Effect of Compost from Cardoon Biomass, as Partial Replacement of Peat, on the Production and Quality of Baby Leaf Lettuce

Abstract

The use of peat, the standard substrate used for soilless cultivation of horticultural crops, is becoming of increasing concern as peat is a non-renewable resource and its extraction can degrade wetland ecosystems, creating a strong environmental impact. For this reason, the search for organic materials that can totally or partially replace peat has become increasingly important. In this research, three types of composts (C1, C2, C3), derived from cardoon biomass mixed in different volumes with woody and/or fruit wastes, were utilized as the constituents of growing media, at two dilution rates with peat (60:40 and 30:70 v:v), to assess their effect on the growth and quality of baby leaf lettuce in a greenhouse trial. The two cultivars Imperiale and Verde d’Inverno, belonging to the butterhead and romaine lettuce types, respectively, were employed. Plant performance and yield were unaffected or were positively affected by compost-containing growing medium compared to the control. The cultivars responded differently to the growing medium; the Imperiale showed the highest yield with C1 compost at a 60% rate while the Verde d’Inverno with the C2 was at 30%. The total chlorophyll, carotenoids, and ascorbic acid were found higher in the Verde d’Inverno than in the Imperiale variety while the total polyphenols, flavonoids, and antioxidant activity were lower. Also, the content of chlorophylls as well as of antioxidant compounds and antioxidant activity were differently affected by the growing medium, depending on the lettuce cultivar. The results obtained indicate that cultivated cardoon waste-based compost is a promising constituent of the growing media for baby leaf production. The specific varietal response observed should be considered to optimize both yield and product quality.

1. Introduction

Peat is the most common growing medium constituent for soilless cultivation of horticultural crops due to its beneficial physical properties and chemical characteristics for commercial plant production [1]. Peat, a non-renewable resource, is formed by the partial decomposition of plants typical of a water-saturated environment with low nutrients and pH, under low temperatures and anaerobic conditions, so it consists of dead biomass with high carbon concentration forming peatland soil [1,2,3]. Peat soils are unique ecosystems that account for about 10% of the available freshwater resources [4,5,6] and are the most important terrestrial carbon reservoirs on Earth [7]. Peat extraction results in the release of high CO₂ emissions due to aeration and mineralization of the peat, which contributes to the greenhouse gas effect [2,8].

The extensive use of peat for horticulture and the subsequent depletion of peatlands has led to growing environmental concerns about peat extraction [9]. For this reason, the search for sustainable substitutes that can totally or partially replace peat, a non-renewable resource, has become increasingly important.

Various materials, such as municipal solid waste, sawdust, biomass waste, digestate, and biochar after proper transformative processes (such as composting, pyrolysis, etc.), were favorably valued when mixed with peat in horticultural media [10,11,12,13,14,15]. Among them, compost is successfully used as a replacement for peat to produce seedlings of tomato (Solanum lycopersicum L.) in nurseries [16] and plants of tomato and basil (Ocimum basilicum L.) in pots [17]. It has been indicated that compost and peat mixtures are a useful growing medium for the greenhouse production of zucchinis (Cucurbita pepo L.), peppers (Capsicum annuum L.), and tomatoes [18].

The use of good growing media is very important for achieving positive results [19]. Although there are various types of compost that have been evaluated to avoid the consumption of peat, to our knowledge there is no information regarding the use of compost derived from cultivated cardoon (Cynara cardunculus var. altilis DC.) crop residues.

Cultivated cardoon is a perennial herbaceous plant with an annual development cycle [20,21]. Cardoon provides high biomass yields with low nitrogen inputs and drought tolerance, protects the soil from erosion, and its cultivation can lead to the improvement of soil fertility [22].

Nowadays, cardoon has gained interest as a novel crop for multipurpose use that includes energy and biofuels, cellulosic pulps, phytochemicals as well as food and feed [20,23,24]. It has recently been reported that cardoon crop residues can also be an optimal growing medium for edible Pleurotus eryngii mushrooms, commonly known as king trumpet [25]. Cardoon is widely considered a strategic crop for bioenergy production while promoting new opportunities for economic improvement in depressed rural areas of the Mediterranean basin, thanks to its excellent adaptation and components of biomass utilized as feedstock for multipurpose applications [22,24].

The possibility of using compost from cardoon crop residues as a component of cultivation substrates for horticultural crops would allow for a reduction in peat consumption and the conversion of waste into a re-evaluated product, in line with sustainability and circular economy principles. In this regard, positive results were obtained from our investigation on using cardoon-based compost mixed with peat for the production of tomato seedlings [26] and potted plants such as rosemary (Rosmarinus officinalis L.), laurel (Laurus nobilis), and cherry laurel (Prunus laurocerasus) [27].

Lettuce (Lactuca sativa L.) is considered the most important crop in the leafy vegetable sector of horticultural production [28]. It is an annual species belonging to the Asteraceae family, characterized by considerable morphological and genetic variation and comprising seven main groups of cultivars differing phenotypically, usually described as morphotypes [29]. Lettuce consumption has been steadily increasing in mixed salads because it is a minimally processed food product with a long shelf life [30,31] as well as the ongoing concern of consumers to the health qualities and sustainability of food products [32].

Lettuce has high nutritional value and remarkable health properties since it is low in both calories and sodium, and high in vitamins A and C, fiber, and folate. Also, lettuce has high levels of antioxidant compounds such as polyphenols [33,34] whose composition is influenced by the environment, agricultural practices, and the type of cultivar [35,36]. Over the past decade, consumers have increased their attention to the potential benefits of eating vegetables which are an important supply of healthy compounds like polyphenols. Indeed, the current literature suggests that long-term consumption of polyphenol-rich diets protects against the development of certain cancers, cardiovascular disease, diabetes, osteoporosis, and neurodegenerative diseases [37,38,39,40].

The consumption of the so-called “baby leaf” has been experiencing an ever-increasing demand in all European countries for several years now [41]. Many lettuce types and cultivars represent the main ingredients of salad [32,42].

The implementation of soilless cultivation systems is expanding for many horticultural crops to increase yield and quality [12,43] and, among them, lettuce is preferably grown in a soilless cultivation system [44]. Baby leaf lettuce is characterized by a short cycle and there is a considerable interest in promoting its production and quality. It is harvested at an early vegetative phase when seedlings have reached the stage of the first four-six true leaves [45]. The content of vitamins, minerals, and other bioactive compounds may be significantly greater in baby leaf lettuce than in leaves when mature [33,46,47].

The objective of this research was to evaluate the effect of three types of composts from cardoon waste, as a partial replacement of peat, on the production and quality of baby leaf lettuce. The choice of this crop was based on the potential benefits in terms of environmental impact (significant reduction in the use of peat) and human health (quality of the product increasing market demand). Moreover, since lettuce is considered a saline-sensitive species [48], this crop was also considered a well-founded choice from this perspective because cardoon-based compost, being originated from green waste, does not have excessive salinity (such as municipal solid organic waste compost).

2. Materials and Methods

2.1. Compost Preparation and Analysis

The preparation of the composts used for the experiment was carried out at the CREA-CI experimental site of Caserta. Three types of compost, named C1, C2, and C3, were made using the following raw materials mixed in different volumes: shredded, dried cardoon biomass, fresh fruit waste (plums and peaches), and lignocellulosic waste from pruning (

Table 1. Composition of piles for composting in percentage (v:v).

