Antioxidant and Nutrient Profile of Tomato Processing Waste from the Mixture of Indigenous Croatian Varieties: Influence of Drying and Milling

Abstract

Tomato processing waste (TPW) represents a valuable but underutilized by-product of the food industry with potential for valorization within bioeconomy models. This study investigated the chemical composition, antioxidant profile, and sanitary safety of TPW, analyzing the whole TPW; its fractions (peels and seeds) and oil are obtained from TPW seeds. All samples showed contaminant levels within regulatory limits, confirming their safety for further applications. Various drying methods (air-drying at 70 °C and at 50 °C, lyophilization and vacuum drying) and grinding intensities were evaluated to determine their impact on TPW bioactive compounds retention and organoleptic characteristics. TPW exhibited valuable nutritional properties, particularly high protein and dietary fiber content while TPW oil was characterized with high monounsaturated fatty acid content. Results demonstrated that drying method and particle size significantly influenced the yield of bioactive compound and organoleptic properties, with either lyophilization or vacuum drying and finer milling generally enhancing the recovery of polyphenols, β-carotene, and lycopene and improving color intensity. This research provides the first characterization of the TPW obtained from Croatian indigenous tomato varieties, establishing a scientific foundation for its sustainable valorization and, in broader terms, supporting circular economy objectives and contributing to more resource-efficient food systems.

1. Introduction

Bioeconomy is an innovative model of the economy that implies the transition of the production model from linear to circular and is based on the sustainable use of renewable sources, taking into account biodiversity and environmental protection [1]. The transition to bioeconomy and circular economy models is critically urgent for mitigating climate change and biodiversity loss, a necessity now embedded in EU legislation through frameworks which mandates food waste reduction and resource circularity [2,3]. Converting food waste into raw materials is among the pivotal strategies for achieving sustainability goals, enabling resource efficiency and reducing environmental footprints across the food chain. In the EU, food processing waste—specifically from the manufacture of food products and beverages—was estimated at 11 million tons in 2022, representing 19% of total food waste generated that year [4]. One of the widely available and underutilized renewable raw materials is the remaining waste after processing tomato (Solanum lycopersicum L.). Globally, approximately 130 million tons of tomatoes are produced each year—including 19,400,000 tons in the EU—resulting in substantial amounts of tomato processing waste (TPW) [5]. The disposal of TPW has substantial economic and environmental impacts. Due to its poor biological and oxidative stability, it is highly susceptible to microbial degradation, leading to the release of greenhouse gases that contribute substantially to global warming and climate change [6]. Moreover, the high costs associated with waste treatment impose a considerable economic burden on the industry.

Despite representing an economic loss and posing environmental challenges, TPW has significant potential as a resource due to its high content of bioactive substances that could be utilized in the food and pharmaceutical industries [7,8]. Although TPW is still primarily used as animal feed, fertilizer, or simply disposed of in landfills [9], its value as a bioresource is gaining increasing recognition. Recent research highlights TPW as a promising source for the recovery of carotenoids, polyphenols, and dietary fiber [10,11,12]. In addition to these compounds, TPW is rich in other nutrients, including proteins (15.4–23.7%), lipids (5.4–20.5%), and minerals (4.4–6.8%) [13]. As a result, it is also being explored as an alternative source of macronutrients, especially proteins, usually derived from the seed fraction [14].

The chemical composition of TPW is significantly influenced by the tomato variety as well as the soil and agroclimate characteristics of the geographical region of cultivation [15,16,17], which consequently reflects on the chemical composition of TPW [18]. Hence, site-specific compositional analysis is a prerequisite for enabling more precise utilization strategies and optimizing the valorization of TPW. To our knowledge, there are no studies that have yet explored the chemical composition of indigenous Croatian tomato varieties (or remaining TPW) nor the chemical composition of the tomatoes/TPW produced in Croatia. Available research data report on the differences among tomato cultivars, chemical composition of processing by-products, or the importance of growth conditions, but without geographic specificity to Croatia. Therefore, despite the fact that tomato remains a key vegetable crop in Croatia (340,000 t produced annually, and more than 30,000 t processed annually) [5], producing sufficient amounts of TPW, due to a lack of data, it is impossible to accurately estimate its potential for reutilization of TPW within the Croatian context.

For the safe reutilization of agricultural waste in various value-added applications, it is fundamental to ensure its sanitary safety in terms of controlling contaminant and residue levels (pesticides, mycotoxins, heavy metals, etc.) that pose risks to human health, animal health, and the environment if not properly managed. This is particularly concerning by-products intended for use as raw materials for the extraction of bioactive compounds, as even low levels of contaminants may compromise product safety as well as regulatory compliance. To our knowledge, such data are not available for TPW in Croatia.

Fresh TPW typically contains high moisture content, up to 70–80%, depending on the source and processing conditions, making it highly perishable and unsuitable for direct use or storage. Drying is necessary to reduce moisture, prevent microbial spoilage, and stabilize TPW, and it is also crucial for the preservation of heat-labile nutrients (such as lycopene), prevention of color changes, and retention of reconstitution, sensorial, or antioxidative properties. Grinding of plant-based raw materials is another crucial pretreatment step for optimizing TPW utilization. It reduces particle size and disrupts the cell structures, so it can positively influence the extractability of bioactive compounds by facilitating solvent penetration and enzymatic or chemical action. On the other hand, it can also increase oxidation and degradation of unstable compounds [19]. The impact of drying or milling on the extraction yields of bioactive compounds from TPW is still scarce [20,21]. Further research is required to develop tailored pre-processing protocols that optimize the extraction yields and quality of specific categories of bioactive compounds from TPW [10].

This study was designed to provide novel insights into the chemical composition and food safety characteristics of tomato processing waste (TPW) as a whole, as well as its key components—tomato peels and seeds—derived from a blend of eight distinct indigenous Croatian tomato varieties. In addition, the research systematically evaluated how varying drying conditions and milling intensities influence the yield of valuable bioactive compounds and organoleptic properties of TPW. By addressing these critical processing parameters, the investigation offers a unique foundation for optimizing pre-processing techniques, thereby enhancing the efficient and sustainable utilization of TPW-derived bioactive resources.

2. Materials and Methods

2.1. Samples

Ripe tomatoes belonging to 6 indigenous tomato varieties from the Republic of Croatia were collected from experimental fields of the Faculty of Food Technology Osijek, Josip Juraj Strossmayer University of Osijek. Tomato varieties were assigned by accession numbers VEG00356-VEG00362, available at the Croatian Plant Genetic Resources Database [22]. Tomatoes were washed, chopped, and submitted to a hot break process (60 min of boiling) and subsequent sieving to separate the tomato sauce. TPW remaining after juice extraction and sieving consisted of seeds, peel, and parts of the pericarp and placenta of the fruit. Different dried conditions were applied: convective air drying at 70 °C and 50 °C in a laboratory dryer; for 36 and 48 h, respectively (Inko, Zagreb, Croatia); vacuum dryer at 35 °C for 72 h (Fratelli Galli, 21GV, 130 Milan, Italy) and lyophilization for 20 h (Martin Christ, Epsilon 2-4 LSC plus, Osterode am Harz, Germany) Drying times were chosen to achieve the final moisture content of <10%. The 50–70 °C temperature range is commonly applied for air drying plant materials because it represents the balance between drying efficiency, enzyme inactivation, and preservation of bioactive compounds. Inefficient air-drying, due to too low temperatures, might lead to the spoilage of the raw material or enzymatic degradation of bioactive compounds, while exposure to too high drying temperatures can cause thermal degradation of valuable compounds such as polyphenols and carotenoids, particularly lycopene [23]. The grinding process was carried out in a Polymix^® PX-MFC 90 D mill (Kinematica AG, Malters, Switzerland) at 6000 rpm with a 2.00 mm sieve, resulting in a fine powdered material, to conduct physicochemical composition of TPW. In addition, to investigate the impact of grinding degree and particle size on the content of TPW bioactive substances, the dried sample was ground for 5 s, 9 s, 18 s, and 40 s. The grinding duration was selected based on preliminary experiments, which demonstrated that a grinding time of 40 s resulted in sufficiently low Dx 90 values (529 µm). Grinding beyond 40 s led to noticeable heating of the sample. Therefore, 40 s was established as the maximum grinding time to prevent unnecessary heat buildup. Shorter grinding intervals were also tested and selected randomly, guided by preliminary findings indicating that the most significant reduction in particle size occurs within the first 15–20 s of grinding.

