rbz Revista Brasileira de Zootecnia R. Bras. Zootec. 1516-3598 1806-9290 Sociedade Brasileira de Zootecnia 02609 10.37496/rbz5520250108 Non-ruminants Performance, quality, and oxidative stability during storage of eggs from white laying hens fed diets supplemented with carotenoids 0000-0002-5553-9471 Santana Patrícia Gomes Conceptualization Investigation Writing – original draft 1 0000-0003-3739-2522 Garcia Elis Regina de Moraes Investigation Methodology Project administration Supervision Writing – original draft Writing – review & editing 1 * 0000-0003-0239-1132 Andrade Gislaine da Cunha de Investigation Methodology Writing – original draft 1 0000-0002-0538-2867 Ferraz André Luiz Julien Data curation Methodology Writing – original draft 1 0000-0002-4346-9373 Souza Andréia Fróes Galuci Oliveira de Investigation Methodology Writing – original draft 2 0009-0004-3181-8435 Melo Evilásio Pontes de Conceptualization Methodology Writing – original draft 3 0000-0001-9622-2844 Kiefer Charles Methodology Writing – original draft Writing – review & editing 4 Universidade Estadual de Mato Grosso do Sul Aquidauana MS Brasil Universidade Estadual de Mato Grosso do Sul, Aquidauana, MS, Brasil. Universidade Estadual de Mato Grosso do Sul Maracaju MS Brasil Universidade Estadual de Mato Grosso do Sul, Maracaju, MS, Brasil. Vetscience Bio Solutions Vet Science Nutracêuticos Ltda Maringá PR Brasil Vetscience Bio Solutions. Vet Science Nutracêuticos Ltda, Maringá, PR, Brasil. Universidade Federal de Mato Grosso do Sul Campo Grande MS Brasil Universidade Federal de Mato Grosso do Sul, Campo Grande, MS, Brasil. ermgarcia@uems.br

Ines Andretta

Alícia Zem Fraga

The authors declare no conflict of interest.

 08 07 2026 2026 55 e20250108 4 06 2025 13 03 2026 This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. ABSTRACT

The aim of the study was to evaluate the inclusion of carotenoids in the diet of layers on the production performance, quality and oxidative stability of eggs during storage. A total of 240 Dekalb White laying hens, 76 weeks of age, were used. The hens were allocated in completely randomized design with eight diets (control diet (CD); astaxanthin (AXT), 10 mg kg⁻1; lutein (LUT), 10 mg kg⁻1; lycopene (LYC), 10 mg kg⁻1; AXT + LUT, 10 mg kg⁻1 each; AXT + LYC, 10 mg kg⁻1 each; LUT + LYC, 10 mg kg⁻1 each; and AXT + LUT + LYC, 10 mg kg⁻1 each), five replicates, and six birds per experimental unit. Egg quality and oxidative stability were evaluated using a randomized 8 × 3 factorial design (diets × storage period: fresh, 15, and 30 days) with repeated measures. Eggs were stored either at room temperature (RT, 25 °C) or under refrigeration (RF, 8 °C) for the evaluation of egg quality and lipid stability. Dietary supplementation with astaxanthin (AXT; 10 mg kg⁻1), lutein (LUT; 10 mg kg⁻1), and lycopene (LYC; 10 mg kg⁻1), either individually or in combination, did not affect the zootechnical performance or overall egg quality of laying hens. However, AXT supplementation improves the oxidative stability of eggs compared with LYC, suggesting a greater protective effect against oxidative deterioration. Combined supplementation with AXT + LUT, AXT + LYC, and LUT + LYC enhances yolk pigmentation, whereas the simultaneous inclusion of all three carotenoids reduces pigmentation intensity. Regardless of dietary treatment, egg storage for up to 30 days at room temperature (25 °C) or under refrigeration (8 °C) negatively affects the internal quality of eggs.

 Keywords: astaxanthin color lutein lycopene storage CAPES 001 The authors thank the Vet Science Bio Solutions, Universidade Estadual de Mato Grosso do Sul (UEMS), Programa de Pós-Graduação em Zootecnia/UEMS, and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES; Finance Code 001) for the financial support in the execution of the research project. 1. Introduction

Eggs provide several health benefits throughout the human lifespan (Puglisi and Fernandez, 2022). They are considered one of the most nutritionally complete foods for human consumption (Zaheer, 2015), containing several essential nutrients, particularly vitamin D, vitamin B12, selenium, choline, and essential amino acids (Rafed et al., 2024). To fully capitalize on its nutritional potential, preserving eggs during the marketing period is crucial, given the weeks that may pass between laying, acquisition, and preparation. Extended periods negatively impact egg quality, with a continuous decline after laying (Carvalho et al., 2007).