Compost		C1	C2	C3
	cardoon biomass	50.0	68.0	33.3
Feedstocks (% v:v)	fresh fruit waste	25.0	32.0	33.3
	waste from pruning	25.0	0	33.3

C1, C2, and C3 are the codes for the composts obtained from the different mixtures of raw material.

). Three static piles of 2 m³ were prepared and composted for 132 days. The size of particles of cardoon and pruning wastes from fruit trees and garden plants were predominantly in the range of 5–15 cm in length while whole, unmarketable fruits of peach and plum were mixed as such. Composting piles were built manually by arranging in alternate layers different wastes, but taking care that shredded lignocellulosic or cardoon particles were the first layer at the bottom of the pile to drain water and leachate from the pile. Oxygenation was carried out with a flow of air blown into the base of the pile by PVC pipes; three manual turnings to ensure new substrates for the active microbial component and to achieve homogeneity of the composition were carried out. The humidity of the piles was measured weekly and maintained above 50% through watering. Temperature monitoring was carried out using thermoresistance sensors (Datalogger Watchdog series 1000, Spectrum Technologies Inc., 3600 Thayer Court, Aurora, IL, USA) placed in different areas within each pile. Twenty days of thermophilic phase were followed by a mesophilic and slow maturation phase of 112 days. In the thermophilic phase, the temperature was in the range of 50–65 °C for more than 5 days, and in conditions allowing for sanitization [49].

Homogenous samples were collected from each compost and prepared to perform the following physico-chemical analyses. Total carbon (C) and nitrogen (N) determinations were performed by an Elementar Analyzer CN802 (VELP, 20865 Usmate Velate, Italy). Humic and fulvic acids (HA + HF) were measured according to ANPA 3/2001 [50]. pH and electrical conductivity (EC) were determined on water extracts (1:10, w/v) of the composts by a conductivity/pH meter Crison, (HACH LANGE, Barcellona, Spain). The ash content is determined according to the gravimetric procedure suggested in the ANPA 3/2001 [50]. All analyses were performed in triplicate. TC, N, and HA + HF were expressed as % of dry matter (d.m.).

2.2. Compost Phytotoxicity Assessment

The phytotoxicity of the composts was evaluated by cress (Lepidium sativum L.) germination bioassays according to APAT 20/2003 [51]. Petri dishes were prepared by placing 10 cress seeds on sterile filter paper soaked with aqueous extracts of compost at different concentrations (50% and 75%) and incubated at 25 °C for 72 h. Distilled water was used as a control. The bioassay had five replications per treatment. The germination percentage and root length were then measured. For each concentration, the germination index (GI) was estimated using the following formula:

GI% = 100 × (Gc × Lc/Gt × Lt)

where Gc and Gt are the mean number of germinated seeds in the sample and in the control, respectively. Lc and Lt are the mean root length of the sample and of the control, respectively. The germination index (GI) value is then determined by the average of the GI values obtained at the 75% and 50% concentrations. GI values above 65% indicate no phytotoxicity, whereas if the GI is below 40%, the test material is considered phytotoxic [52].

2.3. Experimental Design and Treatments

The experiments were conducted in winter at the CREA-CI of Caserta (lat. 41°04′26.1″ N; long. 14°19′05.5″ W). Two lettuce cultivars, Imperiale and Verde d’Inverno, belonging to the butterhead and romaine types, respectively, from Topseed s.r.l., Sarno (SA), Italy, were grown in a floating system. The experiment was carried out in an unheated, polycarbonate-covered greenhouse under natural daylight conditions.

Seeds were sown on 12 January 2021 in 336-cell polystyrene trays filled with the 6 compost-growing media (C1-60, C1-30, C2-60, C2-30, C3-60, C3-30) prepared by mixing the three composts (C1, C2, and C3) with commercial peat (carbon 51% d.m. and nitrogen 1.2% d.m.) Plantobalt substrate, Plantaflor Humus Verkaufs-GmbH from Vechta, Germany) at two rates (60% and 30% on a volume basis), after sieving to 1.5 cm. A commercial peat-based substrate (Neuhaus Humin-substrate n°17, Klasmann-Deilmann, Geeste, Germany) was used as a control (TC).

The experiment was arranged according to a randomized block experimental design with two cultivars and seven growing media (

Table 2. Composition of the Growing Media utilized.

Growing Medium	Coding
Peat-based substrate	TC
60% Compost 1 + 40% Peat	C1-60
30% Compost 1 + 70% Peat	C1-30
60% Compost 2 + 40% Peat	C2-60
30% Compost 2 + 70% Peat	C2-30
60% Compost 3 + 70% Peat	C3-60
30% Compost 3 + 40% Peat	C3-30

) with three replications and a plant density of 1926 plants per square meter.

Three weeks after sowing, the water in the tanks was substituted with 0.5 g L⁻¹ Hydrofert (Biochim Spa, Medicina, Italy) nutrient solution containing 100, 43.6, and 83.0 mg L⁻¹ of N, P, and K, respectively.

During the crop cycle, air temperatures and relative humidity were registered. All the measurements were collected on a data logger Testo Mod. 175-H1 (Testo SE & Co. KGaA, Titisee-Neustadt, Germany).

In Figure 1, daily maximum, mean, and minimum values of air temperature (a) and relative humidity (b) recorded inside the greenhouse during the growing period of lettuce were reported. The minimum temperature did not fall below 0 °C, while the maximum reached 30 °C degrees in the middle and at the end of the growing cycle.

2.4. Growth Analysis, Yield, Harvest, and Quality Analysis Sampling

Ten days after sowing observation on seed germination was performed for each replicate and treatment and calculated the percentage of seed germination. During the growth, three surveys were carried out at 33, 43, and 54 days after sowing (DAS), sampling 10 plants for each replicate and treatment. The performance parameters measured were the number of true leaves, plant height, and dry weight of organs (root, stem, and leaves). For dry weight determination, fresh samples were placed in an oven at 60 °C for 72 h until constant weight.

Harvest was carried out on 12 March 2021, 54 days after sowing, and the yield was determined as kg of fresh weight (f.w.) m⁻².

At harvest, leaf samples were collected from 10 plants of each replicate, and treatments to determine the leaf content of chlorophyll a, chlorophyll b, and carotenoids. Similarly, leaf samples were gathered and frozen (−80 °C) to determine the content of ascorbic acid, total phenols, total flavonoids, and antioxidant activity.

2.5. Qualitative Analysis

2.5.1. Chemicals and Instruments

All chemicals were purchased from Sigma-Aldrich MERCK KGaAaffilate (Darmstadt, Germany) and CARLO ERBA Reagents s.r.l. (Cornaredo, Italy). with analytical reagent grade ≥ 98%. The colorimetric measurements were performed using a UV-Vis Spectrophotometer Beckman mod. DU64 (5350 Lakeview Parkway S Drive, Indianapolis, IN, USA). Ultrapure water was prepared by a Milli-Q ion exchange system, (Millipore, Merck group, Darmstadt, Germany).