As presented in Figure 1, the study was conducted in two phases. In the first phase, TPW was separated into peel (TPW-PEEL) and seed fractions (TPW-SEED), which were then used to assess and compare their suitability as secondary raw materials. This evaluation was based on several criteria: (1) sanitary safety, (2) nutritional composition, (3) bioactive compounds content, and (4) seed oil characteristics. To investigate the effect of higher drying temperatures on specific properties, the entire TPW was also lyophilized (freeze-dried) (TPW-LIO), and the results of these analyses were compared with those from samples dried at 70 °C (TPW-70). For this part of the investigation, we used TPW collected in the season 2022/2023. Given the significant differences observed in the content of certain compounds between TPW-70 and TPW-LIO, the second phase of the study compared the impact of various drying methods and TPW particle size distributions on the content/extractability of the bioactive TPW compounds (total antioxidants, total polyphenols (TPs), particular phenolic compounds, β-carotene, and lycopene). For this part, TPW collected in the season 2023/2024 was used.

2.2. Particle Size Characterization

The particle size, expressed as a volume distribution, was assessed for samples grounded under different conditions and was measured via laser diffraction with a Mastersizer 3000 instrument (Malvern Instruments, Worcestershire, UK), which incorporates a Hydro SV dispersion unit featuring an integrated magnetic stirrer. For this analysis, a small quantity of each sample was suspended in 10 mL of distilled water and introduced into the Hydro SV unit until an obscuration level of 10–20% was achieved. Based on obtained data, the span of the particle size distribution was calculated according to Equation (1):

$$SPAN={\displaystyle \frac{Dx90-Dx10}{Dx50}}$$

(1)

where Dx 10 is the particle diameter below which 10% of the sample’s total volume of particles is found; Dx 50 is median diameter and Dx 90 is the diameter below which 90% of the sample’s total volume of particles is found.

2.3. Determination of TPW Sanitary Safety

For determination of sanitary safety, TPW was analyzed for the content of heavy metals (lead (Pb), cadmium (Cd), arsenic (As)); pesticide residues; mycotoxins (patulin, ochratoxin, aflatoxin B1, the sum of aflatoxins B1, G1, B2, G2); and perchlorate as recommended by the applicable legal regulations that define maximal residue levels (MRLs) in different food categories [24]. Since there were no defined MRLs for residues and contaminants in TPW, the obtained levels were compared with MRLs defined for similar food groups (

Table 1. Contaminant residues in TPW.

Contaminant	Content	MRL *	Technique	Category *
Patulin (µg/kg)	<5.00	25	HPLC	Solid apple products placed on the market for the final consumer, except products listed in 1.3.4 and 1.3.5
Ochratoxin (µg/kg)	<0.01	2.0	LC-MS/MS	Other dried fruits *
Aflatoxin B1 (µg/kg)	<0.01	2.0	LC-MS/MS	Ingredients or processed products from dried fruits, placed on the market for the final consumer or used as an ingredient in food, except products listed in 1.1.3
Aflatoxin B1, G1, B2, G2 (µg/kg)	<0.01	4.0	LC-MS/MS
Pb (mg/kg)	ND	0.05	AAS	Fruiting vegetables, except products listed in 3.1.4.2
Cd (mg/kg)	ND	0.02	AAS	Fruiting vegetables, except products listed in 3.2.4.2
As (mg/kg)	ND	0.15	AAS	Non-parboiled milled rice (polished or white rice)
Perchlorate (mg/kg)	<0.05	0.05	LC-MS/MS	Fruits and vegetables, except products listed in 6.3.1.1 and 6.3.1.2

* Taken from [23]. Levels are not determined for vegetables/tomatoes, so we compared the obtained values with MRLs determined for dried fruits. MRLs: maximal residue levels; HPLC: high-performance liquid chromatography; LC-MS/MS: liquid chromatography–mass spectrometry; AAS: atomic absorption spectrometry.

). Methods of analysis of chosen contaminants and residues were in line with the recommendation of European legislation [25]. Cd was determined by flame atomic absorption spectrometry (FAAS) and Pb and As were determined by total reflection X-ray fluorescence spectrometry (TXRF), as described in detail in 2.5. Pesticide residues in TPW were determined either by UPLC-MS/MS, HPLC-MS/MS, or GC-MS/MS by the method in an accredited laboratory [26], which is in line with maximum residue levels of pesticides in food and feed [27]. Briefly, the samples were extracted with acetonitrile, followed by partitioning with magnesium sulfate and sodium chloride. The extract was then cleaned using dispersive solid-phase extraction (QuEChERS) and analyzed. Other residues were determined by validated in-house HPLC (patulin) or LC-MS-MS method (other analyzed mycotoxins and perchlorates) of the referent laboratory for mycotoxin determination in the Republic of Croatia [28].

2.4. Proximate Composition of TPW

Nutritive composition of TPW was analyzed using established analytical protocols. Ash content was determined gravimetrically by incinerating the sample in a muffle furnace at 550 °C until constant weight, following the AOAC method 942.05 [29]. Moisture was measured gravimetrically by drying the sample at 105 °C to constant weight, as described in AOAC method 934.01 [29]. Protein content was determined using the Buchi Kjehldal system for determination of total nitrogen (Buchi scrubber B-414; Buchi distillation unit B-324, Buchi digestion units K-424; New Castle, DE, USA), and a conversion factor of 6.25 was used to calculate crude protein, in accordance with the AOAC method 2001.11 [29]. Fat content was extracted using Soxhlet apparatus with petroleum ether according to AOAC method 920.39 [29]. The contents of total, soluble, and insoluble dietary fiber were measured using a Total Dietary Fiber Assay Kit (Megazyme K-TDFR-100A, Neogen, Lansing, MI, USA) by enzymatic-gravimetric method following AOAC method 991.43 [29]. Available carbohydrates were calculated by difference, subtracting the sum of moisture, ash, protein, fat, and total dietary fiber from the total sample weight, as recommended by the FAO/WHO [30], commonly applied in food analysis. Finally, energy content was estimated using the Atwater factors (4 kcal/g for protein and carbohydrates, 9 kcal/g for fat, and 2 kcal/g for dietary fiber), as described by Merrill and Watt [31] and adopted in food composition databases. Moisture content of the samples was below 10% and data are not presented in the manuscript but were used for expressing obtained results on a dry matter basis.

2.5. Mineral Composition of TPW

A multielement analysis of TPW and determination of Pb and As (in the framework of investigation of sanitary safety) was performed by Total Reflection X-ray Fluorescence Spectrometry (TXRF). TXRF system (S2 PICOFOX, Bruker AXS Microanalysis GmbH, Berlin, Germany) was equipped with a 40 W X-ray tube with a molybdenum (Mo) anode and a multilayer monochromator (17.5 keV). The characteristic radiation of elements present in the sample is detected by a silicon detector with an active area of 10 mm² and a resolution of 139.43 eV (Mn Kα). Measurements were carried out at 50 kV and 750 μA with a measurement time of 600 s. Determination of Cd content was carried out using an FAAS (Analyst 800 PerkinElmer, Waltham, MA, USA) with deuterium background correction under optimized measurement conditions with suitable hollow cathode lamps and at optimum flame height (air–acetylene). The results were recorded and processed using AAWinlab 32 software (PerkinElmer). Prior to analysis, samples were subjected to microwave-assisted digestion using a microwave closed vessel system (Ethos Easy, Milestone, Sorisole, Italy). For this procedure, approximately 0.5 g of dried TPW was digested with a mixture of concentrated HNO₃ and H₂O₂, following established protocols for sample mineralization and matrix destruction. For the TXRF analysis, 1 g of the mineralized sample solution is transferred into an Eppendorf^® tube, and 0.05 g of the internal standard is added (Ga, 200 mg/kg). The sample was homogenized using a vortex mixer (~15 s). Then, 10 µL of the prepared solution was transferred onto the center of a quartz carrier. The same procedure was repeated for each of the digested samples, after which the carriers were placed on a heating plate at approximately 40 °C. After drying, the samples were introduced into the instrument and analyzed.

