 d	63.08 ± 2.26 c,d	67.30 ± 1.22 c	63.07 ± 2.25 e	69.20 ± 2.47 d,e
D3	144.61 ± 2.53 a	86.46 ± 3.51 a	125.91 ± 3.93 a	125.24 ± 4.56 a	109.58 ± 2.08 a	110.42 ± 2.20 a	128.32 ± 5.05 a	88.37 ± 3.82 a	85.38 ± 1.99 a	85.03 ± 2.05 a	97.59 ± 5.30 a

GCT: Garden cress type. Levels not connected by same letter are significantly different (p ≤ 0.05). B: Bahar garden cress cultivar, D: Dadaş garden cress cultivar, B0–D0: T 100% (without drought treatment, control group), B1–D1: T 85% (mild drought stress), B2–D2: T 75% (moderate drought stress), and B3–D3: T 55% (severe drought stress).

and Figure S5). Dadaş cress was quite rich in amino acids, and non-drought Dadaş cress contained twice the amount of all amino acids compared to Bahar cress (p ≤ 0.05). The highest amount of phenylalanine was 115.27 ± 3.73 nmol/µL in Dadaş cress and 54.09 ± 2.25 nmol/µL in Bahar cress. The lowest amino acid levels were lysine (65.33 ± 2.08 nmol/µL) in Dadaş cress and valine (31.89 ± 1.32 nmol/µL) in Bahar cress. Significant changes in amino acid composition were observed in cress cultivars following drought stress. Unlike the previously mentioned phenolic compounds, sugars, vitamins, anthocyanins, and antioxidant enzymes, the greatest increase in amino acid levels occurred after severe drought. While amino acid concentrations increased following mild and moderate drought, the most substantial rise was seen after severe drought. Notably, there was more than a 2.5-fold increase in amino acids in Bahar cress. For example, asparagine increased about 2.76-fold, from 32.67 ± 1.36 nmol/µL to 90.19 ± 3.42 nmol/µL, and leucine rose approximately 2.67-fold, from 47.38 ± 1.97 nmol/µL to 126.83 ± 10.16 nmol/µL. The increase in amino acid levels in Dadaş cress was not as high as in Bahar cress, indicating that cress cultivars respond differently to various drought levels.

4. Discussion

In plants exposed to drought stress, the production of ROS increases. As a response, plants boost the production of antioxidant molecules such as phenolic compounds, leading to higher concentrations of these substances [27]. Additionally, Selmar and Kleinwächter [28] described the biochemical changes in phenolic compounds under drought stress. When stomata close due to high light intensity, CO₂ uptake is significantly reduced. As a result, the consumption of reducing equivalents (NADPH+H⁺) for CO₂ fixation via the Calvin cycle drops significantly, creating an excess of NADPH+H⁺. This surplus directs metabolic processes toward the synthesis of highly reduced compounds like isoprenoids, phenols, or alkaloids. Espadas et al. [27] reported that drought stress increases both the antioxidant capacity and the diversity of phenolic compounds in Carica papaya. Mohagheghian et al. [24] observed that the levels of syringic acid, gallic acid, ellagic acid, and vanillic acid in barley (Hordeum vulgare L.) genotypes doubled due to drought stress. Sarker and Oba [29] applied different levels of drought stress—mild stress (90% FC), moderate stress (60% FC), and severe stress (30% FC)—to Amaranthus tricolor. Their findings showed that moderate and severe drought stress significantly boosted the concentrations of sixteen phenolic compounds. Conversely, prolonged drought stress was reported to cause a notable decrease in total phenolic content and phenolic acid levels in the leaves and roots of grapevine seedlings [30]. Historically, drought stress was seen as a negative factor causing serious yield losses in agriculture [31]. However, recent research suggests that applying moderate drought stress can improve product quality in spices and medicinal plants and promote the synthesis of health-promoting compounds [31]. This approach has been studied across many plant species. The results of this study indicate that drought stress increases the phenolic content in cress cultivars, highlighting the importance of applying controlled moderate drought stress for producing cress harvests rich in phenolic compounds.