2.5.2. Estimation of Chlorophyll Pigments and Total Carotenoids

The chlorophyll and carotenoid content of the leaves was calculated with the Wellburn formulae [51]. Fresh leaf samples (0.100 g) were extracted in dimethyl sulfoxide (DMSO, 10 mL) under shaking overnight. Thus, absorption was measured at 665, 649, and 480 nm. The chlorophyll (a and b) and total carotenoid content of the leaves were calculated according to the following Formulas (1)–(4):

Chlorophyll a (mg/g) = [(12.19 × A₆₆₅) − (3.45 × A ₆₄₉)] × V/(1000 × W)

(1)

Chlorophyll b (mg/g) = [(21.99 × A₆₄₉) − (4.68 × A ₆₆₅)] × V/(1000 × W)

(2)

Total chlorophyll = chlorophyll a + chlorophyll b

(3)

Total carotenoids (mg/g) = [[(1000 × A₄₈₀) − (2.14 × Ca) − (70.16 × Cb)]/220] × V/(1000 × W)

(4)

where V: Total volume of dimethyl sulfoxide (L); W: Weight of fresh leaf (g); A: Absorbance at the wavelength indicated; Ca: chlorophyll a (μg mL⁻¹); and Cb: chlorophyll b(μg mL⁻¹).

2.5.3. Evaluation of Antioxidant Compounds and Antioxidant Activity

The methods utilized for the determination of ascorbic acid, carotenoids, total phenols (TP), total flavonoids, and antioxidant activity have already been comprehensively described in Morra et al. [53]. The following is a brief description of the procedures used for the determination. Each was performed with three replications.

For ascorbic acid analysis, an aliquot of sample homogenate (2 g) was extracted with 20 mL of a mixture consisting of 30 g L⁻¹ metaphosphoric acid (MPA), 80 mL L⁻¹ acetic acid, and 1 mmol L⁻¹ EDTA [54]. Folin–Ciocalteu phenol reagent was used for subsequent spectrophotometric quantization in accordance with the method described by Jagota and Dani [55]. The results were expressed as mg ascorbic acid per 100 mg of fresh weight.

The same alcoholic extract carried out according to Kaur et al. [56] with minor modifications was used for the analysis of total phenols, flavonoids, and antioxidant activity. Approximately 2 g of lettuce leaf was homogenized in 20 mL of 80:20 methanol/water and then placed in an ultrasonic bath (Elmasonic P, Elma Schmidbauer GmbH, Singen, Germany) at 40 °C for 1 h in the darkness. After centrifugation for 20 min at 10,000 at 4 °C the solution containing the extracted metabolites was recovered and stored at −20 °C until analyses were carried out.

Total phenols were determined utilizing the method of Folin–Ciocalteu [57]. Color development is carried out at t.a. for 120 min in dark glass vials where 1 mL H₂O, 100 μL of extract, 100 μL of Folin–Ciocalteu reagent, and 800 μL of Na₂CO₃ 75 g L⁻¹ are successively mixed. The reading wavelength was 765 nm. Analytical values were reported as mg of gallic acid equivalent (GAE) per 100 g of fresh weight.

The total flavonoid content was determined by spectrophotometric reading at 510 nm of the colored complexes that this class of compounds forms with aluminum salts in the presence of nitrite in a basic environment. The procedure described by Zhishen et al. [58], with modifications [59] was performed. The results were indicated as mg catechine equivalent (CE) per 100 g of fresh weight.

The antioxidant activity was determined on lettuce extract using the DPPH radical scavenging method [60,61]. Lettuce extract (0.1 mL), properly diluted, was added to the DPPH solution in methanol (3.9 mL of 0.0634 mM), shaken vigorously, and allowed to stand at room temperature for 60 min, when the reaction kinetic reached a steady state. The decrease in absorbance at 515 nm, proportional to the antioxidant power of the sample, was recorded. The antioxidant activity was estimated as the equivalent concentration of Trolox used as a reference standard for constructing the calibration line (μmol TE 100 g⁻¹ fw).

2.6. Statistical Analysis

All biometrical, agronomical, and qualitative data were subjected to Shapiro–Wilk and Levene tests for normality and homogeneity of variance, respectively. Two-way analysis of variance (ANOVA) for the two factors, cultivar and growing medium, according to a randomized block experimental design with three replications was performed. When the effect of a source of variability was significant, means were separated by the Tukey HSD test to a 0.05 probability level. STATISTICA Software Version 7.1 for Windows (2000 StatSoft, Inc., Tulsa, OK, USA) was used [62]. Principal Component Analysis (PCA) was conducted using the statistics software R (version 4.1.2) using the package “FactoMineR” on mean-centered and scaled data [63].

3. Results and Discussion

3.1. Compost and Growing Medium Characteristics

The main chemical characteristics of the composts obtained are shown in

Table 3. Main chemical characteristics of the composts used.

Compost	C % d.m.	N % d.m.	C/N	HA + HF % d.m.	pH	EC dS/m	Ash % d.m.	GI %
C1	37.9 ± 0.5	3.4 ± 0.1	11.0 ± 0.3	18.4 ± 5.1	7.9 ± 0.2	0.90 ± 0.1	21.5 ± 2.1	124.1
C2	35.2 ± 0.6	2.2 ± 0.1	16.3 ± 0.7	11.8 ± 0.0	8.0 ± 0.4	0.77 ± 0.2	32.3 ± 1.9	132.8
C3	41.1 ± 0.1	2.6 ± 0.1	15.8 ± 0.2	21.8 ± 0.1	7.9 ± 0.3	0.82 ± 0.3	22.1 ± 2.2	118.8

. Slight differences in total carbon and total nitrogen content were observed but in a range similar to that of other plant-based composts [13,14,15,64]. The carbon/nitrogen ratio was lower in C1 compost (11.0) than in C2 and C3 composts (16.3 and 15.8, respectively), however, with values suggesting good compost maturation [15]. In compost C2, the humic and fulvic acids were lower and the ash content was higher than in the other two composts, likely a function of the composition of the compost heap, which did not contain lignin-rich pruning waste. However, all were between 10 and 20 percent of dry matter, a range considered desirable for a quality compost. The pH and conductivity values were comparable among the composts and in a range suitable for use as a growing medium component [11,13].

The germination indices (GI) found, being higher than 65%, indicated the absence of phytotoxicity in the three composts [52]. Interestingly, values of over 100% suggest a stimulating effect on Lepidium sativum L. germination of composts.

In addition, the temperature trend recorded in the thermophilic phase of the transformative process of feedstocks (composting) was in the range of 50–65 °C for more than 3–5 days, showing the occurrence of conditions that, as reported by Aguilar et al. [49], allow for sanitization of compost.

In

Table 4. Main chemical characteristics of the growing media utilized.