A promising alternative is the use of technologies aimed at maintaining egg quality after laying, thereby extending the shelf life of eggs and egg-derived products. In the context of animal production and food technology, considerable research has focused on antioxidants and their effects on the shelf life and nutritional quality of food (Hayat et al., 2010). Eggs contain substantial amounts of unsaturated fatty acids, which highly susceptible to lipid oxidation processes, limiting their conservation capacity (Pita et al., 2004). Currently, there is a tendency to substitute synthetic antioxidants with natural alternatives due to concerns about liver alterations the formation of cancer cells associated with the exposure of animals to synthetic compounds (Jardini and Mancini Filho, 2007).

Carotenoids, when used as feed additives in poultry diets, enhance yolk color and increase the boost antioxidant capacity, protecting eggs from the harmful effects of reactive oxygen species and free radicals (Nabi et al., 2020). In terms of oxygen-sequestration ability of carotenes and xanthophylls, lycopene, AXT or canthaxanthin, β-carotene or bixin, lutein, and crocin follow an ascending order (Fontana et al., 2000). The most commonly used carotenoids are lycopene, lutein, astaxanthin, β-carotene, fucoxanthin, and canthaxanthin (Darvin et al., 2022). Carotenoids not only intensify egg yolk pigmentation but also exhibit multiple biological roles that contribute to therapeutic effects, including anticancer, immunomodulatory, anti-inflammatory, antibacterial, antidiabetic, and neuroprotective activities (Nabi et al., 2020).

Utilizing antioxidants, including carotenoids such as AXT, LUT, and lycopene, represents an alternative strategy to strengthen the enzymatic antioxidant system against free radical attacks. Moreover, synergistic interactions among antioxidants may enhance the body’s defense mechanisms and help prevent excessive free radical production, as these compounds possess distinct characteristics and modes of action. This may contribute to the inhibition of lipid peroxidation in egg yolk, thereby improving the oxidative stability of eggs (Panaite et al., 2021). Therefore, this study aimed to evaluate the effects of dietary carotenoid supplementation on the performance, egg quality, and oxidative stability of eggs from white laying hens stored for up to 30 days either at room temperature (RT) or under refrigeration (RF).

 2. Material and methods 2.1. Ethical matters

Research on animals was conducted according to guidelines of the Ethics Committee on Animal Use of the Universidade Estadual de Mato Grosso do Sul (UEMS) (protocol number 038/2017).

 2.2. Experiment design and treatments

The experiment was conducted at the Poultry and Animal Products Quality Laboratory of the Universidade Estadual de Mato Grosso do Sul, Aquidauana, MS, Brazil (20°20' S, 55°48' W, altitude of 174 m). The study involved 240 Dekalb White-strain laying hens, aged 76 weeks, evaluated over a 112-day experimental period divided into four 28-day cycles. The birds were housed in galvanized wire cages measuring 25 × 40 × 45 cm in a conventional laying house. A continuously supplied water trough was positioned along the front of the cages and cleaned daily. Feed was supplied ad libitum at 08:00 and 16:00 h in feeders located below the water troughs.

The lighting program adopted was 17 h daily (natural + artificial lighting). Thermal conditions in the aviary were assessed using a thermohygrometer at 08:00 and 16:00 h, with records of average (30.6 °C), minimum (24.0 ± 0.8 °C), and maximum temperatures (35.8 ± 2.7 °C), as well as relative humidity (83.8 ± 6.0%).

A completely randomized design was adopted with eight diets, five replications, and six birds per experimental unit. Diets included: control diet (CD) without carotenoids, CD with 10 mg kg−1 of AXT, CD with 10 mg kg−1 LUT, CD with 10 mg kg−1 LYC, CD with 10 mg kg−1 of AXT and 10 mg kg−1 of LUT, CD with 10 mg kg−1 of AXT and 10 mg kg−1 of LYC, CD with 10 mg kg−1 of LUT and 10 mg kg−1 of LYC, and CD supplement with 10 mg kg−1 of AXT, 10 mg kg−1 of LUT and 10 mg kg−1 of LYC. The experimental diets, based on corn and soybean meal, were formulated to be isoenergetic and isonutritive (Table 1) to meet the nutritional requirements of the hens, according to the strain manual (Dekalb White, 2009) and Rostagno et al. (2017). No antioxidants were included in the CD. Carotenoids were incorporated into the feed in powder form.

Percentage and calculated compositions of the control diet1
Ingredient Control diet
Corn 62.00
Soybean meal, 46.5% 22.91
Soybean oil 2.07
Calcitic limestone 10.30
Dicalcium phosphate 1.58
L-Lysine HCl, 78.5% 0.10
DL-Methionine, 99% 0.33
L-Threonine, 98% 0.06
Salt 0.50
Mineral and vitamin supplement2 0.10
Calculated values  
ME (kcal kg−1) 2,900
CP (%) 15.61
Digestible methionine + cystine (%) 0.76
Disgetible threonine (%) 0.60
Digestible lysine (%) 0.78
Calcium (%) 4.43
Available phosphorus (%) 0.37
Sodium (%) 0.21

1 Each carotenoid was added to the control diet in an amount of 10 mg kg−1 individually (Astaxanthin - AXT, Lutein - LUT, Lycopene - LYC) or combined (AXT + LUT, AXT + LYC, LUT + LYC and AXT + LUT + LYC).