Plants tolerant to water stress increase their antioxidant capacity by reprogramming their metabolism, while sensitive plants activate their antioxidant systems. Antioxidant activity is not only important during severe drought stress but also influences recovery from water deprivation and dehydration [32]. Another study determined that APX, POD, and CAT activities in samples of different Eruca sativa genotypes under drought stress increased significantly. Under drought stress, the highest APX activity values were found in G54, the lowest in G23 and G11. CAT values ranged from 0.17 units mg/protein (G32) to 0.22 units mg/protein (G37). POD activity values ranged from 0.19 (G7) to 0.30 (G56) µmol min/mg/protein [33]. Hou et al. [34] determined that under mild drought stress, CAT, POD, and GPX activities increased in Carex duriuscula leaves, while CAT and POD activities increased in roots. Under severe drought, CAT, POD, SOD, and GPX activities increased in both leaves and roots. DaCosta and Huang [35] examined differences in antioxidant enzyme levels in response to drought in colonial twistgrass (Agrostis capillaris L.), creeping twistgrass (A. stolonifera L.), and velvet twistgrass (A. canina L.), the species with the highest drought tolerance. They determined that prolonged drought stress results in decreased antioxidant enzyme activity and causes oxidative damage, while velvet twistgrass maintains its antioxidant enzyme activity longer under drought stress than the other two species. In this study, it was observed that the Bahar cultivar exhibited higher antioxidant capacity than the Dadaş cultivar under stress conditions in terms of antioxidant content. It has been determined that the antioxidant properties, and therefore the quality, of the aromatic plant species mint, basil, marjoram, and thyme can be significantly enhanced by deliberately applying drought stress. However, this increase is accompanied by a decrease in plant growth, and therefore the advantages and disadvantages of deliberately applying drought stress to each aromatic plant species should be carefully evaluated [36].

Studies indicate that drought stress significantly impacts anthocyanin content and related biosynthetic pathways in plants. Sperdouli and Moustakas [37] reported a notable increase in anthocyanin levels in Arabidopsis thaliana when subjected to moderate drought stress compared to control plants. Similarly, Chen et al. [38] studied a purple-leafed Brassica napus (rapeseed) genotype and found that drought stress promotes anthocyanin accumulation, which enhances the plant’s stress tolerance. This research showed that anthocyanin buildup, especially cyanidin glucosides, improves drought resistance by strengthening the plant’s water-holding capacity and antioxidant defenses. Ünal and Okatan [39] also observed that mild drought stress applied to strawberry (Fragaria × ananassa) plants (cv. Festival, Benicia, and Monterey) increased total anthocyanin levels. Medina-Lozano et al. [40] reported that drought stress raised the anthocyanin content across various lettuce species (Lactuca sativa). Hinojosa- Gómez et al. [41] found that applying drought stress to different Hibiscus cultivars increased their anthocyanin levels, with significant rises in cyanidin, delphinidin 3-O-glucoside, cyanidin 3-O-glucoside, and cyanidin 3-O-sambubioside, particularly at 65% humidity irrigation. Conversely, a decrease in anthocyanin content was observed in all Hibiscus cultivars under water stress at 33% humidity.

Anthocyanins offer several benefits to plants. They improve visual appeal, boost resistance to abiotic stresses, and help regulate osmotic pressure. They also aid in photosynthesis, reinforce antioxidant defenses, neutralize ROS, and influence antioxidant enzymes such as CAT and SOD [38]. The increase in anthocyanin levels in plants under drought is controlled by various genetic, hormonal, and metabolic processes [38,42]. For instance, to reduce oxidative stress caused by ROS, plants boost anthocyanin accumulation during drought stress [38]. Abscisic acid (ABA) is a crucial plant hormone involved in anthocyanin biosynthesis. When plants experience drought stress, ABA levels rise, which activates genes responsible for producing anthocyanins and results in higher anthocyanin levels [42]. Alternatively, drought stress triggers transcription factors (TFs) such as MYB, bHLH, and WD40, which are key proteins that activate structural genes involved in anthocyanin biosynthesis, leading to increased anthocyanin accumulation [42]. Therefore, the increase in anthocyanin levels in plants under drought stress results from a complex and interconnected set of mechanisms, including ROS scavenging, activation of genetic and hormonal signaling pathways, photoprotection, and osmotic regulation.