Growing Medium	C % d.m	N % d.m.	pH	EC dS/m	Ash % d.m.
TC	51.0 ± 0.2	1.4 ± 0.02	6.0 ± 0.1	0.35 ± 0.0	9.6 ± 0.8
C1-60	42.2 ± 0.1	2.3 ± 0.09	7.2 ± 0.2	0.68 ± 0.2	18.2 ± 1.6
C1-30	46.9 ± 0.9	1.8 ± 0.10	6.4 ± 0.2	0.52 ± 0.1	10.7 ± 1.3
C2-60	36.0 ± 0.8	2.7 ± 0.10	7.3 ± 0.3	0.60 ± 0.2	26.6 ± 0.9
C2-30	45.7 ± 0.2	1.6 ± 0.01	6.4 ± 0.1	0.47 ± 0.1	15.5 ± 1.8
C3-60	41.8 ± 0.6	2.7 ± 0.11	7.1 ± 0.3	0.50 ± 0.1	17.8 ± 1.0
C3-30	47.3 ± 0.3	1.9 ± 0.01	6.4 ± 0.2	0.63 ± 0.2	13.3 ± 1.1

The values are means ± standard deviation: TC: Peat-based substrate; C1-60: 60% Compost 1 + 40% Peat; C1-30: 30% Compost 1 + 70% Peat; C2-60: 60% Compost 2 + 40% Peat; C2-30: 30% Compost 2 + 70% Peat; C3-30: 30% Compost 3 + 70% Peat; and C3-60: 60% Compost 3 + 40% Peat.

, the main chemical characteristics of the growing media utilized are reported. The values of parameters measured in all compost-containing substrates were in a range not detrimental to germination and initial seedling growth, similar to peat-based substrate [12,13].

3.2. Effect of Growing Medium on the Growth and Yield

Cultivar and growing medium did not influence seed germination, as was expected for the latter based on the absence of phytotoxicity of the three composts, and the pH and EC values measured in all compost-containing substrates, which, compared to those of the control, could be presumed not to inhibit germination.

Statistical analysis performed by two-way ANOVA on the growth components i.e., number of true leaves, plant height, and dry weight of organs (root, stem, and leaves), at 33 days after sowing, showed that the parameters considered were affected by the cultivar and growing medium and their interaction, except for the root dry weight.

Imperiale (IMP) lettuce showed significantly higher leaf number than Verde d’Inverno (VDI) (3.0 vs. 1.9), while lower height (6.9 vs. 8.7 cm). The dry weight of stem and leaves was significantly higher for VDI compared to IMP (10.60 vs. 5.27, 19.14 vs. 15.98, respectively).

In

Table 5. Effect of the cultivar (C) and growing medium (GM) interaction on the number of leaves; plant height; and root, stem, and leaves dry weight per plant at 33 days after sowing.

C × GM	Leaves			Plant Height				Root			Stem				Leaves
	n°			(cm)				(mg d.w.)			(mg d.w.)				(mg d.w.)
IMP × TC	3.0	±	0.5	7.0	±	0.7	cde	5.4	±	1.9	6.7	±	1.2	cde	20.0	±	0.1	abc
IMP × C1-60	2.9	±	0.5	6.6	±	0.9	de	5.2	±	2.2	4.8	±	0.4	de	14.4	±	2.4	c
IMP × C1-30	2.9	±	0.3	6.5	±	0.8	e	6.0	±	2.1	5.9	±	0.8	cde	15.8	±	2.4	bc
IMP × C2-60	3.0	±	0.3	7.5	±	0.7	bc	5.0	±	2.5	4.9	±	0.4	de	17.5	±	0.2	abc
IMP × C2-30	3.0	±	0.3	7.1	±	0.7	cde	5.3	±	0.7	4.7	±	0.5	de	16.3	±	3.4	abc
IMP × C3-60	3.0	±	0.4	6.8	±	0.5	cde	7.5	±	2.4	4.1	±	0.6	e	13.8	±	1.8	c
IMP × C3-30	3.0	±	0.3	6.8	±	0.9	cde	5.4	±	2.0	6.0	±	1.0	cde	14.3	±	2.0	c
VDI × TC	1.9	±	0.3	8.3	±	0.9	a	5.1	±	0.5	8.1	±	0.2	bcd	14.6	±	1.3	c
VDI × C1-60	2.0	±	0	7.2	±	0.8	cd	6.7	±	0.3	8.7	±	2.5	bc	16.1	±	3.6	abc
VDI × C1-30	2.0	±	0	8.2	±	0.7	a	7.4	±	0.1	12.1	±	3.3	a	22.7	±	3.1	a
VDI × C2-60	2.0	±	0	8.4	±	0.7	a	5.8	±	0.9	12.3	±	0.5	a	20.2	±	3.3	abc
VDI × C2-30	1.8	±	0.4	8.0	±	1.3	ab	6.5	±	1.8	9.5	±	0.8	abc	18.6	±	1.5	abc
VDI × C3-60	2.0	±	0	7.9	±	0.6	ab	6.6	±	1.4	12.0	±	0.1	a	19.6	±	1.2	abc
VDI × C3-30	2.0	±	0.3	8.0	±	0.7	ab	8.8	±	1.4	11.6	±	0.3	ab	22.3	±	1.1	ab
Significance		ns			**				ns			**					***

The values are means ± standard deviation. Within each column, different letters indicate significant differences according to Tukey’s test at p ≤ 0.05. IMP: Imperiale lettuce cv; VDI: Verde d’Inverno lettuce cv; TC: Peat-based substrate; C1-60: 60% Compost 1 + 40% Peat; C1-30: 30% Compost 1 + 70% Peat; C2-60: 60% Compost 2 + 40% Peat; C2-30: 30% Compost 2 + 70% Peat; C3-30: 30% Compost 3 + 70% Peat; C3-60: 60% Compost 3 + 40% Peat; ns: not significant; significance **: p < 0.01; ***: p < 0.001.

, the effect of the cultivar (C) and growing medium (GM) interaction on leaf number; plant height; and root, stem, and leaves dry weight 33 days after sowing is reported. The plant height of IMP lettuce was not affected by the growing medium, while VDI plants grown in C1-60 (VDI × C1-60) showed a lower height compared with those grown in control (VDI × TC). Similar values of dry weight (root, stem, leaf) of IMP plants grown with the different substrates were found; instead, VDI × C1-30, VDI × C2-60, and VDI × C3-60 showed higher mean values of stem dry weight, compared with VDI × TC. VDI × C1-30 also positively influenced leaf dry weight.

At 43 days after sowing, at 33 DAS, VDI showed higher mean values than IMP for all the parameters considered, except for leaf number. Indeed, VDI was high at 8.8 while IMP was 7.6 cm, and the dry weight of roots, stems, and leaves of VDI compared with that of IMP was 26.2 vs. 18.1, 16.20 vs. 13.2, and 70.9 vs. 51.1 mg d.w., respectively.

The effect of the cultivar (C) and growing medium (GM) interaction was significant for all considered parameters, as shown in

Table 6. The main effect of the cultivar (C) and growing medium (GM) interaction on the number of leaves, plant height, and root, stem, and leaves dry weight per plant at 43 days after sowing.