2 Composition per kg of feed: vitamin A, 9,637 IU; vitamin D3, 2,409 IU; vitamin E, 36.1 IU; vitamin K3, 1.93 mg; vitamin B1, 2.59 mg; vitamin B2, 6.44 mg; nicotinic acid, 39.2 mg; pantothenic acid, 12.95 mg; vitamin B6, 3.614 mg; vitamin B12, 0.0159 mg; folic acid, 0.903 mg; biotin, 0.090 mg; choline, 392 mg; Cu, 8.37 mg; Fe, 41.68 mg; I, 0.843 mg; Mn, 58.36 mg; Se, 0.250 mg; Zn, 54.21 mg.

2.3. Evaluation of performance and quality characteristics of eggs

Animal performance was evaluated through feed intake (g hen−1day−1), egg production (%), egg mass (g hen−1), and feed conversion ratio (kg kg−1 and kg dz−1). For egg quality assessment during the last four days of each cycle, six eggs by diet were randomly selected to measure average weight (g) and specific gravity (g cm−3). Of these, three eggs were used to evaluate shell thickness, shell percentages, albumen, yolk, yolk pH, and albumen pH. The remaining eggs were used to measure the Haugh Unit (HU), yolk index (mm), and raw yolk color.

Using data from the total weight and number of eggs, average egg weight was calculated. For composition and pH analysis, eggs were individually weighed and broken into non-toxic polystyrene containers, previously identified and weighed, with complete separation of the albumen, yolk, and shell. The weight and percentage of each constituent relative to egg weight were calculated. The pH of the albumen and yolk was determined with a bench-top pH meter (Hanna Instruments®). Specific gravity was determined by the water displacement method (Hamilton, 1982).

Eggs were broken onto a smooth glass surface, albumen and yolk heights (mm) were measured using a digital caliper. Using albumen height (mm) and egg weight (g), HU values were calculated using the equation described by Silversides and Budgell (2004):

HU = 100 log ⁡ ( H + 7.75 − 1.7 W 0.37 ) ,

in which H = albumen height (mm) and W = egg weight (g).

The yolk diameter was measured horizontally with a digital caliper (± 0.05 mm), and the yolk index (mm) (height/diameter) was calculated based on mean values. Raw yolk color analysis was evaluated using a DSM colorimetric fan (Yolk Color Fan®) on a scale from 1 to 15, reflecting varying yellow tones. Shells were washed, dried at RT for 48 hours, and subsequently weighed to determine external quality using shell weight (g) and thickness (mm). Shell thickness, including membranes, was measured at four distinct equatorial points using a digital micrometer (± 0.001 mm) and averaged per experimental unit.

To evaluate egg quality and oxidative stability, a randomized design was adopted in an 8 × 3 factorial arrangement (experimental diets × storage periods), with repeated measures, using two eggs diet⁻1 day⁻1. Internal quality parameters, including HU, yolk index, yolk color, and albumen and yolk pH, were evaluated for each storage period at RT (25 °C) and RF (8 °C). Lipid oxidation was assessed using the TBARS method according to the methodologies described by Ramanathan and Das (1992) and Vyncke (1970), using three eggs diet⁻1 day⁻1 for each storage period.

 2.4. Statistical analysis

The data were submitted to the Shapiro-Wilk normality test. For normally distributed residues, analysis of variance was applied, and the means were compared using the Tukey test. The TBARS variable, which did not present a normal distribution, was analyzed using a generalized linear model with a gamma distribution and inverse link function, and means were compared using the t-test. Statistical analyses were performed using the R software. The following mathematical model was adopted:

Y i j = μ + α i + ε i j ,

in which Yij = dependent variable, μ = mean of the variable, αi = effect of diets, and εij = random error.

For quality analyses, the experimental design utilized was completely randomized in a 8 × 3 factorial scheme. The statistical model utilized was:

Y i j k = μ + D i + S j + ( D S ) i j + ε i j k ,

in which Yijk = represents the observation obtained from hens subjected to the i-th diet (i = 1-8) and j-th storage period (1-3); μ = is the overall mean; Di = is the effect of diets; Sj = is the effect of storage period; (DS)ij = is the interaction effect between diets and storage period; and εijk = is the random error associated with each observation. Statistical significance was declared at P<0.05.

 3. Results

There was no significant effect (P>0.05) on the zootechnical performance of laying hens with the inclusion of AXT, LUT, and LYC, either individually or in combination (Table 2). With respect to egg quality variables, most showed no differences among experimental diets, except for raw yolk color. LYC supplementation and the combinations of AXT and LUT, AXT and LYC, and LUT and LYC resulted in more pigmented yolks compared to the CD (Table 3). No significant effects of carotenoid inclusion, whether individually or combined, were observed for the percentages of shell, albumen, and yolk, as well as specific gravity, shell thickness, and percentage of shell per surface of area in the diets of laying hens (Table 4). Raw egg yolks from hens fed the combination containing AXT and LUT, AXT and LYC, and LUT and LYC were more pigmented than those from hens on the combination of AXT, LUT, and LYC (Table 5). Differences in the concentration of malonaldehyde (MDA) and raw yolk color were observed between experimental diets for RF eggs (8 °C) (Table 6). Yolk indexes for eggs from laying hens fed the CD supplemented with AXT and LUT, as well as AXT and LYC, decreased after 15 days of storage (Table 7).