Drought stress is one of the most significant abiotic stresses that limits plant growth and yield, causing notable changes in plant carbon metabolism [43]. The reviewed studies show that the effect of drought stress on sugar content in plants is not one-sided; it can lead to either sugar accumulation or a decrease, depending on factors such as plant species, the organ examined, the developmental stage, and the stress severity. This results from the complex and multifaceted adaptation mechanisms plants develop to cope with drought. One of the common plant responses to drought is the accumulation of soluble sugars to maintain osmotic pressure. Du et al. [43] reported significant increases in soluble sugar and sucrose levels in the leaves and roots of soybean seedlings under drought stress. Similarly, Dong et al. [44] also indicated that drought triggers the buildup of sucrose, fructose, and glucose in soybeans during early stages. These findings suggest that soluble sugars serve as osmoprotectants, helping to maintain cell turgor and reduce osmotic stress. Moreover, a study on apple leaves by Yang et al. [45] found that sugars like sorbitol, glucose, and fructose accumulated under drought stress, while sucrose, a primary product of photosynthesis, decreased significantly. This reduction is linked to the conversion of sucrose into hexoses. However, in some cases, drought stress results in a decline in sugar content. It has been observed that total soluble sugar levels decrease with drought, especially in rice leaves and sheaths [46], Lanzhou lily [47], and during the pod-filling period of soybeans, starch, fructose, and glucose levels also decline [48]. The main reason for this decrease is that drought stress limits CO₂ intake by closing stomata, leading to reduced photosynthesis [44]. As a result, sugar metabolism under drought stress involves not just accumulation but a dynamic process of synthesis, degradation, and transport. Although the changes in sugar levels (either increase or decrease) depend on the plant’s genetic traits and the severity and duration of stress, the core mechanism revolves around maintaining osmotic balance and optimizing energy use [43,44,46].

Literature data show that drought stress can affect the vitamin content of plants in different ways, either increasing or decreasing it. For example, in a study on lettuce (Lactuca sativa) and its wild relatives, Medina-Lozano et al. [40] reported that drought stress lowered the total ascorbic acid content. Similarly, Shin et al. [49] observed a decrease in ascorbic acid in lettuce seedlings under drought conditions. These results suggest that the plant either rapidly uses up existing ascorbic acid stores to fight drought-induced oxidative stress or that drought suppresses how ascorbic acid is made [50]. This aligns with how ascorbic acid works as an antioxidant, meaning it gets oxidized while neutralizing ROS, which reduces its overall amount. On the other hand, Aziz et al. [50] found that ascorbic acid levels increased in quinoa (Chenopodium quinoa) when drought stress was applied. Šola et al. [51] reported that Chinese cabbage (Brassica rapa ssp. pekinensis) grown under drought conditions had more ascorbic acid than plants in normal conditions. Increased levels of ascorbic acid and beta-carotene have also been found in drought-stressed Amaranthus leafy vegetables [29]. This indicates that ascorbic acid isn’t just a passive protector but also actively produced and accumulated in response to drought as part of a defense mechanism [50]. This helps the plant preserve cellular integrity by boosting its antioxidant defenses during prolonged stress. Therefore, some plants quickly exhaust their vitamin resources as an immediate defense, while others boost vitamin production for longer-term adaptation. Understanding these mechanisms provides important insights for developing drought-tolerant and nutrient-rich plant varieties.