C × GM	Leaves				Plant Height				Root				Stem				Leaves
	(n°)				(cm)				(mg d.w.)				(mg d.w)				(mg d.w.)
IMP × TC	3.5	±	0.6	e	7.6	±	0.7	e	16.9	±	2.6	c	12.2	±	0.3	e	44.8	±	2.5	d
IMP × C1-60	4.2	±	0.6	a	8.4	±	1.1	bcd	21.1	±	1.5	abc	15.4	±	2.2	a-e	58.9	±	7.9	bcd
IMP × C1-30	3.8	±	0.4	cde	7.3	±	0.7	e	18.8	±	1.3	bc	11.3	±	1.4	e	48.8	±	0.4	d
IMP × C2-60	4.1	±	0.6	ab	7.8	±	1.0	de	17.1	±	1.2	bc	12.6	±	1.8	de	49.6	±	5.7	d
IMP × C2-30	4.1	±	0.5	abc	7.4	±	0.6	e	16.9	±	4.4	c	13.1	±	1.0	cde	53.4	±	3.5	cd
IMP × C3-60	4.0	±	0.4	a-d	7.2	±	0.7	e	16.9	±	4.1	c	14.9	±	1.1	b-e	47.7	±	0.7	d
IMP × C3-30	3.8	±	0.4	b-e	7.4	±	0.6	e	18.9	±	5.7	bc	13.3	±	1.7	cde	54.8	±	5.1	cd
VDI × TC	4.0	±	0	a-d	9.2	±	1.0	a	32.6	±	4.4	a	11.3	±	1.9	e	77.7	±	7.2	ab
VDI × C1-60	4.0	±	0.2	a-d	8.8	±	0.8	abc	28.6	±	8.0	abc	17.5	±	2.8	a-d	70.3	±	9.4	abc
VDI × C1-30	3.7	±	0.5	de	8.7	±	0.9	abc	22.3	±	2.0	abc	18.0	±	0.1	abc	70.5	±	8.7	abc
VDI × C2-60	4.0	±	0.2	a-d	9.1	±	0.9	ab	29.3	±	4.0	ab	20.3	±	1.4	a	80.7	±	13	a
VDI × C2-30	4.1	±	0.3	ab	9.3	±	0.7	a	28.8	±	5.6	abc	18.5	±	1.1	ab	83.3	±	5.2	a
VDI × C3-60	4.0	±	0.2	a-d	8.3	±	0.8	cd	22.3	±	4.2	abc	13.5	±	3.1	b-e	63.4	±	4.0	a-d
VDI × C3-30	3.9	±	0.3	a-d	8.3	±	0.6	cd	19.8	±	1.9	bc	13.0	±	0.7	cde	50.3	±	7.6	d
Significance		***				***				*				***				***

The values are means ± standard deviation. Within each column, different letters indicate significant differences according to Tukey’s test at p ≤ 0.05. IMP: Imperiale lettuce cv; VDI: Verde d’Inverno lettuce cv; TC: Peat-based substrate; C1-60: 60% Compost 1 + 40% Peat; C1-30: 30% Compost 1 + 70% Peat; C2-60: 60% Compost 2 + 40% Peat; C2-30: 30% Compost 2 + 70% Peat; C3-30: 30% Compost 3 + 70% Peat; C3-60: 60% Compost 3 + 40% Peat; *: p < 0.05; ***: p < 0.001.

. With regard to the number of leaves present at the time of the survey, IMP × C1-60 showed the highest number of leaves and IMP × TC the lowest. For VDI, significant differences were observed only between C2-30 and C1-30 growing media (4.1 vs. 3.7). C1-60 positively affected the height of IMP lettuce plants, while the other growing media were comparable to the control (IMP × TC). C3-60 and C3-30 gave shorter plants of VDI lettuce than VDI × TC. For the IMP variety, no statistically significant differences were registered between mean values of dry weight of root, stem, and leaves grown on the different growing media. All growing media except C3-30 gave a root dry weight of VDI variety comparable to the control (VDI × TC). VDI plants grown both in C1 and C2 at two rates showed stem dry matter greater than those grown in control (VDI × TC). The leaf dry weight of VDI was negatively influenced only by the C3-30 growing medium.

Statistical analysis performed by two-way ANOVA relative to the survey at 54 days after sowing revealed a significative effect of the cultivar, growing medium, and their interaction on all the growth parameters considered.

As with previous surveys at 33 and 43 DAS, VDI showed higher mean values than IMP for all parameters considered except for the number of leaves. VDI was high at 9.6 cm while IMP was 8.3 cm. The dry weight of the roots, stems, and leaves of VDI compared with that of IMP were 63.4 vs. 41.6, 29.8 vs. 16.5, and 154.4 vs. 132.5 mg d.m., respectively.

In

Table 7. Effect of the cultivar (C) and growing medium (GM) interaction on leaf numbers; plant height; and root, stem, and leaves dry weight per plant at 54 days after sowing.

C × GM	Leaves				Plant Height				Root				Stem				Leaves
	(n°)				(cm)				(mg d.w.)				(mg d.w)				(mg d.w.)
IMP × TC	5.0	±	0.6	ef	7.8	±	1.1	f	34.0	±	4.1	c	13.5	±	1.3	e	111	±	13	c
IMP × C1-60	6.1	±	0.8	a	9.1	±	0.8	cd	46.2	±	8.6	bc	20.1	±	1.2	cde	170	±	18	ab
IMP × C1-30	4.9	±	0.9	ef	8.2	±	0.9	f	30.9	±	4.3	c	17.1	±	0.9	de	128	±	6.5	bc
IMP × C2-60	5.5	±	0.5	bcd	9.1	±	0.7	cd	54.4	±	3.5	abc	19.5	±	0.1	cde	141	±	23	bc
IMP × C2-30	5.5	±	0.8	bcd	8.1	±	0.7	f	36.4	±	4.1	c	14.2	±	0.7	e	118	±	5.2	bc
IMP × C3-60	5.7	±	0.6	ab	8.3	±	0.7	ef	48.2	±	2.8	abc	16.9	±	1.7	de	145	±	19	bc
IMP × C3-30	5.6	±	0.5	bc	7.7	±	0.8	f	41.2	±	5.6	bc	14.0	±	0.8	e	115	±	9.2	bc
VDI × TC	5.1	±	0.3	def	10.1	±	0.8	ab	58.8	±	2.9	abc	30.5	±	4.1	ab	139	±	3.8	bc
VDI × C1-60	5.2	±	0.4	c-f	9.6	±	0.6	abc	62.6	±	15	abc	28.9	±	2.0	abc	144	±	9.0	bc
VDI × C1-30	5.1	±	0.4	c-f	9.4	±	1.1	bcd	79.3	±	19	a	32.3	±	1.8	ab	171	±	21	ab
VDI × C2-60	5.0	±	0.5	ef	10.3	±	0.6	a	62.5	±	7.4	abc	28.3	±	8.5	abc	155	±	23	abc
VDI × C2-30	5.4	±	0.6	b-e	10.0	±	0.8	ab	72.9	±	16	ab	35.7	±	4.6	a	202	±	34	a
VDI × C3-60	5.0	±	0.2	ef	9.1	±	0.7	cd	56.4	±	6.1	abc	24.7	±	4.5	bcd	132	±	32	bc
VDI × C3-30	4.8	±	0.5	f	8.9	±	0.7	de	51.2	±	9.2	abc	28.2	±	2.6	abc	137	±	12	bc
Significance		***				***				*				**				***

The values are means ± standard deviation. Within each column, different letters indicate significant differences according to Tukey’s test at p ≤ 0.05. IMP: Imperiale lettuce cv; VDI: Verde d’Inverno lettuce cv; TC: Peat-based substrate; C1-60: 60% Compost 1 + 40% Peat; C1-30: 30% Compost 1 + 70% Peat; C2-60: 60% Compost 2 + 40% Peat; C2-30: 30% Compost 2 + 70% Peat; C3-30: 30% Compost 3 + 70% Peat; C3-60: 60% Compost 3 + 40% Peat; not significant; significance *: p < 0.05; **: p < 0.01; ***: p < 0.001.