2.6. Extraction and Characterization of TPW-SEED Oil

Cold-pressed TPW-SEED oil was obtained by pressing dried seeds (400 g) using a screw press [32]. Insoluble impurities were removed from the crude oil by sedimentation (over 14 days) followed by vacuum filtration. Freshly produced cold-pressed tomato seed oil was evaluated for quality parameters to determine compliance with the Regulation on Edible Oils and Fats [33]. Properties of TPW-SEED oil were analyzed according to available ISO methods for particular analyses: peroxide value (PV) [34]; acid value [35]; iodine value [36]; saponification value [37]; and p-anisidine value (AV) [38]. Totox value was calculated as 2PV + AV. All determinations were carried out in triplicate.

The fatty acids methyl esters (FAME) were prepared with cold methanolic potassium hydroxide solution according to the procedure described in Annex X B of the Commission Regulation No 796/2002 [39]. FAME were afterwards separated by gas chromatograph equipped with a flame ionization detector (Shimadzu GC-2010 Plus, Shimadzu, Kyoto, Japan) and fitted with an SH-FAMEWAXTM capillary column (30 m, 0.32 mm i.d., and 0.25 µm thick stationary phase). Nitrogen was used as carrier gas, flowing at the constant linear velocity of 1.26 mL/min. The split/splitless injector was set at 240 °C, split ratio was 1:100, and the injection volume 2 µL. Initial column temperature of 120 °C was held for 5 min, then gradually increased 5 °C/min until temperature of 220 °C that was held for 20 min. Flame ionization detector temperature was 250 °C. Identification of separated FAME in samples was achieved based on the comparison of retention times with the retention times of certified reference standard (Supelco F.A.M.E. Mix, C4-C24, St. Louis, MO, USA) analyzed under the same conditions. The results were expressed as a percentage of identified fatty acid on total fatty acids (%).

2.7. Antioxidants in TPW

2.7.1. Determination of Antioxidant Activity

Extraction of antioxidants from TPW was conducted by conventional solvent extraction. Conditions of extraction were as follows: 250 mg of the sample was weighed and extracted with 10 mL of 1 M H₂SO₄ for 90 min at 90 °C in closed vials, on a magnetic stirrer with continuous stirring. After extraction, reaction mixtures were filtered and after cooling made up to exactly 10 mL with 1 M H₂SO₄. Clear extracts were used for determination of antioxidant activity and HPLC determination of particular polyphenolic compounds using LC-MS-MS (2.6.2). Optimal sample-to-solvent ratio, temperature, and duration of extraction were identified during preliminary experiments and the solvent (0.1 M H₂SO₄) was chosen based on the available literature data. Extractions were conducted in duplicates, and each extract was analyzed in triplicate (n = 6). Antioxidant activity of TPW was assessed by three methodological approaches—determination of total reducing capacity/total phenols (TPs) by the Folin–Ciocalteu method [40]; TEAC (Trolox Equivalent Antioxidant Capacity) measuring radical scavenging activity against 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonate) radical cation (ABTS·⁺) [41] and Oxygen Radical Antioxidant Capacity (ORAC) [42]. For TP determination, an appropriately diluted TPW extract was mixed with Folin–Ciocalteu reagent and 6% Na₂CO₃ solution in the 96-well plate. Reaction mixtures were incubated for 30 min, and absorbance was recorded at 725 nm. A calibration curve was established using gallic acid standards (ranging from 1 to 150 mgL⁻¹), and TP values were reported as gallic acid equivalents (GAEs). For TEAC the reaction mixture contained adequately diluted extract and ABTS·⁺ solution in the 96-well plate. The microplate was put into the microplate reader immediately after the addition of ABTS·⁺ to TPW extracts. Plate was shaken for 60 s and an end-point absorbance was read at 732 nm after an additional 90 s of incubation at 30 °C (in total 3 min of incubation). The percentage of absorbance reduction was calculated according to Equation (2).

$$\%I={\displaystyle \frac{(Amax-A3min)}{Amax}}\times 100$$

(2)

A calibration curve was constructed using Trolox standards (0–40 gL⁻¹), and antioxidant activity was expressed in Trolox equivalents (TEs). ORAC was assessed by utilizing fluorescein as a fluorescent marker. The assay was conducted in a 96-well black microplate. Briefly, 5 µmolL⁻¹ fluorescein was added to each well, followed by phosphate buffer (75 mmolL⁻¹, pH 7.0; blank), Trolox standard (6.25–100 µmolL⁻¹), or diluted sample. The mixture was pre-incubated at 37 °C for 10 min, after which the oxidation reaction was initiated by adding 25 µL of 150 mmolL⁻¹ 2,2′-Azobis(2-methylpropionamidine) dihydrochloride solution (AAPH). Fluorescence intensity (excitation: 493 nm; emission: 515 nm) was monitored over 60 min. The area under the curve (AUC) was calculated for each sample, and the net AUC was determined by subtracting the blank value. A standard curve was generated using known Trolox concentrations, and sample antioxidant activity was reported as TE. Absorbance and fluorescence measurements were performed using multimodal microplate reader (SpectraMax i3x and SpectraMax MiniMax 300 Imaging Cytometer, Molecural Devices, San Jose, CA, USA).

2.7.2. Chromatographic Determination of Individual Polyphenols

TPW extracts obtained for the analysis of antioxidant activity were also used for the analysis of the specific polyphenols, but after the additional filtration through 0.45 µm PTFE syringe filters. Extracts were analyzed in duplicates using ultra-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS). Analyses were performed using an Agilent 1290 RRLC system (Agilent Technologies, Santa Clara, CA, USA) equipped with a 6430 triple quadrupole mass analyzer and an electrospray ionization source, as described in detail previously [43]. Ionization was carried out in both positive and negative polarity modes, scanning a mass range from 100 to 1000 m/z. High-purity nitrogen (Messer, Zagreb, Croatia; 99.999%) served as the nebulizing and collision gas. Instrument parameters included capillary voltages of 4000 V (positive mode) and 3500 V (negative mode), a drying gas temperature of 300 °C, a gas flow rate of 11 L/h, and a nebulizer pressure of 40 psi. Chromatographic separation was achieved on a Zorbax Eclipse Plus C18 column (Agilent, 100 × 2.1 mm, 1.8 µm particle size) maintained at 35 °C, with an injection volume of 2.5 µL. Data acquisition and analysis were managed using Agilent MassHunter Workstation (version B.04.01). Identification and quantification of polyphenols relied on calibration curves generated using authentic standards, while for compounds lacking commercial standards, tentative identification was based on accurate mass and characteristic fragmentation patterns reported in the literature.

2.7.3. Chromatographic Determination of Carotenoids

Carotenoids from TPW were extracted according to the method described by Dzakovich and co-authors [44], with slight modifications. Briefly, 400 mg of the sample was weighed into the extraction vials and extracted with 10 mL of methanol. After the solvent was added, the samples were vortexed briefly, sonicated for 35 s at 40% power (no pulse mode) using a stepped titanium micro tip (OmniRuptor Ultrasonic Homogenizer, Kennesaw, GA, USA) and centrifuged at 2000× g for 5 min (Heraeus SEPATECH Megafuge 1.0, Heraeus, Thermo Scientific, Waltham, MA, USA). The supernatant was collected, and the extraction was repeated with 10 mL of acetone–hexane mixture (1:1, v/v) containing 0.01% of butylhydroxyanisole (BHA), until the supernatant was colorless. Subsequently, 10 mL of water was added to facilitate the phase separation. The organic layer was collected and filtered through a 0.45 µm PTFE syringe filter into glass vials for quantification of β-carotene and lycopene. Extracts were analyzed in duplicate by reversed-phase high-performance liquid chromatography (RP-HPLC) using an Agilent 1260 Infinity II system (Agilent Technologies, Santa Clara, CA, USA), according to the method described by Hostetler and co-authors [45], with minor modifications. Separation was performed on a C30 column (Phenomenex Develosil, 5 µm, 4.6 × 250 mm, Torrance, CA, USA) maintained at 30 °C. The mobile phases were 20 mM ammonium acetate in methanol–water (98:2, v/v; phase A) and methyl tert-butyl ether (MTBE; phase B). The flow rate was set at 1.0 mL/min, and the injection volume at 30 µL. The gradient was used as follows: from 97% A to 92% A over 1 min, from 92% A to 85% A over 7 min, and from 85% A to 0% A over the next 17 min. The column was held at 0% A for 0.5 min and switched to 97% A over 9.5 min for column reconditioning. Spectral data were acquired from 360 to 550 nm. Carotenoids were detected at 450 nm and quantified using external calibration curves.