Animal performance, laying percentage and egg mass of white laying hens fed with diets supplemented with different carotenoids
Variable FI (g hen−1 day−1) FC (kg kg−1) FC (kg dz−1) LP (%) EM (g hen−1)
Control diet 105.21 1.80 1.43 90.57 60.06
AXT 104.91 1.78 1.41 88.97 58.59
LUT 104.24 1.79 1.43 89.32 59.35
LYC 103.04 1.85 1.49 82.73 55.00
AXT + LUT 105.22 1.86 1.45 86.93 56.87
AXT + LYC 104.96 1.81 1.43 90.34 59.02
LUT + LYC 108.00 1.87 1.51 87.45 58.65
AXT + LUT + LYC 105.46 1.92 1.52 87.67 56.79
Mean 105.46 1.81 1.45 87.99 58.01
CV (%) 2.65 5.36 4.25 5.01 6.24
P-value 0.41 0.76 0.29 0.16 0.40

FI: feed intake; FC: feed conversion; LP: laying percentage; EM: egg mass; AXT: astaxanthin; LUT: lutein; LYC: lycopene; AXT + LUT: astaxanthin + lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

Egg quality of white laying hens fed diets supplemented with different carotenoids
Variable EW (g) HU YI (mm) RYC
Control diet 66.15 94.36 0.411 5.56b
AXT 65.44 93.13 0.423 6.01ab
LUT 66.58 95.21 0.414 6.20ab
LYC 66.89 93.13 0.419 6.55a
AXT + LUT 65.59 94.85 0.427 6.51a
AXT + LYC 65.79 93.79 0.423 6.56a
LUT + LYC 67.40 93.58 0.415 6.33a
AXT + LUT + LYC 64.58 94.65 0.419 5.96ab
Mean 66.05 94.08 0.418 6.21
CV (%) 2.59 2.04 4.03 5.43
P-value 0.25 0.54 0.81 <0.05

EW: egg weight; UH: Haugh unit; YI: yolk index; RYC: raw yolk color; AXT: astaxanthin; LUT: lutein; LYC: lycopene; AXT + LUT: astaxanthin + lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

Different letters in the same column differ at P<0.05.

Percentages of shell, albumen and yolk, specific gravity and shell thickness of eggs from white laying hens fed diets supplemented with different carotenoids
Variable Shell Albumen (%) Yolk (%) SG (g cm−3) ST (mm) PSSA (mg cm−2)
Control diet 8.91 73.99 24.85 1.083 0.322 75.28
AXT 8.82 73.67 25.09 1.082 0.321 78.05
LUT 8.67 73.75 25.28 1.083 0.325 75.48
LYC 8.46 73.87 25.27 1.083 0.327 75.44
AXT + LUT 8.97 73.37 25.16 1.082 0.319 74.82
AXT + LYC 8.76 73.14 25.58 1.083 0.325 75.72
LUT + LYC 8.63 73.00 25.86 1.083 0.324 75.91
AXT + LUT + LYC 8.80 72.88 26.24 1.083 0.316 75.78
Mean 8.75 73.45 25.41 1.082 0.322 75.81
CV (%) 5.01 1.42 4.68 0.23 4.26 3.33
P-value 0.67 0.58 0.65 0.99 0.93 0.64

SG: specific gravity; ST: shell thickness; PSAS: percentage of shell per surface of area; AXT: astaxanthin; LUT: lutein; LYC: lycopene; AXT + LUT: astaxanthin + lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

Malonaldehyde concentration, egg weight, Haugh unit, yolk index, pH of albumen and yolk, raw yolk color, of eggs stored for different periods of storage at room temperature, from white laying hens fed diets supplemented with different carotenoids
Variable MDA (mg kg−1) EW (g) HU YI (mm) pH RYC
Yolk Albumen
Control diet 0.174b 62.58 62.76 0.307 6.38 8.78 6.07ab
AXT 0.164b 59.54 54.91 0.280 6.42 8.91 6.40ab
LUT 0.194ab 61.86 59.58 0.305 6.19 8.82 6.25ab
LYC 0.262a 60.21 53.24 0.281 6.37 8.92 6.40ab
AXT + LUT 0.196ab 63.45 57.58 0.288 6.37 8.89 6.75a
AXT + LYC 0.198ab 61.18 58.56 0.285 6.45 8.95 6.69a
LUT + LYC 0.201ab 64.03 58.21 0.290 6.48 8.86 6.69a
AXT + LUT + LYC 0.228ab 59.71 57.70 0.298 6.31 8.91 5.84b
Storage period (SP; days)              
Fresh 0.199b 66.15a 91.60a 0.414a 6.12b 8.00b 5.39c
15 0.201b 61.71b 44.93b 0.276b 6.24b 9.30a 6.54b
30 0.207a 58.22c 41.06b 0.217c 6.75a 9.33a 7.00a
Mean 0.202 61.57 57.82 0.292 6.37 8.88 6.39
P-value*              
Diet 0.03 0.38 0.29 0.11 0.82 0.77 0.03
SP <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001
Diet × SP 0.53 0.94 0.46 0.59 0.86 0.95 0.97