, the effect of the cultivar (C) and growing medium (GM) interaction on the number of leaves; plant height; dry weight of root, stem, and leaves; and yield 54 days after sowing is reported. IMP lettuce grown on all based-compost growing media, except C1-30, gave a greater leaf number than IMP × TC; meanwhile, for VDI, growing media did not influence leaf number. C1-60 and C2-60 growing media positively affected the height of IMP lettuce plants, while the other substrates were comparable to the control (IMP × TC). VDI × C3-60 and VDI × C3-30 were shorter than VDI lettuce × TC.

As in the surveys at 33 and 43 days after sowing, IMP lettuce showed no statistically significant differences in root, stem, and dry weight in dependence by substrates. Instead, at 54 days IMP × C1-60 gave a greater leaf dry matter statistically significant than IMP × TC. For VDI among the compost-based substrates, the best was C2-30. VDI × C2-30 showed higher leaf dry matter values than the control (VDI × TC).

Compost has been shown to improve the amount of nutrients available to plants and enhance their growth with increased dry weight of both shoots and roots [65,66,67]. According to Trevisan et al. [68], compost, over time, is a source of nutrients that are absorbed by plant roots [69], and its beneficial effect on yield is probably related to auxin-like molecules from the humic fraction.

Similar to our results, Moschou et al. [70] found that in hydroponic lettuce (Lactuca sativa var. Tanius) cultivation waste-based compost (from fruits, and vegetable and olive pruning) substrate significantly increased leaf dry weight (19.8%) of the plants compared to the conventional organic growing media constituents (cocodust).

De Falco et al. [15] reported that compost produced from crop waste of vegetable green leaf utilized as a component of substrate to baby leaf species cultivation increased the dry weight of both shoot and root compared to the peat substrate. For lettuce (Lactuca sativa var. Batavia verde Falstaff), the increase in dry weight was only observed for the roots at the lowest dose (C25).

These different results could be due to differences in feedstocks, the transformation process, and the characteristics of the compost and its mixing ratios that influence the properties of the growing media. In addition, environmental conditions and cultivar-specific responses could be taken into account [67,71]. Concerning the latter, our findings on dry matter accumulation at the three surveys conducted at 33, 43, and 54 DAS, indicated varietal-specific behavior of the lettuce cultivars with respect to the growth medium. At 33 and 43 DAS, the compost-containing growing media gave comparable results to the control in IMP lettuce plants; at harvest, C1-60 showed leaf dry weight even greater than the control. Instead, for VDI, already at 33 days after sowing, some compost-based growing media (C2-30, C2-60) gave higher dry matter values than the control.

As for yield, the VDI cultivar resulted in more productivity than IMP (2.6 vs. 1.8 kg f.w. m⁻²). The varieties responded differently to the growing medium: the IMP variety showed the highest production values C1-60 (2.4 kg f.w. m⁻²) as did the VDI with C2-30 growing medium (3.2 kg f.w. m⁻²) (Figure 2).

The average yield found in our experiment was comparable to that measured by Gimenez et al. [72] who reported a yield of about 2.2 and 2.4 kg m⁻² of baby leaf lettuce (Lactuca sativa L. cv Antoria (red lettuce) grown, respectively, in peat and compost (from fruit and vegetable wastes) in a floating system. Our results agree with Gimenez et al. [73] who found no negative influence on plant development by composts (from agricultural wastes) utilized as growing media for the production of baby leaf lettuce (Lactuca sativa L., red lettuce) in a floating system. Moreover, as for our results, some compost-based substrates of cultivation improved plant growth more than the peat-based ones, achieving higher yields [73]. It has been shown that suitable characteristics of water-holding capacity, EC, aeration, and bulk density allow satisfactory conditions for healthy plant growth, which guarantees an increase in the photosynthetic potential of the leaves [11,67,69]. These characteristics are affected by the porosity in the media, which promotes plant growth [69]. Our results indicated that the cardoon-based composts were suitable to meet this condition up to the highest peat replacement volume tested (60%). This is noteworthy since it has been reported that both plant growth and yields were higher when the volume of compost was in a relatively low quantity (25–50%) in the growing medium [15,67]. Barker and Bryson [74] indicated that the optimal amount of compost may depend on its type and reported that a 25% mixture of non-food waste compost and peat resulted in the best growth for lettuce. In addition, in work conducted to evaluate green compost and palm fiber trunk waste as peat replacement for seedling production, Ceglie et al. [71] reported that optimal performance was reached with 20% and 35% peat for tomato and melon seedlings, respectively, while up to 60% peat was required for lettuce.

In a study on the application of compost derived from fresh vegetable residues for the production of lettuce, Porto et al. [75], evaluated different mixtures with the commercial substrate using four different cultivars and reported that the best results were obtained using compost amounts between 20.1 and 26.7% of the growing medium.

Moreover, Porto et al. [75] observed a significant interaction between cultivar and growing medium on plant fresh mass. Also, Marutani and Clemente [76] highlighted the interaction effects of lettuce cultivar type and compost (food waste–based) growing medium on the fresh weight of the shoots, as occurred in our findings. These findings point out the importance of a genotype-production system for this crop and show that some cultivars respond better to the use of compost [75].

3.3. Effect of Growing Medium on the Quality of Lettuce

3.3.1. Effect of Growing Medium on Content of Chlorophylls and Carotenoids

Statistical analysis was performed by two-way ANOVA on leaf chlorophyll a, chlorophyll b, and total chlorophyll. The ratio of chlorophyll a/b and carotenoids at harvesting showed a significant effect of cultivar (C), while no significant effect of growing medium (GM) was observed. Cultivar (C) and growing medium (GM) interaction influenced only chlorophyll b and the ratio chlorophyll a/b.

As for the effect of cultivar, chlorophyll a, total chlorophyll, and carotenoids were higher in the YDI than IMP variety, as shown in

Table 8. Effect of the cultivar (C) on the leaf chlorophyll a, total chlorophyll, and carotenoid content at harvesting.

C	Chlorophyll a (mg 100 g−1 f.w.)	Total Chlorophyll (mg 100 g−1 f.w.)	Carotenoids (mg 100 g−1 f.w.)
IMP	40.1 ± 4.9 b	52.1 ± 3.9 b	13.4 ± 1.0 b
VDI	56.7 ± 7.3 a	82.9 ± 5.9 a	21.2 ± 2.3 a
Significance	***	***	***

The values are means ± standard deviation. Within each column, different letters indicate significant differences according to Tukey’s test at p ≤ 0.05. IMP: Imperiale lettuce cv; VDI: Verde d’Inverno lettuce cv; significance ***: p < 0.001.

. The chlorophyll b content also resulted higher in VDI lettuce than in IMP (26.2 vs. 12.0 mg 100 g⁻¹ f.w.), while the chlorophyll a/b ratio was lower (2.2 vs. 3.3 mg 100 g⁻¹ f.w.).