2.8. Color Analysis

The color parameters of the tomato powder samples were determined using the CIELAB color space. The color measurements were performed with a Minolta Chroma Meter II Reflectance CR-300 (Minolta Co., Milan, Italy) and the results were expressed as L*, a*, and b* values. In this color space, the L* value stands for lightness (from black to white), a* for the red–green component (positive values stand for redness, negative values for greenness), and b* for the yellow–blue component (positive values stand for yellowing, negative values for bluing). The samples were evenly distributed in a Petri dish to ensure uniform surface coverage during the measurement. Each sample was measured in triplicate and the results were expressed as means ± standard deviations. Browning index (BI) was determined to investigate the potential accumulation of brown pigments that might be formed during drying at higher temperatures due to enzymatic or non-enzymatic reactions. The index was calculated as the ratio of absorbance at specific wavelengths (ABS_520nm/ABS_420nm), which reflects the balance between anthocyanin content and brown pigmentation.

2.9. Statistical Analysis

Statistical analysis and plotting were performed using Microsoft Excell (Washington, WA, USA) and GraphPad Prism 10.4.1 (Boston, MA, USA) software. Analyses were performed in duplicates, triplicates, or quadruplicates depending on the type of analysis and presented as means ± standard deviation. The differences between groups were tested using Student’s t-test or one-way ANOVA followed by Tukey’s post hoc multiple comparison test, depending on the number of groups. A p < 0.05 was considered statistically significant.

3. Results and Discussion

3.1. Particle Size Distribution

TPW samples ground under different conditions resulted in significantly different particle size distribution (Figure 2). Prolonged grinding (40 s of grinding) resulted in a significant decrease in the median diameter from 732 μm (no grinding) to 210 μm but did not affect the span which ranged from 2.34 to 2.47 in all ground samples.

3.2. Sanitary Safety of TPW

Although raw materials used in food production are required to comply with established MRLs for contaminants, it remains essential to monitor food processing residues for contaminant levels as well.

In this aspect, TPW was analyzed for various contaminants—mycotoxins (patulin, ochratoxin, aflatoxins); heavy metals (Pb, Cd, As); and perchlorate and obtained values were compared with MRLs set by Commission regulation [24] for different food categories, including fruits and vegetables, which are directly relevant to tomato-derived materials (Table 1). The amount of pesticide residues is presented in Table S1 and the obtained values were compared with MRLs for tomatoes set by the European Pesticide Database [46].

The investigation of contaminant residues in TPW is crucial due to the high prevalence and persistence of pesticide residues in tomatoes and their derivatives. Despite industrial processing steps like washing, peeling, and thermal treatment, some pesticide residues remain detectable in processed tomato products such as extracts, sauces, and ketchups, albeit often below regulatory MRLs [47]. Studies have demonstrated that worldwide, a significant proportion of tomato samples contain multiple pesticide residues, with over 60% of samples exceeding the current European Union MRLs for pesticides such as acetamiprid, chlorpyrifos, lambda-cyhalothrin, and DDT [48]. In tomatoes grown in the EU, the situation is significantly better; according to the 2022 EU report on pesticide residues in food, approximately 1.6% of tomato samples analyzed under the EU-coordinated multiannual control program (EU MACP) exceeded the MRLs, with 0.9% being non-compliant after considering measurement uncertainty [49]. The European Food Safety Agency (EFSA) particularly highlights that chlorates, which can be found in tomatoes, have been detected at levels exceeding MRLs in leafy crops and tomatoes, warranting continued surveillance. The results of our analysis show that the residues of all monitored pesticides were below the current MRLs, including the residues of perchlorates (Table S1 and Table 1). The occurrence of heavy metals and pesticides in tomatoes and their by-products is less frequently studied. However, metals and certain mycotoxins are resilient to common industrial processing methods like heating and pasteurization, leading to their persistence in tomato products and potentially in processing wastes. As presented in Table 1, the levels of mentioned contaminants were below MRLs in the analyzed TPW, indicating its suitability for further valorization.

3.3. Proximate Composition of TPW, TPW-PEEL, and TPW-SEED

Numerous studies have shown that TPW is exceptionally rich in dietary fiber, followed by carbohydrates, proteins, lipids, and minerals, making it a suitable source of fiber, proteins, and lipids [50,51,52]. The proximate composition of individual fractions of indigenous Croatian tomato pomace varieties and the effect of different drying methods on their composition is shown in

Table 2. Proximate composition and energy content of the TPW, TPW-PEEL, and TPW-SEED.

g/100 g Dry Matter	TPW-70	TPW-LIO	TPW-PEEL	TPW-SEED
ash	4.81 ± 0.06 c	4.16 ± 0.04 b	5.09 ± 0.08 d	3.08 ± 0.11 a
fat	7.84 ± 0.02 b	9.21 ± 0.37 c	3.89 ± 0.20 a	25.87 ± 0.18 d
protein	18.00 ± 0.93 b	17.46 ± 0.02 b	14.34 ± 0.29 a	27.12 ± 0.11 c
dietary fiber
total	40.10 ± 1.14 a	41.16 ± 1.37 a	52.06 ± 1.32 b	41.30 ± 1.89 a
soluble	7.53 ± 0.81 a	7.57 ± 0.88 a	9.09 ± 1.46 a	5.40 ± 2.20 a
insoluble	32.57 ± 0.44 a	33.59 ± 0.50 b	42.97 ± 0.17 d	35.90 ± 0.31 c
available carbohydrates	29.25 ± 2.04 c,d	28.01 ± 1.66 b,c	24.62 ± 1.67 b	2.62 ± 0.00 a
energy (kcal/100 g)	339.74 b	347.09 c	294.99 a	434.44 d

The data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Letters next to value indicate statistically significant differences between groups for each element. TPW-70: tomato processing waste air dried at 70 °C; TPW-LIO: tomato processing waste dried by lyophilization; TPW-PEEL: peel fraction of tomato processing waste dried air dried at 70 °C; TPW-SEED: seed fraction of tomato processing waste air dried at 70 °C.

.

In this context, the goal of our research was to determine the macronutrient composition of individual fractions of tomato pomace, as well as to examine the effect of different drying methods on the composition, especially since the study focused on a mixture of indigenous Croatian tomato varieties for which no nutritional data is currently available. The analysis results presented in Table 2 showed that TPW contains the highest proportion of dietary fiber, followed by available carbohydrates (40% and 29%, most abundant in the peel); proteins and lipids (18% and 7.8%, most abundant in the seeds); and minerals (4.8%). Previous studies have shown that lipid content can vary from 2 to 16.24%, depending on the variety, ripeness, climate conditions, and extraction method [53]. Given that 7.8% lipids were detected in our samples, we can conclude that indigenous Croatian varieties have medium values compared to those recorded in the literature. The soluble fiber fraction, with a particular emphasis on pectin, which is important for the pharmaceutical industry, makes up to 7.5% of the total composition. The literature reports that pectin content can reach up to 15% of dry matter [53], indicating that our samples contain approximately half the amount of maximum pectin concentration in TP. A significantly higher presence of insoluble compared to soluble fiber was also observed (ratio 5.3–7.6:1, depending on the fraction). The peel contained the highest proportion of total (52%) and insoluble fiber (43%), while the seeds had the highest energy value (424 kcal/100 g), consistent with their high lipid content.

The influence of drying method on the nutrient composition of TPW was investigated by comparing TPW-70 and TPW-LIO samples. The method of drying clearly affected the content of specific components.