In plants, drought stress triggers a complex series of physiological and biochemical responses aimed at maintaining osmotic balance and minimizing cellular damage. Central to these responses are changes in amino acid metabolism. Studies show that drought stress generally tends to increase the amount of free amino acids in plants. This increase serves both as part of osmotic regulation and as a defense mechanism against stress-induced cellular damage. Mild drought stress conditions in strawberry (Fragaria × ananassa) have also been shown to increase protein levels and the content of many amino acids, including L-alanine, L-aspartic acid, L-glutamic acid, phenylalanine, lysine, histidine, valine, leucine, threonine, serine, proline, and arginine [39]. Another study by Hu et al. [52] reported that total amino acid content and the quantities of most individual amino acids increased in leaves of Zanthoxylum bungeanum under drought stress. The mechanism behind this rise in amino acids is quite complex. Proline is one of the amino acids most abundantly accumulated in response to drought stress and serves multiple roles, including osmotic protection, scavenging of ROS, and maintaining protein structures [52,53]. In their study on licorice (Glycyrrhiza glabra L.), Haghighi et al. [54] reported increased proline accumulation under drought stress, linking it to plant stress tolerance. Similarly, Qu et al. [55] discovered that the glutamic acid-mediated proline biosynthesis pathway in tea (Camellia oleifera) was activated during drought stress, resulting in higher proline levels. Arginine can also act as a precursor for proline synthesis, and regulation of this pathway plays a vital role in plant stress responses [54,56]. Aromatic amino acids such as phenylalanine, tyrosine, and tryptophan serve as starting molecules in the biosynthesis of phenolic compounds and phytoalexins, which are secondary metabolites in plants [52]. These secondary metabolites play important roles in plant defense by functioning as antioxidants and signaling molecules. A study on soybean (Glycine max) roots by Wang et al. [56] revealed that the phenylalanine, tyrosine, and tryptophan metabolic pathways are particularly important in drought resistance. Under drought stress, the accumulation of these amino acids enhances the plant’s defense mechanisms, increasing its chances of survival. As a result, drought stress causes plants to accumulate amino acids. This build-up helps with water regulation through amino acids like proline and arginine, while others, such as aromatic amino acids, promote the production of secondary metabolites and boost cellular defense systems. This metabolic shift is a key way for plants to adapt to tough environmental conditions like water shortage.

This study clearly demonstrated that the Bahar and Dadaş cress cultivars respond differently to drought stress. For example, the Bahar cultivar exhibited higher antioxidant enzyme activity, while the Dadaş cultivar generally showed increased levels of phenolic compounds, anthocyanins, and sugars. These differences suggest that the two cultivars adopt different strategies to cope with stress. These strategies may be due to genetic variations between the varieties or physiological differences such as root system efficiency, stomatal behavior, or metabolic regulation. The Bahar cultivar primarily activates enzymatic defense mechanisms (e.g., GST, 6GPD) against stress, whereas the Dadaş cultivar focuses on osmotic regulation and ROS scavenging by increasing the production of non-enzymatic antioxidants (like phenolics and anthocyanins). This indicates that the plants prioritize different defense pathways, allocating their energy accordingly. Drought tolerance is a complex trait governed by many genes. Therefore, the genetic differences between the two cultivars probably explain these observed variations.

5. Conclusions

This study thoroughly examined the effects of various water restriction levels (100%, 85%, 75%, and 55%) on the nutritional and bioactive compounds in Bahar and Dadaş cress cultivars. The results clearly showed that cress plants develop adaptive mechanisms to drought stress, which can enhance their nutritional value. Specifically, it was found that mild and moderate water restrictions (85% and 75% moisture levels) significantly increased phenolic compounds, antioxidant enzymes, anthocyanins, and sugar contents. These increases indicate that the plants strengthen their defense systems against oxidative stress and boost secondary metabolite production. The study also revealed that severe drought stress (55% moisture level) may reduce some compound levels; however, they still remained higher than the control group. Interestingly, amino acid levels showed the most notable increase under severe drought conditions, suggesting that the plant accumulates amino acids for survival and protein synthesis during harsh stress. In conclusion, these findings imply that water restriction techniques can be incorporated into sustainable farming practices to produce cress with higher nutritional value and increased functional compounds. On the other hand, implementing controlled mild to moderate drought conditions in real-world settings can be challenging due to varying environmental factors such as rainfall differences and soil variety. Although regular water restrictions can improve nutrient quality, severe drought may reduce yield. Therefore, future field-based studies are essential to verify these benefits and investigate how controlled drought stress can be incorporated into commercial growing systems.