MDA: malonaldehyde concentration; EW: egg weight; HU: Haugh unit; YI: yolk index; pH: pH values of yolk and albumen; RYC: raw yolk color; AXT: astaxanthin; LUT: lutein; LYC: lycopene; AXT + LUT: astaxanthin + lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

* Values with P>0.05 present normal distribution by the Shapiro Wilk test.

Different letters in the same column differ at P<0.05.

Malonaldehyde concentration, egg weight, Haugh unit, yolk index, pH of albumen and yolk, raw yolk color, of eggs stored for different periods of storage under refrigeration, from white laying hens fed diets supplemented with different carotenoids
Variable MDA (mg kg−1) EW (g) HU YI (mm) pH RYC
Yolk Albumen
Control diet 0.146b 61.24 79.15 0.416 6.16 8.66 6.00bc
AXT 0.151b 65.39 80.27 0.416 6.30 8.76 6.30abc
LUT 0.189ab 59.99 81.40 0.413 6.22 8.79 6.08bc
LYC 0.228a 59.86 81.54 0.409 6.35 8.74 6.50ab
AXT + LUT 0.194ab 64.86 82.97 0.407 6.58 8.78 6.82a
AXT + LYC 0.206ab 63.95 80.05 0.405 6.10 8.80 6.72a
LUT + LYC 0.226a 65.56 80.29 0.404 6.28 8.69 6.81a
AXT + LUT + LYC 0.246a 62.88 84.73 0.404 6.60 8.73 5.79c
Storage period (SP; days)              
Fresh 0.194 66.15a 91.60a 0.421a 6.12b 8.00b 5.59b
15 0.197 63.85a 79.98c 0.411ab 6.30ab 9.06a 6.75a
30 0.204 59.99b 79.96b 0.395b 6.56a 9.17a 6.88a
Mean 0.198 62.96 81.30 0.409 6.32 8.74 6.37
P-value*              
Diet 0.04 0.13 0.48 0.86 0.19 0.88 0.02
SP 0.17 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001
Diet × SP 0.50 0.94 0.95 0.019 0.11 0.99 0.41

MDA: malonaldehyde concentration; EW: egg weight; HU: Haugh unit; YI: yolk index; pH: pH values of yolk and albumen; RYC: raw yolk color; AXT: astaxanthin; LUT: lutein; LYC: lycopene; AXT + LUT: astaxanthin + lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

* Values with P>0,05 present normal distribution by the Shapiro Wilk test.

Different letters in the same column differ at P<0.05.

Yolk index of eggs from white laying hens fed diets with different sources of carotenoids stored up to 30 days under refrigeration
Diet Storage period (days)
Fresh 15 30
Control diet 0.448a 0.400b 0.393b
AXT 0.411 0.397 0.402
LUT 0.421 0.411 0.413
LYC 0.423 0.410 0.383
AXT + LUT 0.449a 0.399b 0.373b
AXT + LYC 0.442a 0.393b 0.391b
LUT + LYC 0.406 0.421 0.384
AXT + LUT + LYC 0.431 0.409 0.404
Mean 0.411 0.392 0.422

AXT: astaxanthin; LUT: luten; LYC: lycopene; AXT + LUT: astaxanthin + Lutein; AXT + LYC: astaxanthin + lycopene; LUT + LYC: lutein + lycopene; AXT + LUT + LYC: astaxanthin + lutein + lycopene.

Different letters in the same row differ at P<0.05.

4. Discussion

The inclusion of AXT, LUT, and LYC, alone or in combination, did not affect (P>0.05) the zootechnical performance of laying hens, corroborating previous studies using low to moderate levels of AXT and LUT (Leeson and Caston, 2004; Yang et al., 2006; Leeson et al., 2007). In contrast, improvements in performance have been reported at higher inclusion levels of AXT and LYC, suggesting that the lack of response in the present study may be related to the dietary levels evaluated (Garcia et al., 2015; Magnuson et al., 2018).

The absence of improvements in the performance of laying hens supplemented with carotenoids in the present study may be associated with the inclusion levels evaluated, since positive responses have been reported in studies using higher supplementation levels than those tested here.