Ash content is an important parameter in quality control of plant material, as excessive ash may indicate the presence of impurities or contaminants, potentially pointing to poor quality or harmful substances. For example, increased ash content may suggest soil contamination used in cultivation, such as heavy metals or other pollutants. In our study, determining ash content was crucial to confirm the quality of the prepared TPW samples and to accurately calculate total carbohydrates. The ash content was higher in the TPW-70 (4.8% ± 0.1%) compared to TPW-LIO (4.2% ± 0.0%), consistent with previous findings [52]. However, although statistically significant, the increase in ash content is negligible and likely the result of intrinsic variation in the composition of the plant material, as previously noted by Lemus-Mondaca and co-authors [54]. Determining ash content was crucial to confirm the quality of the prepared TPW samples, as excessive ash may indicate the presence of impurities or contaminants, potentially pointing to poor quality or harmful substances. For example, increased ash content may suggest soil contamination used in cultivation, such as heavy metals or other pollutants [55]. Lipid content was significantly higher in TPW-LIO (9.2% ± 0.4%) compared to TPW-70 (7.8% ± 0.0%). This result may be attributed to the greater porosity of the freeze-dried material, which allows for more efficient extraction of lipids but also highly valuable lipophilic micronutrients in TPW such as lycopene, beta-carotene, lutein, phytoene, and phytofluene [56]. Protein content did not differ significantly between the two drying methods (TPW-70: 18% ± 0.9%, TPW-LIO: 17% ± 0.0%), which is expected given the analysis method used (Kjeldahl). These values are consistent with previous studies that report a range from 15.08 to 24.67% [50]. Dietary fiber content was also not statistically significantly different between samples dried using different methods. The content of insoluble fiber was 33% ± 0.4% for TPW-70 and 34% ± 0.5% for TPW-LIO, while the content of soluble fiber was 7.5% ± 0.8% and 7.6% ± 0.9%, respectively. These values are consistent with the literature for insoluble fiber, while the values for soluble fiber are somewhat higher [57]. Carbohydrate content was 23% ± 2.0% for TPW-70 and 20% ± 1.7% for TPW-LIO, with no statistically significant difference, aligning with the literature data [58].

3.4. Mineral Composition of TPW, TPW-PEEL, and TPW-SEED

The mineral composition of TPW is presented in

Table 3. Mineral composition of TPW-70, TPW-LIO, TPW-PEEL, and TPW-SEED.

Element (mg/kg)	TPW-70	TPW-LIO	TPW-PEEL	TPW-SEED
Cr	1.24 ± 0.65 a	1.06 ± 0.08 a	1.17 ± 0.75 a	0.54 ± 0.01 a
Mn	15.3 ± 2.06 b	18.17 ± 1.38 b	25.97 ± 4.01 c	1.0 ± 0.0 a
Ni	2.03 ± 1.01 a	1.66 ± 0.22 a	1.42 ± 0.39 a	2.15 ± 0.23 a
Br	3.07 ± 0.27 b	3.23 ± 0.21 b	5.09 ± 0.35 c	2.11 ± 0.18 a
Rb	3.73 ± 0.09 b	3.56 ± 0.09 b	8.98 ± 0.83 c	0.24 ± 0.01 a
Sr	2.29 ± 0.09 b	2.85 ± 0.32 b	4.85 ± 0.92 c	0.0 ± 0.0 a
Fe	117.5 ± 34.65 b	131.36 ± 0.68 b	202.91 ± 12.57 c	21.6 ± 0.7 a
K (g/kg)	10.79 ± 0.73 b	11.77 ± 3.36 b	19.21 ± 1.28 c	0.92 ± 0.6 a

The data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Letters next to value indicate statistically significant differences between groups for each element. TPW-70: tomato processing waste air dried at 70 °C; TPW-LIO: tomato processing waste dried by lyophilization; TPW-PEEL: peel fraction of tomato processing waste dried air dried at 70 °C; TPW-SEED: seed fraction of tomato processing waste air dried at 70 °C.

. It is evident that different fractions of the material have distinct mineral profiles. TPW-PEEL had the highest contents of Fe, Mn, and K. The peel was especially rich in Fe and Mn, while TPW-SEED contained lower amounts of Fe, Mn, as well as Br, Rb, Se, and K. Previous research reports variable mineral composition of TPW [56,59], which is expected, as it depends on genetic variation, fertilization, irrigation, cultivation method, and ripening stage among other factors [60,61,62,63,64]. To the authors knowledge there is no other research concerning the mineral content of indigenous Croatian tomato varieties.

Since TPW is relatively rich in minerals, there have been efforts to use it in enrichment of food products. For example, Mehta and co-workers [65] incorporated TPW in the production of muffins and bread which resulted in higher content of K, Mg, Ca, and Fe. They also observed lower sodium content, which might prove as a valuable outcome considering the excessive consumption of sodium (table salt) in many parts of the world. The products also had a softer texture and improved shelf life compared to those without the addition of TPW. Isik and Topkaya [66] prepared crackers containing tomato pomace that had significantly higher content of Mg, Ca, K, P, Mn, Zn, and Fe compared to controls. The respondents reported certain bitterness in the taste; however, they did not report significant decrease in sensory attractiveness. An interesting study was performed by Isik and Yapar [67], who made tarhana (fermented wheat/yogurt food product) comprising seed from tomato by-products to improve its characteristics. They reported the increase in the content of all the analyzed minerals in the study—Mg, CA, K, P, Mn, Zn, Fe, and Cu. Tarhana containing 15% tomato seeds also received the highest like scores by the respondents in sensory analysis. There are also examples of adding TPW to animal feed to improve the mineral content of meat [68].

3.5. Total Polyphenols, Carotenoids, and Antioxidant Capacity of TPW, TPW-PEEL, and TPW-SEED

TPW has been recognized as a valuable but highly underutilized source of antioxidants, polyphenols and carotenoids, whose content highly depends on the tomato variety, origin, and the type of processing [10,11]. Research on carotenoids is considerably more extensive than that on polyphenols, and, to date, the antioxidant profile and content of antioxidants across various fractions of TPW derived from Croatian indigenous varieties remains unexplored. Antioxidant activity and the content of TP, β-carotene, and lycopene in TPW is presented in Figure 3. Antiradical activity was assessed by two approaches; even though ORAC is considered more biologically relevant, it focuses more on hydrogen atom transfer antioxidants, so TEAC was also performed to take into account electron transfer reactions [69]. TP, reflecting total reducing power, was significantly higher in TPW-PEEL compared to TPW-SEED (213.5 ± 1.5 and 69.3 ± 0.1 mg/100 g GAE, respectively), which is consistent with the available literature data (Figure 3a). Sarkar and Kaul [70] showed that the TP content of tomato peel was 66.5% higher compared to seeds and similar observations were confirmed by other authors investigating the distribution of phenolic compounds in ripe tomato [71]. A similar effect of TPW fractionation was observed in the antioxidant activity of TPW-PEEL and TPW-SEED measured by both ORAC (111.4 ± 9.0 and 52.4 ± 10.8 g/kg TE, respectively) and TEAC assays (55.2 ± 0.6 and 25.0 ± 0.2 mg/100 g TE, respectively). This outcome was anticipated, as previous studies have demonstrated a strong correlation between antioxidant capacity (e.g., TEAC) and polyphenol content, indicating that polyphenols are the primary contributors to the hydrophilic antioxidant activity in tomatoes [72]. Although vitamin C also plays a role in the antioxidant activity of tomatoes, its content in TPW obtained after subjecting ripe tomatoes to thermal processing at 90 °C is expected to be significantly reduced, minimizing its contribution in this context. Carotenoids also exhibit strong antioxidant activity that can be measured by the ORAC assay, offering detailed insight into radical chain-breaking potential. However, their contribution to observed ORAC levels in this case is minimal because the detection of lipid-soluble antioxidants by ORAC requires an extraction protocol specifically designed for lipophilic compounds. This was not the case in our study because applied extraction protocols for antioxidants primarily targeted hydrophilic substances; therefore, the observed ORAC values predominantly reflect the polyphenol content rather than carotenoids.

TPW-70 contained somewhat higher levels of total polyphenols compared to TPW-LIO (221.4 ± 7.0 vs. 194.4 ± 8.7 mg/100 g GAE, respectively), which reflected on ORAC values (139.7 ± 29.1 vs. 95.4 ± 2.2 g/kg TE, respectively). This is consistent with available investigations showing that exposure of tomatoes to heat treatment significantly increases the content of total phenols, more specifically caffeic and p-coumaric acid, whereas the content of ferulic acid slightly decreases. Also, chlorogenic acid is not detected in fresh tomatoes, while it was the predominant phenolic acid in TPW [73].