The average values of total chlorophyll found in this study were in the range of 51.4 to 86.1 mg 100 g⁻¹ f.w. These data are in agreement with the results of Martinez et al. [47], who reported in 11 baby leaf lettuce varieties mean values from 33.9 to 60.6 mg 100 g⁻¹ f.w. Similar results were found by Amanda et al. [77] and Di Mola et al. [78], who found in baby leaf lettuce an average chlorophyll content, respectively, of 49 and 50.8 mg 100 g⁻¹ f.w. As for carotenoids, Martinez [47], reported in 11 baby leaf lettuce varieties, an average carotenoid content of 36.76 μg g⁻¹ f.w. in the range from 1.85 to 126.98 μg g⁻¹ f.w.

The differences found in chlorophyll and carotenoid content in lettuce cultivars are likely related to the structure of the head, which differs between IMP, butterhead type (closed head), and VDI, romaine type (open head). It has been reported that the different morphology of lettuce heads influences some nutritional and bioactive compounds; lettuce morphotypes with open heads have a greater surface area exposed to light that promotes photosynthetic activity and thus the synthesis of sugars, chlorophylls, and other relevant compounds, while cultivars with closed heads have a smaller surface area leading to lower content of metabolites [79]. Mou and Ryder [80] also report variations in carotenoid content depending on head morphology in lettuce cultivars.

In Figure 3, the effect of the cultivar (C) and growing medium (GM) interaction on chlorophyll b and the ratio of chlorophyll a/b at harvesting is shown. In the leaves of VDI grown on C1-60 and C2-60, chlorophyll b content was higher than those grown on TC while for Imperiale lettuce, chlorophyll b leaf content was not affected by growth medium. Significant differences in the leaf chlorophyll a/b ratio were observed for cv VDI that showed the highest values of the ratio when grown on control (TC). The chlorophyll a/b ratio of the leaves of the two lettuce varieties grown on TC was comparable; however, unlike VDI, no significant differences in the leaf chlorophyll a/b ratio were observed for cv Imperiale in dependence on the growing medium.

Chlorophyll and carotenoids are both chromophores involved in photosynthesis. Chlorophyll a is an essential photosynthetic pigment present in the photosynthetic reaction centers. Generally, the ratio of chlorophyll a to chlorophyll b is one to three. Carotenoids are plant pigments involved in the photoprotection of chlorophylls. The health status of plants can be assessed indirectly on the basis of chlorophyll concentrations [81]. Our findings highlighted a similar content of chlorophyll a and carotenoids in the leaf of plants grown in medium with compost with respect to control, indicating a comparable health status.

The ratio of chlorophyll a to chlorophyll b can give indications about the physiological state of the plant as it can change with the type and degree of abiotic stress [81]. In our study, the VDI cultivar showed a lower value of the chlorophyll a to b ratio when grown on a substrate containing compost compared to TC indicating a response cultivar-specific to compost indicative of stress conditions.

3.3.2. Effect of Growing Medium on the Leaf Antioxidant Compounds and Antioxidant Activity

Statistical analysis performed by two-way ANOVA on the leaf antioxidant compounds and antioxidant activity at harvesting showed a significant effect of cultivar (C), growing medium (GM), and their interaction.

The content of ascorbic acid was found higher in VDI than in the Imperiale variety (67.3 vs. 56.1 mg 100 g⁻¹ f.w.). The leaf content of this compound was affected by the interaction of the cultivar (C) with the growing medium (GM). The average values of total ascorbic acid found in this study were in the range of 53.6 to 71.8 mg 100 g⁻¹ f.w. These data are in agreement with the results of Martinez et al. [47] who reported in 11 baby leaf lettuce varieties mean values from 39.2 to 86.5 mg 100 g⁻¹ f.w.

In Figure 4, the effect of the cultivar (C) and growing medium (GM) interaction on the ascorbic acid at harvesting is represented. VDI showed the highest values in plants grown in the C2-60 and C3-30 substrates. No significant differences in the leaf ascorbic acid content were observed for cv IMP in dependence on the growing medium. The leaf ascorbic acid content resulted in a comparison between the two cultivars when grown on a control growing medium, TC.

The observed increase in ascorbic acid agrees with previous results in which the ascorbic acid concentration was higher with compost in strawberries, peppers, and rocket salad [82], and in which a cultivar-specific response to lettuce in closed soilless cultivation was found [83].

Ascorbic acid is a potent biological antioxidant involved in both pathogen defense mechanisms and growth modulation via phytohormone signaling [36]. The observed increase in ascorbic acid could be related to the presence of compounds in the compost that increase the antioxidant capacity of plants, as suggested by Lakhdar [84]. This beneficial effect is important both for the nutritional properties of the lettuce and for the marketing of the product. In fact, based on Saini et al. [85], the shelf life of baby leaf lettuce is favored by high concentrations of ascorbic acid and tocopherols by safeguarding phenolic compounds.

The polyphenols, flavonoids, and antioxidant activity were found higher in IMP than VDI cultivar (respectively, 488.6 vs. 435.9 mg GAE 100 g⁻¹ f.w., 292 vs. 274.1 CE mg 100 g⁻¹ f.w., and 2337.7 vs. 2016.1 µmol TE 100 g⁻¹ f.w.).

In the present work, the polyphenol content was in the range of 367.1 to 551.4 mg GAE 100 g⁻¹ f.w., resulting in slightly higher than the ones found by Gimenez et al. [73] and Martinez et al. [47], taking into account that the average dry matter (d.w.) value found in our samples was 15%. Indeed, these authors reported an average value, respectively, of 2467.8 mg GAE 100 g⁻¹ d.w. and 18.43 mg GAE g⁻¹ d.w. According to Yang et al. [34], several studies indicate that lettuce genotypes may have different polyphenol contents and levels of specific phenolic compounds, and they reported that in a study in which 25 lettuce cultivars were examined, the polyphenols ranged between 104 and 857 mg GAE per 100 g f.w.

In Figure 5, the effect of the cultivar (C) and growing medium (GM) interaction on the polyphenols (a) and flavonoids (b) at harvesting is shown. IMP × TC showed the highest polyphenol content while VDI × TC was the lowest (551.1 vs. 367.1 mg 100 g⁻¹ f.w.). Leaf polyphenols of IMP × C1-60 and IMP × C2-60 were lower than those of IMP × TC; instead, the content of VDI × C1-60 and VDI × C2-60 were higher than VDI × TC. VDI × C2-30 e VDI × C3-60 did not differ from VDI × TC (Figure 5a).

Polyphenols, a family of naturally occurring organic substances comprising a large number of highly bioactive polyfunctional compounds, are involved in the response to abiotic and biotic stresses and are induced by genetic, environmental, and eliciting factors. The increased polyphenol content could be linked to the activation of phenylalanine ammonia-lyase (PAL), the regulatory enzyme of phenylpropanoid metabolism [86].

Also with regard to flavonoid content, IMP × TC showed the highest average value (331.9 mg 100 g⁻¹ f.w.) while VDI × TC was the lowest (209.5 mg 100 g⁻¹ f.w.). The flavonoid content of IMP × C1-60 (261.4 mg 100 g⁻¹ f.w.) was lower than that of IMP × TC; VDI × C2-30 (255.1 mg 100 g⁻¹ f.w.), which did not differ from VDI × TC (Figure 5b).

The average values of total flavonoid content found in this study were in the range of 272.2.6 to 331.9 mg 100 g⁻¹ f.w. These data are in agreement with the results of Gimenez et al. [73] who reported baby leaf mean values of about 2300 to 2400 mg kg⁻¹ f.w.