Most egg quality traits were unaffected by dietary treatments, except for yolk color, which was enhanced by LYC and by carotenoid combinations. These results agree with previous findings showing increased yolk pigmentation without effects on egg weight or internal quality parameters (Yang et al., 2006; Olson et al., 2008; Garcia et al., 2015). Leeson et al. (2007) observed no differences in egg and yolk weights with LUT, but reported increased yolk color with 125 mg kg−1LUT supplementation. Magnuson et al. (2018) reported that dietary supplementation with AXT at 40 mg kg−1 significantly enhanced egg yolk color, which is consistent with the results observed in the present study for LYC. As hens do not synthesize carotenoids endogenously, yolk pigmentation is directly influenced by dietary carotenoid intake (Leeson and Caston, 2004), with corn-based diets already contributing lutein and zeaxanthin (Amaya et al., 2014).

No effects were observed on shell and external egg quality traits, consistent with previous studies (Leeson et al., 2007; Spada et al., 2012), although thicker shells have been reported with 40 mg kg−1of AXT supplementation (Magnuson et al., 2018). Under room temperature storage, carotenoid supplementation did not affect internal egg quality; however, higher yolk MDA levels were observed in LYC-supplemented diets, likely due to the greater susceptibility of LYC to oxidative degradation (Rao and Agarwal, 2000). Leeson et al. (2007) noted that eggshell quality tended to decline towards the end of the laying period, posing challenges in demonstrating the positive effects of carotenoids on shell characteristics. Carotenoids did not affect external quality, likely due to their incorporation into chylomicrons and subsequent transport through the bloodstream (Parker, 1996).

For eggs stored at RT (25 °C), carotenoid supplementation did not affect characteristics like egg weight, HU, yolk index, or the pH of the albumen and yolk. However, MDA concentration in the yolk was higher in LYC-supplemented diets compared with the control diet and AXT-supplemented diets. This can be attributed to the greater susceptibility of LYC to oxidative degradation and isomerization (Rao and Agarwal, 2000).

Egg yolks from hens fed combinations of two carotenoids (AXT + LUT, AXT + LYC, or LUT + LYC) showed greater pigmentation than those receiving the triple combination, likely due to competition among carotenoids during intestinal uptake and metabolism, which can limit absorption and deposition in the yolk (Kostic et al., 1995). This competitive interaction can prevent the absorption and transport of carotenoids to target tissues (Salter-Venzon et al., 2017), potentially explaining the differences observed in yolk color.

Results showed varying MDA concentrations in egg yolks stored at RT, with AXT exhibiting superior antioxidant activity compared with other substances (Palozza et al., 2008). The instability of LYC resulted in higher MDA values in yolks, influenced by its chemical properties and susceptibility to oxidation or isomerization (Shi et al., 2002). Xanthophylls, such as LUT, are incorporated into the membrane with their terminal groups anchored in polar regions. In turn, LYC, due to the absence of hydrophilic substituents, remains entirely within the membrane. Therefore, carotenoids’ specific structural characteristics may contribute to distinct protection mechanisms that achieve synergistic properties. The triple combination’s inability to enhance yolk color may be attributed to carotenoid competition during absorption, leading to reduced deposition in the intestinal tract and subsequent intestinal absorption (Maiani et al., 2009).

Regarding individual carotenoids, LYC at 420 mg kg−1 efficiently improved yolk pigmentation by 4 mg kg−1 (Olson et al., 2008), and LUT at 500 mg kg−1 resulted in 2.2 mg of LUT in eggs (60 g), with increased yolk pigmentation (Leeson and Caston, 2004). Egg quality declined throughout RT storage, as evidenced by the reduction in egg weight caused by water loss from the albumen (Tainika et al., 2024).

Values of HU and YI decreased with storage due to carbonic acid (H2CO3) dissociation, breakdown of the ovomucin bonds, and increased pH in the yolk and albumen (Li-Chan et al., 1995). The RYC intensity decreased with storage, attributed to the rapid transfer of iron from the yolk to the egg white during storage, as well as to protein penetration into the yolk (Santos et al., 2009).

Under RF (8 °C), diets supplemented with LUT + LYC and AXT + LUT + LYC resulted in higher MDA concentrations than the CD and the AXT-supplemented diet. Yolk color increased with AXT + LUT, AXT + LYC, and LUT + LYC compared with AXT + LUT + LYC. Lower MDA concentrations in AXT-supplemented yolks were linked to the waterproofing effect of the unique chemical structure of AXT, which limits lipid peroxidation (Goto et al., 2001).

Carotenoid antioxidant activity is dose-dependent, and at lower concentrations these compounds may exhibit pro-oxidant effects. In this context, Garcia et al. (2015) demonstrated that dietary supplementation with 400 and 800 mg kg⁻1 of LYC in laying-hen diets significantly reduced MDA concentrations in eggs, indicating improved oxidative stability. Similarly, using seaweed as a dietary source of AXT, Magnuson et al. (2018) reported dose-dependent increases in AXT levels in plasma and liver of laying hens fed diets containing 10 and 80 mg kg⁻1, demonstrating the hens’ capacity to absorb, transport, and store algae-derived phytochemicals.