Figure 3b shows the content of major carotenoids in TPW-lycopene and β-carotene. To the best of our knowledge, this is the first study to report carotenoid content in TPW derived from indigenous Croatian tomato varieties. The values we obtained fall within the ranges reported in the literature; however, they should be interpreted with caution. Namely, even though different genotypes exhibit varying levels and profiles of carotenoids, different environmental and agronomic factors often modulate carotenoid accumulation to an even greater extent [74]. As expected, the amounts of lycopene were significantly higher compared to β-carotene, and the pigment was predominantly found in TPW-PEEL, while the amounts in TPW-SEED were significantly lower (214.5 ± 0.9 mg/100 g and 51.6 ± 0.2 mg/100 g, respectively). Similar differences, even though less pronounced, were obtained for β-carotene content that ranged from 10.3 ± 0.1 mg/100 g in TPW-SEED to 18.7 ± 0.2 in TPW-PEEL. Our results are consistent with the recent review [52] reporting that tomato peels are rich in lycopene, accounting for 80–90% of total carotenoids, with lycopene content ranging from 50 to 1930 mg/100 g whereas tomato seeds contain much lower lycopene levels, around 22–37 mg/100 g. As seen from the presented data, type of TPW drying (air drying at 70 °C or lyophilization) did not have significant effect on the amounts of β-carotene (12.8 ± 0.3 and 12.8 ± 0.1 mg/100 g, respectively) but it significantly affected lycopene content (79.4 ± 0.8 and 99.9 ± 2.5 mg/100 g, respectively) suggesting more pronounced thermal instability of lycopene. This is consistent with the available literature data showing that during thermal processing of tomato pastes, lycopene content decreases with extended heating times, while β-carotene, though present in lower amounts, remains relatively more stable under the same conditions [75].

3.6. Characterization of TPW-SEED Oil

As mentioned previously, TRP-SEED oil was obtained by cold pressing of dried TPW seeds followed by sedimentation and removal of crude oil by vacuum filtration. This is the method where extraction is conducted at low temperatures (typically below 27 °C), without the use of heat or chemical solvents. Yields obtained in our work were 17.4% ± 0.7% which is comparable with the available literature data related to yields obtained by cold pressing (

Table 4. Yield and physical–chemical composition of TPW-SEED oil.

	TPW-SEED Oil	Literature Data	EU Regulation
Yield (%)	17.4 ± 0.7	17–18 [76]
Density (g/cm3)	0.92 ± 0.01	0.81–0.89 [32]
Peroxide value (mmol O2/kg)	0.00 ± 0.00	2.5–2.6 [32]	<20.0 [77]
p-anisidine value	0.62 ± 0.02		<10.0 [78]
Total oxidation value	0.62 ± 0.02
Free fatty acid (%)	0.44 ± 0.01	1.4–1.5 [32]	<0.80 [77]
Saponification number (g KOH/kg)	204.3 ± 5.1	192–193 [32]
Iodine number (g I2/kg)	118.3 ± 2.1	121–131 [32]

The data are presented as mean ± SD. TPW-SEED: seed fraction of tomato processing waste air dried at 70 °C.

), and a little lower compared to yields obtained by applying higher temperatures and organic solvents (yields 21–23%) [76].

The key parameters for assessing the quality of TPW-SEED oil include the iodine number (which measures the degree of unsaturation of fatty acids and reflects oil stability and drying properties); the saponification number (that gives information on the fatty acid chain length); the peroxide value (indicating the extent of primary oxidation and freshness by quantifying peroxides); the p-anisidine value (that gives information on secondary oxidation products, such as aldehydes, that affect flavor and shelf life); and the total oxidation number (which is a combined indicator of overall oxidative rancidity calculated from peroxide and p-anisidine values). Additionally, the free fatty acid (FFA) content represents the amount of hydrolytic degradation, impacting taste and shelf life, while the iodine number is also used to characterize the oil’s unsaturation profile. Obtained data, presented in Table 4, are compared to the available literature data as reviewed by Sangeetha and co-authors [32] and with the current legal regulations. In the EU the quality of tomato seed oil and other edible oils is regulated by specific physical–chemical parameters to ensure safety, authenticity, and nutritional value [79]. The analytical results demonstrate that the oil was of excellent quality: the peroxide value (PV) was 0 mmol O₂/kg, indicating no oxidative deterioration of the oil during storage of the dried seeds. As peroxide value measures the concentration of peroxides and hydroperoxides formed during the initial stages of fat oxidation, this is a strong indicator of high quality and proper storage, Low p-anisidine value indicates minimal presence of secondary oxidation products (mainly aldehydes), which means the oil has not developed off-flavors or odors and maintains good sensory qualities. The oil acidity, expressed as free fatty acid content, was 0.44%. All the characteristics mentioned comply with the current legal regulations and are significantly below permissible limits and lower compared to the available literature data indicating very high quality. Density (0.92 g/cm³) is typical for vegetable oils and indicates a light oil, which is easy to handle and blend in both food and cosmetic formulations. A saponification number (204 mg KOH/g) is considered moderately high. It fits the typical range of values characteristic for common edible oils and is consistent with available data for tomato seeds oil.

Fatty acid compositions are predominantly polyunsaturated and monounsaturated fatty acids, with 52.3% of omega-6 (linoleic acid) followed by 25.8% of omega-9 (oleic acid) as the major components (Figure 4). Saturated fatty acids such as palmitic acid (13.4%) and stearic acid (5.7%) are present in moderate amounts, while other fatty acids like myristic (0.12%), palmitoleic (0.29%), linolenic (2.16%), arachidic (0.19%), and gadoleic acid (0.12%) occur in smaller quantities. This is comparable to other frequently consumed edible oils such as sunflower or corn oil. Comparison of the fatty acid composition of TPW-SEED oil with the other available literature data is presented in Table S2. It is obvious from the presented data that the obtained values are consistent with the data obtained by other authors [32].

3.7. Impact of Drying Method and Grinding Degree on the Content of Total Antioxidants, Polyphenols, β-Carotene, and Lycopene

As mentioned previously, TPW offers considerable promise as a rich source of carotenoids and polyphenols, which are highly valued in the pharmaceutical and cosmetic industries. However, more research is necessary to expand its industrial use, particularly in terms of scaling extraction methods from laboratory settings to full-scale production and investigating more thoroughly how processing and pretreatment of the raw material affect the quantity and characteristics of target compounds [10]. The results from the initial phase of the study demonstrated that drying conditions have a significant impact on the levels of bioactive compounds in TPW, particularly lycopene, as well as other antioxidants. These changes consequently affect the overall antioxidant capacity of the samples. Building on these findings, the second phase of the investigation expanded to include a broader range of drying conditions and a more comprehensive analysis of the polyphenolic content.

As presented in

Table 5. Impact of drying on the content of polyphenols, carotenoids, and the antioxidant activity of TPW.

Compound	TPW-LIO	TPW-VAC	TPW-50	TPW-70
β-carotene (mg/100 g)	12.8 ± 0.1 b	14.1 ± 0.2 c	11.9 ± 0.3 a	12.8 ± 0.3 b
Lycopene (mg/100 g)	99.9 ± 0.0 b	164.8 ± 3.5 c	83.1 ± 3.1 a	79.4 ± 0.8 a
TP (mg GAE/100 g)	194.4 ± 8.7 a,b	204.9 ± 10.7 b,c	181.2 ± 8.6 a	221.4 ± 7.0 c
TEAC (mg TE/100 g)	72.8 ± 9.6 b	58.5 ± 6.6 a,b	51.8 ± 2.4 a	63.4 ± 8.4 a,b
Rutin (mg/kg)	27.3 ± 0.9 b	15.1 ± 6.6 a	13.7 ± 2.5 a	16.7 ± 2.2 a
Kaempferol-3-rutinoside (mg/kg)	2.4 ± 0.2 a	1.9 ± 0.0 a	2.4 ± 0.5 a	0.9 ± 0.5 a
Catechin (mg/kg)	15.2 ± 1.4 b	7.4 ± 9.4 a,b	5.0 ± 4.4 a,b	2.4 ± 0.2 a
Chlorogenic acid (mg/kg)	11.2 ± 0.2 c	10.7 ± 1.5 b,c	9.3 ± 0.7 b	6.7 ± 0.2 a
Caffeic acid (mg/kg)	12.3 ± 2.6 b	7.2 ± 0.2 a	6.5 ± 0.3 a	6.5 ± 0.8 a

The data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Letters next to values indicate statistically significant differences between groups for each element (within the row). TPW-70: tomato processing waste air dried at 70 °C; TPW-LIO: tomato processing waste dried by lyophilization; TPW-PEEL: peel fraction of tomato processing waste air dried at 70 °C; TPW-SEED: seed fraction of tomato processing waste air dried at 70 °C; TP: total phenols; TEAC: Trolox equivalent antioxidant capacity.