Flavonoids constitute a large and diverse group of phenolic compounds ubiquitous in higher plants in which they fulfill many functions. Flavonoids are involved in mechanisms of response to biotic and abiotic stresses, color and chemical signaling, and other auxiliary roles. They constitute the first line of defense against photodamage by UV [87]. As well as polyphenols, we observed a rise of flavonoid content in VDI lettuce when grown in medium containing compost that presumably led to cultivar-specific activation of a relative biosynthetic pathway.

With regards to the antioxidant activity, the highest value was found in IMP × C3-60 (2769.3 µmol TE 100 g⁻¹ f.w.) and the lowest in VDI × TC (1548.7 µmol TE 100 g⁻¹ f.w.). The antioxidant activity of IMP × C1-60 and IMP × C1-30 was lower than those of IMP × TC; VDI × C2-60 and VDI × C2-30 did not differ from VDI × TC (Figure 6).

Leaf antioxidant activity found in this study was in the range from 1548.7 to 2769.3 µmol TE 100 g⁻¹ f.w. (from 32 to 69 expressed as % of inhibition). These data agree with the results of Martinez et al. [47], who reported in 11 baby leaf lettuce varieties mean values from 32.61 to 87.02% of inhibition. Similar results were found by Di Mola et al. [78], who found in baby leaf lettuce (cv Zarina) an antioxidant activity of 26.2 mmol TE 100 g⁻¹ d.w. Indeed, our results were in the range from 10.3 to 18.5 expressed as mmol TE 100 g⁻¹ d.w., taking into account that the average dry matter (d.w.) value found in our samples was about 15%.

Antioxidant capacity is related to the phenolic compound content. IMP lettuce displayed higher antioxidant activity in terms of radical scavenging activity on the DPPH free radical compared to VDI, but the first cultivar did not show an increase of antioxidant activity when grown in a growing medium containing compost differently from the second. This agrees with the content of antioxidant compounds that have been found.

It has been reported that flavonoids, a class of polyphenols, are selectively absorbed by roots and are able to move long distances within the plant, so the content of phenolic compounds in the compost may play a role in the transport and synthesis of antioxidant molecules in lettuce [88,89]. Santos et al. [88] conducted a study to assess the effect of compost polyphenols on the phenolic composition, vitamin C, and carotenoid contents of lettuce and reported that the yield and phenolic compounds of lettuce are significantly influenced by the type of compost and that the lettuce’s response to the growing medium is highly dependent on genetics. This agrees with our findings which highlighted a higher total polyphenol content in the VDI cultivar when grown in compost-containing media, indicating a cultivar-specific activation of the phenylpropanoid biosynthetic pathway. Interestingly, the VDI cultivar in compost-containing media, while showing a total chlorophyll content not different from the control, had a lower chlorophyll a to b ratio indicating a stress condition. It should be pointed out that cardoon biomass is particularly rich in polyphenols [90] and since the three composts, C1, C2, and C3, were obtained using 50%, 68%, and 33% of cardoon biomass, respectively, they may have a different polyphenol content, potentially impacting differently on lettuce yield and quality. It should be noted that for the VDI cultivar, in general, the lowest values of the growth components were shown by C3, which was obtained with the lowest amount of cardoon.

3.4. Principal Component Analysis

The data obtained from measurements of plant growth components and biochemical analyses were statistically analyzed using the principal component analysis (PCA) method for each cultivar. For the IMP cultivar (butterhead lettuce), the outcome of the analysis is given in Figure 7 (Figure 7a represents the scores of the samples grown on different growing media on the first two principal components; Figure 7b shows the vector of each variable on the first two principal components). The two first principal components represented 65.1% of the variance. The PC1 axis, which explained the 38.7% of the variability, was positively correlated with all growth parameters, which was correlated negatively with polyphenols, flavonoids, antioxidant activity, and only slightly with ascorbic acid. The PC2 axis, which explained the 26.4% of the variability, instead was strongly correlated with chlorophylls a and b, carotenoids, and in part with the other biochemical parameters except the antioxidant activity, while the correlation with morphological parameters was almost always low. A clear separation of individuals was observed between growing media with compost and those without compost. Samples grown on growing medium without compost, in fact, evidenced higher content of biochemicals, above all of the polyphenols, flavonoids, ascorbic acid, and in some cases of chlorophyll b, while samples grown on substrates with compost evidenced higher values of morphological parameters. It is to be noted, furthermore that C1 and C2 at dose 60 had the higher values of grown components.

Figure 8 shows the PCA outcome of the cultivar VDI (romaine lettuce). The two first principal components, PC1 and PC2, represented 57.9% of the variance. The PC1 axis, which explained the 32.4% of the variability, was correlated positively with growth parameters, and slightly with chlorophyll a, and the chlorophyll a/chlorophyll b ratio, while it was correlated negatively with polyphenols, flavonoids, antioxidant activity, ascorbic acid, and chlorophyll b. The PC2 axis, which explained the 25.5% of the variability, instead was correlated negatively with chlorophylls a, carotenoids, and total chlorophyll. For the VDI, a clear separation of individuals was observed between samples grown on growing media with compost and samples grown on growing medium without compost. Samples grown on growing media with compost, except for C2-30, however, contrary to the cultivar IMP evidenced higher content of some bio-chemicals, above all polyphenols, flavonoids, ascorbic acid, antioxidant activity, while samples grown on substrates without compost stood out for the higher content of chlorophyll a and the higher chlorophyll a/chlorophyll b ratio. The C2 at dose 30 differed from the other growing media with compost because had higher values of grown components.

These results indicate that cardoon-based compost allows for comparable or even better plant growth than that observed for peat-based growing media and supports the importance of the cultivar factor as a key tool for obtaining healthy food crops [73,88,89].

To this end, according to Santos [88], composting could become an important component of strategies to maximize the levels of bioactive molecules in lettuce.

4. Conclusions

The three composts, derived from as many different admixtures containing cardoon crop residues, did not affect or positively affected lettuce baby leaf growth and production compared to the control. In addition, the specific varietal response observed should be considered to optimize both yield and product quality.

Indeed, it was shown that the effect of compost on plant growth is cultivar-specific and depends on the percentage of cardoon residues in the composting process. The Imperiale lettuce showed the highest leaf dry matter in the presence of Compost 1 (from 50% cardoon), at the peat replacement of 60%, while the Verde d’Inverno in Compost 2 (from 68% cardoon) with a peat replacement of 30%. Also, the content of chlorophylls as well as of antioxidant compounds and antioxidant activity were affected by growing medium differently depending on the lettuce cultivar.

Our overall findings broaden knowledge on the use of compost to replace peat in horticulture, a topic of growing environmental concern, highlighting the suitability of cardoon crop residues-based compost as a constituent of growing media to produce baby leaf lettuce, in agreement with our previous studies with other crops. Along with this, the conversion of waste into a re-evaluated product for innovative use allows a production circuit characterized by a virtuous flow of resources.

In conclusion, our findings suggest that compost from cardoon waste may play a role in sustainable soilless cultivation, in line with the circular economy principles.

Looking forward, on this basis, further research will focus on a detailed physical and chemical characterization of cardoon-based compost in order to deepen the knowledge of its properties and to promote its spread in soilless horticulture.