Phytochemicals are transported to different tissues, particularly to the liver where they can be stored or transported to other tissues. AXT may not be metabolized by animals, unlike most carotenoids, which indicates a unique retention of this compound in tissues and may explain the lower concentration of this carotenoid in egg yolks (Magnuson et al., 2018). Carotenoids’ instability and susceptibility to oxidation contribute to yolk color loss during storage, a process that is accelerated by light, temperature, and the presence of metallic catalysts (Sarantópoulos et al., 2001).

The egg yolks produced by the hens supplemented with the combinations of AXT and LUT, AXT and LYC, and LUT and LYC showed greater pigmentation than those from hens fed the CD, LUT supplementation alone, or the combination of AXT + LUT + LYC. This result can be explained by the fact that hens primarily absorb substances present in LUT, known as oxycarotenoids, contained in the diets, especially in corn grains (Hudon, 1994).

In laying hens, oxycarotenoids stored in muscle and skin tissues are mobilized to the ovaries upon the onset of sexual maturity, and a portion of these compounds is subsequently deposited in the egg yolk, thereby determining yolk coloration (Goodwin, 1986). In general, animals are capable of absorbing, metabolizing, and depositing dietary carotenoids in various tissues, including the skin, beak, feet, and egg yolk (Breithaupt, 2007).

It has been shown that dietary supplementation with LUT at 250 mg kg−1 significantly intensified egg yolk color compared with a control diet without carotenoid inclusion, highlighting the strong pigmentation potential of this compound (Englmaierová and Skrivan, 2013). Similarly, Magnuson et al. (2018) reported increased egg yolk pigmentation in laying hens receiving AXT supplementation.

As expected, egg quality worsened during storage under RF. However, the loss of quality occurred less markedly in eggs stored under refrigeration (10 to 15 °C), according to Lana (2000). Egg weight (EW) and HU decreased, while the pH of the yolk and albumen increased with extended storage periods. These results were expected, as studies evaluating the effects of the tropical climate on eggs stored under RF have demonstrated that HU values decrease markedly during refrigerated storage, whereas yolk and albumen pH values increase due to the loss of carbon dioxide through the pores of the shell (Xavier et al., 2008). Jones et al. (2002) stated that storage time and temperature are important factors and must be controlled during storage. Thus, higher temperatures and longer storage times lead to a decline in internal egg quality, mainly indicated by lower HU values.

This fact accelerates the physicochemical reactions causing the degradation of the protein structure present in the albumin, resulting in the release of water previously bound to large protein molecules, which subsequently migrates to the yolk by osmosis (Leandro et al., 2005). The increase in yolk pH with longer storage periods aligns with findings by Shang et al. (2004). Similarly, the increase in albumen pH observed after 30 days of storage is associated with carbon dioxide loss through the enlarged pores of the shell (Li-Chan et al., 1995).

Yolk indexes obtained for eggs from laying hens that received the CD and those supplemented with AXT + LUT and AXT + LYC decreased after 15 days of storage. The progressive decrease in YI values during storage is explained by the gradual weakening of the vitelline membrane and the liquefaction of the yolk, which is mainly caused by the diffusion of water from the albumen (Akyurek and Okur, 2009).

The YI is based on the relationship between yolk height and diameter; thus, increased storage temperature may negatively affect this parameter, as observed in the present experiment. Eggs stored for up to six days after laying maintain high quality when kept at RT (25 °C), whereas eggs stored under RF (8 °C) maintain excellent quality for up to 30 days (Lana et al., 2018). Therefore, although yolk indexes decrease as storage periods increase, eggs maintained under RF tend to exhibit more stable YI values.

According to Santos et al. (2009), YI values should range between 0.35 and 0.49, indicating that all eggs analyzed when fresh and after 15 and 30 days of storage were within the range considered indicative of good quality. This variable has been recommended as one of the best indicators of egg quality when compared with HU and pH (Spada et al., 2012). Research on the inclusion of two antioxidants in the diets of laying hens is scarce. However, in the present study, the association of AXT and LUT, as well as the association of AXT and LYC, showed greater stabilization of YI values during storage.

 5. Conclusions

Dietary supplementation with astaxanthin (AXT; 10 mg kg−1), lutein (LUT; 10 mg kg−1), and lycopene (LYC; 10 mg kg⁻1), either individually or in combination, did not influence the zootechnical performance or overall egg quality of laying hens. Dietary supplementation with AXT improves the oxidative stability of eggs compared with LYC, suggesting a greater protective effect of this carotenoid against oxidative deterioration. Combined supplementation with AXT + LUT, AXT + LYC, and LUT + LYC enhances yolk pigmentation, whereas the simultaneous inclusion of all three carotenoids reduces pigmentation intensity. Regardless of dietary treatment, egg storage for up to 30 days, both at room temperature (25 °C) and under refrigeration (8 °C), negatively affects the internal quality of eggs. These findings highlight the potential of specific carotenoid combinations to improve yolk pigmentation and oxidative stability, while also reinforcing the importance of appropriate storage conditions to preserve egg quality.