, vacuum drying preserves the highest amounts of carotenoids, and the impact is particularly emphasized in the case of lycopene. While β-carotene levels are affected moderately (with values ranging from 11.9 ± 0.3 mg/100 g up to 14.1 ± 0.2 mg/100 g), lycopene content varied from 79.4 ± 0.8 mg/100 g in TPW-70 up to 164.8 ± 3.5 mg/100 g, confirming again the sensitivity of lycopene to thermal degradation, primarily heating but also freezing during the process of lyophilization. Compared to conventional air drying, both lyophilization and vacuum drying provide protection from heat-induced degradation of thermally sensitive compounds. Additionally, lyophilization leads to ice crystal formation that disrupts cellular structures, which might have positive effect on the subsequentional extraction of carotenoids and other bioactive compounds. However, during the freeze-drying process, a temperature of −80 °C to −20 °C is usually maintained which might lead to the loss of particularly thermally sensitive compounds such as lycopene (as consistent with our results) [80]. Vacuum drying has been proven to be the method of choice for preservation of lycopene, which is consistent with the available literature data focused on optimization of drying of lycopene-rich carrots [81]. This is probably due to the fact that vacuum drying is performed at sub-atmospheric pressure, which lowers the boiling point of water and enables drying at reduced temperatures. Additionally, the low-oxygen environment minimizes oxidation, making vacuum drying ideal for heat- and oxidation-sensitive raw materials [82].

Interestingly, the TP content was the highest in the TPW-VAC and TPW-70 samples, measuring 204.9 ± 10.7 and 221.4 ± 7.0 mg/100 g GAE, respectively. This highlights the complexity and heterogeneity of polyphenolic compounds in tomatoes. This phenomenon can be attributed to the thermal sensitivity of certain polyphenols, which may be completely degraded under thermal stress [83,84]. On the other hand, exposure to higher temperatures can break down complex polyphenolic compounds into smaller molecules that remain detectable by the total polyphenol assay which can lead to an apparent increase in the total phenolic content [85]. Furthermore, heat treatment can enhance the extractability of phenolic compounds, contributing to the increased TP in thermally treated samples and inhibit polyphenol oxidase that degrades polyphenols [86,87]. This explains the high TP values obtained in TPW-70 and is consistent with a similar investigation that revealed that increasing oven drying temperature from 40 °C to 60 °C significantly increased polyphenol content in dried tomatoes [88].

Rutin, kaempferol-3-rutinoside, catechin, chlorogenic acid, and caffeic acid were determined as major phenolic compounds in TPW, which is mostly consistent with the available literature data. Fuentes and coauthors [73] determined chlorogenic acid, caffeic acid and p-coumaric acid as major phenolic constituents of tomato pomace. Abbasi-Parizad and co-workers [89] determined gallic acid, chlorogenic acid, cinnamic acid, naringenin chalcone, quercetin, and kaempferol as the major polyphenolic compounds in TPW. Observed differences can be explained by the differences in numerous factors that can affect the phenolic composition of TPW: tomato sort, agroclimatic conditions, type of TPW processing, and type of extraction. The highest amounts of particular polyphenolic compounds were found in lyophilized samples and were degraded significantly when submitted to higher drying temperatures.

The results demonstrate that the choice of drying technique markedly affects the chemical composition of the dried raw material and should thus be aligned with the intended application. For TPW, conventional air drying at 70 °C effectively maintains high polyphenol content, strong antioxidant activity, and satisfactory β-carotene levels. However, if lycopene preservation is a priority, vacuum drying is the preferred method, while retention of thermally labile polyphenolic compounds is best achieved through lyophilization. At the industrial scale, the selection of a drying method should be guided by an objective assessment of the benefits and total costs associated with each process.

Grinding is another aspect of TPW processing that can influence the content of polyphenols and carotenoids in the extracts obtained by affecting the particle size, surface area, and cell wall disruption, which in turn impacts the efficiency of solvent penetration and compound release [90]. Grinding facilitates disruption of cell walls and membranes, and reduces sample particle size, consequently increasing extraction yields. However, excessive grinding or very fine particle sizes may lead to oxidation and degradation of sensitive compounds like polyphenols due to increased exposure to oxygen and enzymes, so an optimal particle size is important [91]. To our knowledge, the effect of grinding on the extractability of bioactive compounds from tomato pomace has not been investigated yet.

Table 6. Impact of grinding degrees on the content of polyphenols, carotenoids, and the antioxidant activity of TPW.

Compound Dx 50	TPW-70-M0 732 µm	TPW-70-M5 542 µm	TPW-70-M9 431 µm	TPW-70-M18 388 µm	TPW-70-M40 210 µm
β-Carotene (mg/100 g)	11.1 ± 0.6 a	11.3 ± 0.4 a	11.9 ± 0.4 a	12.8 ± 0.3 b	11.9 ± 0.1 a
Lycopene (mg/100 g)	75.9 ± 2.5 a,b	72.5 ± 0.5 a	73.2 ± 3.3 a	79.4 ± 0.8 b	74.1 ± 0.3 a
TP (mg GAE/100 g)	174.9 ± 2.8 a	211.8 ± 12.8 b	209.9 ± 0.8 b	221.4 ± 7.0 b	230.2 ± 17.7 b
TEAC (mg TE/100 g)	58.4 ± 5.3 a	54.2 ± 0.7 a	49.8 ± 2.6 a	63.4 ± 8.4 a	61.7 ± 9.7 a
Rutin (mg/kg)	14.7 ± 2.5 a	15.0 ± 0.2 a	13.8 ± 0.8 a	18.2 ± 0.1 b	18.1 ± 1.3 b
Kaempferol-3-rutinoside (mg/kg)	0.81 ± 0.41 a	0.81 ± 0.34 a	1.26 ± 0.29 a	1.36 ± 0.20 a	1.06 ± 0.13 a
Catechin (mg/kg)	6.46 ± 0.39 d	3.50 ± 0.67 c	3.83 ± 0.12 c	2.41 ± 0.16 b	0.91 ± 0.28 a
Chlorogenic acid (mg/kg)	8.85 ± 1.15 b	7.17 ± 0.47 a	6.63 ± 0.44 a	6.67 ± 0.21 a	7.30 ± 0.03 a
Caffeic acid (mg/kg)	7.03 ± 0.84 a	7.49 ± 0.44 a	8.83 ± 0.23 b	7.96 ± 0.04 a,b	7.22 ± 0.33 a

The data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Letters next to values indicate statistically significant differences between groups for each element (within the same row). TPW-70-Mx: tomato processing waste air dried at 70 °C and milled for x sec; Dx 50: median diameter of particles.

shows the results of investigating the influence of TPW median particle size on the extractability of TP, total antioxidants, carotenoids, and particular polyphenolic compounds. Their yields were significantly influenced by the degree of milling.

The highest β-carotene and lycopene content was obtained in samples with a median diameter of 388 µm, obtained after 18 s of grinding under predefined conditions. Prolonged milling (40 s) and reduction in median diameter decreased the yields of both carotenoids, probably causing degradation. Distinct trends were observed concerning the extraction of total phenolics (TPs) and total antioxidant capacity (TEAC). Correlation analysis revealed a significant negative correlation between median particle diameter (Dx 50) and total phenolic content (r = −0.9386, p = 0.0181), indicating that a reduction in particle size enhances the extraction of phenolic compounds. In contrast, TEAC values were maximized at Dx 50 = 388 µm, yet further decreases in particle size through prolonged grinding did not adversely affect antioxidant yields. This suggests that the compounds responsible for antioxidant activity, as measured by TEAC, possess greater stability against oxidative or thermal degradation than carotenoids. However, chromatographic analysis indicated that specific polyphenols can be susceptible to adverse effects from extended grinding. In particular, a positive and statistically significant correlation was found between Dx 50 and catechin content (r = 0.9629, p = 0.0085), implying that excessive reduction in particle size leads to catechin loss, likely due to its heightened sensitivity to oxidation [92,93].

3.8. Impact of Drying Method and Grinding Degree on Color Characteristics

Instrumental color measurements of tomato powder samples, expressed in CIELAB coordinates, as well as the values of BI, showed remarkable differences in color attributes depending on the drying method and the degree of grinding (

Table 7. Impact of drying method and grinding degree on color parameters of TPW.

Sample	L*	a*	b*	C*	h°	BI
	Impact of drying technique
TPW-LIO	56.0 ± 0.1 b,c	28.2 ± 0.0 c	52.1 ± 0.1 d	59.2 ± 0.1 b	61.6 ± 0.0 d	0.76 ± 0.00 a
TPW-VAC	48.6 ± 0.0 a	31.2 ± 0.0 d	50.6 ± 0.1 c	59.4 ± 0.1 b	58.3 ± 0.1 a	0.74 ± 0.01 a
TPW-50	57.8 ± 1.8 c	25.4 ± 0.1 a	45.4 ± 0.1 b	52.0 ± 0.2 a	60.8 ± 0.0 c	0.71 ± 0.06 a
TPW-70	55.4 ± 0.0 b	26.0 ± 0.1 b	44.8 ± 0.2 a	51.8 ± 0.2 a	59.8 ± 0.0 b	0.73 ± 0.01 a
	Impact of grinding degree
TPW-70-M0	49.3 ± 0.1 a	21.9 ± 0.2 a	32.5 ± 0.1 a	39.2 ± 0.2 a	56.0 ± 0.1 a	0.56 ± 0.00 a
TPW-70-M5	54.1 ± 0.4 c	26.4 ± 0.2 c	45.2 ± 0.1 c,d	52.3 ± 0.3 b	59.7 ± 0.1 c	0.71 ± 0.02 b
TPW-70-M9	52.6 ± 0.1 b	28.0 ± 0.0 e	45.5 ± 0.2 d	53.4 ± 0.2 c	58.4 ± 0.1 b	0.73 ± 0.01 b
TPW-70-M18	55.4 ± 0.0 d	26.0 ± 0.1 d	44.8 ± 0.2 b,c	51.8 ± 0.2 b	59.8 ± 0.0 c	0.73 ± 0.01 b
TPW-70-M40	61.2 ± 0.0 e	23.5 ± 0.0 b	49.0 ± 0.2 e	54.4 ± 0.1 d	64.4 ± 0.0 d	0.72 ± 0.02 b

The data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Letters next to values indicate statistically significant differences between groups for each element (within the same column). TPW-70: tomato processing waste air dried at 70 °C; TPW-LIO: tomato processing waste dried by lyophilization; TPW-PEEL: peel fraction of tomato processing waste air dried at 70 °C; TPW-SEED: seed fraction of tomato processing waste air dried at 70 °C; TPW-70-Mx: tomato processing waste air dried at 70 °C and milled for x sec; L*: lightness value; a*: green/red axis value; b*: yellow/blue axis value; C*: chroma (overall color saturation); h°: hue angle; BI: browning index.

).

The lightness values (L*) ranged from 48.6 ± 0.0 in sample TPW-VAC to 61.2 ± 0.0 in TPW-70-M40. Higher L* values indicate a lighter appearance, while lower values reflect darker color shades, probably due to pigment degradation or surface changes during processing. Vacuum drying produced samples with the lowest L values and they decreased gradually as the grinding degree increased. The a* values (red/green axis) varied from 21.9 ± 0.2 (TPW-70-M0) to 31.2 ± 0.0 (TPW-VAC) and reflect the intensity of red coloration. The samples dried under vacuum conditions showed the most intense red coloration. The values increased with the increased degree of milling, but only up to a certain point (18 s of milling). The b* values (yellow/blue axis) ranged from 32.5 ± 0.1 (TPW-70-M0) to 52.1 ± 0.1 (TPW-LIO), indicating dominant yellow hues in all samples. The most vivid yellow was observed in the samples exposed to mild drying conditions (vacuum drying and lyophilization) and an increased degree of milling. Chroma (C*), representing overall color saturation, was highest in samples exposed to mild drying conditions (vacuum drying and lyophilization), while the lowest chroma was measured in the non-grinded sample exposed to drying at 70 °C, indicating that vacuum drying/lyophilization and finer grinding can improve color brilliance. The hue angle (h°) ranged from 56.0° ± 0.1 (TPW-70-M0) to 64.4° ± 0.0 (TPW-70-M40), showing the variation in predominant hue from orange tones to more orange–yellow tones. Browning index was low, indicating the absence of the formation of Maillard reaction products under the tested drying conditions. Significantly lower BI of intact (ungrounded) sample might be because grinding increases the surface area exposed to heat and oxygen, enhancing the rate of Maillard reactions and pigment degradation [94].

The observed variation in the colorimetric parameters shows the considerable influence of the drying conditions and the grinding intensity on the final appearance of the tomato processing waste powder. Samples processed at higher temperatures (70 °C) and coarser particle size (M40) tended to have higher L* values, possibly due to surface lightening or degradation of pigments during prolonged exposure to heat [95]. In contrast, powders obtained by vacuum drying and fine grinding (M18) showed stronger red (a*) and yellow (b*) tones, indicating better preservation of carotenoids, especially lycopene [96]. This is consistent with our data showing significantly lower lycopene content in samples dried at 70 °C compared to other drying techniques (Table 7).

The a* values indicating red coloration were the highest in samples dried under mild conditions and were positively associated with preservation of carotenoid content (Table 4) as indicated by mild positive correlation between a* and β-carotene and (to a lesser degree) lycopene (Figure S1).

The higher chroma (C*) observed in finely ground samples can be attributed to the larger surface area and uniformity of the particles, which improves color perception [97]. Furthermore, the shift in hue angle (h°) reflects a subtle balance between red and yellow pigments, which is influenced by drying kinetics and oxygen exposure during processing [98]. Overall, the visualization of the CIELAB color space allows a clear distinction between the tested samples and supports the use of instrumental color measurement for the objective evaluation of color and quality of tomato powders. These findings are consistent with previous studies reporting that vacuum drying and finer grinding improve the color retention and visual appearance of fruit and vegetable powders [96,98].

4. Conclusions

The macronutrient composition of TPW obtained from indigenous Croatian tomato varieties is characterized by a high proportion of dietary fiber (up to 52%) and available carbohydrates (up to 29%), which are the most abundant in the TPW-PEEL. Proteins (18%) and lipids (7.8%) are predominantly found in TPW-SEED, while minerals account for 4.8% of the composition. Lyophilization, as opposed to air drying at 70 °C, positively influences certain nutrient preservation. TPW is relatively rich in minerals, particularly iron (Fe), manganese (Mn), selenium (Se), and potassium (K), with distinct mineral distributions between the seed and peel fractions. Bioactive compounds are more abundant in the peel compared to the seed fraction, especially carotenoids. Lycopene is the carotenoid present in the highest concentration, while rutin, kaempferol-3-rutinoside, catechin, chlorogenic acid, and caffeic acid were identified as the major phenolic compounds. Their content is significantly affected by drying conditions and grinding intensity. Vacuum drying is optimal for preserving carotenoids and total phenols, whereas lyophilization yields the highest TEAC and better preserves specific phenolic compounds. The optimal median particle diameter for achieving optimal yields of bioactive compounds is 388 µm. Sample color is also influenced by drying and grinding degree-vacuum-dried samples exhibit the most intense red and vivid yellow hues, with color brilliance further enhanced by finer milling. Overall, the analyzed TPW is a safe, nutrient-rich resource with substantial potential for valorization in food, feed, and nutraceutical industries. The choice of drying and milling protocols significantly affects bioactive compound yield and TPW organoleptic characteristics; therefore, tailored pre-processing can enhance sustainable TPW utilization in line with circular economy principles.

This research shows that TPW obtained from the mixture of indigenous Croatian tomato varieties represents a valuable resource owing to its high content of lycopene and β-carotene, strong antioxidant activity, and distinctive reddish color. These attributes make it well-suited for use as a natural colorant and antioxidant in the food, cosmetic, and nutraceutical industries. Tomato seeds, notable for their rich mineral and protein content, offer potential for food fortification, animal feed, or as a source for tomato seed oil. The oil, distinguished by its favorable fatty acid composition and high nutritional quality, can be applied across food, cosmetic, and industrial sectors. Despite these advantages, large-scale utilization of TPW in Croatia is hindered by several challenges such as variability in TPW chemical composition and the economic limitations associated primarily with availability of the sufficient amounts of TPW.