 Acknowledgments

The authors thank the Vet Science Bio Solutions, Universidade Estadual de Mato Grosso do Sul (UEMS), Programa de Pós-Graduação em Zootecnia/UEMS, and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES; Finance Code 001) for the financial support in the execution of the research project.

 References Akyurek, H. and Okur, A. A. 2009. Effect of storage time, temperature and hen age on egg quality in free-range layer hens. Journal of Animal and Veterinary Advances 8:1953-1958. Akyurek H. Okur A. A. 2009 Effect of storage time, temperature and hen age on egg quality in free-range layer hens Journal of Animal and Veterinary Advances 8 1953 1958 Amaya, E.; Becquet, P.; Carné, S.; Peris, S. and Miralles, P. 2014. Carotenoids in animal nutrition. Fefana, Brussels. Amaya E. Becquet P. Carné S. Peris S. Miralles P. 2014 Carotenoids in animal nutrition Fefana Brussels Breithaupt, D. E. 2007. Modern application of xanthophylls in animal feeding: a review. Trends in Food Science and Technology 18:501-506. https://doi.org/10.1016/j.tifs.2007.04.009 Breithaupt D. E. 2007 Modern application of xanthophylls in animal feeding: a review Trends in Food Science and Technology 18 501 506 https://doi.org/10.1016/j.tifs.2007.04.009 Carvalho, F. B.; Stringhini, J. H.; Jardim Filho, R. M.; Leandro, N. S. M.; Café, M. B. and Deus, H. A. S. B. 2007. Qualidade interna e da casca para ovos de poedeiras comerciais de diferentes linhagens e idade. Ciência Animal Brasileira 8:25-29. Carvalho F. B. Stringhini J. H. Jardim R. M. Filho Leandro N. S. M. Café M. B. Deus H. A. S. B. 2007 Qualidade interna e da casca para ovos de poedeiras comerciais de diferentes linhagens e idade Ciência Animal Brasileira 8 25 29 Dekalb White. 2009. Manual de manejo das poedeiras Dekalb White. Modelo de Revisão. 39p. Dekalb White 2009 Manual de manejo das poedeiras Dekalb White. Modelo de Revisão 39 Englmaierová, M. and Skrivan, M. 2013. Effect of synthetic carotenoids, lutein, and mustard on the performance and egg quality. Scientia Agriculturae Bohemica 44:138-143. Englmaierová M. Skrivan M. 2013 Effect of synthetic carotenoids, lutein, and mustard on the performance and egg quality Scientia Agriculturae Bohemica 44 138 143 Darvin, M. E.; Lademann, J.; von Hagen, J.; Lohan, S. B.; Kolmar, H.; Meinke, M. C. and Jung, S. 2022. Carotenoids in human skin in vivo: Antioxidant and photo-protectant role against external and internal stressors. Antioxidants 11:1451. https://doi.org/10.3390/antiox11081451 Darvin M. E. Lademann J. von Hagen J. Lohan S. B. Kolmar H. Meinke M. C. Jung S. 2022 Carotenoids in human skin in vivo: Antioxidant and photo-protectant role against external and internal stressors Antioxidants 11 1451 https://doi.org/10.3390/antiox11081451 Fontana, J. D.; Mendes, S. V.; Persike, D. S.; Peracetta, L. and Passos, M. 2000. Carotenóides cores atraentes e ação biológica. Biotecnologia Ciência & Desenvolvimento 13:40-45. Fontana J. D. Mendes S. V. Persike D. S. Peracetta L. Passos M. 2000 Carotenóides cores atraentes e ação biológica Biotecnologia Ciência & Desenvolvimento 13 40 45 Garcia, E. R. M.; Cruz, F. K.; Kiefer, C.; Avila, L. R. and Souza, R. P. P. 2015. Minerais orgânicos e licopeno na alimentação de poedeiras: desempenho zootécnico e qualidade dos ovos. Arquivo Brasileiro de Medicina Veterinária e Zootecnia 67:1703-1710. https://doi.org/10.1590/1678-4162-8418 Garcia E. R. M. Cruz F. K. Kiefer C. Avila L. R. Souza R. P. P. 2015 Minerais orgânicos e licopeno na alimentação de poedeiras: desempenho zootécnico e qualidade dos ovos Arquivo Brasileiro de Medicina Veterinária e Zootecnia 67 1703 1710 https://doi.org/10.1590/1678-4162-8418 Goodwin, T. W. 1986. Metabolism, nutrition and function of carotenoids. Annual Review of Nutrition 6:273-297. https://doi.org/10.1146/annurev.nu.06.070186.001421 Goodwin T. W. 1986 Metabolism, nutrition and function of carotenoids Annual Review of Nutrition 6 273 297 https://doi.org/10.1146/annurev.nu.06.070186.001421 Goto, S.; Kogure, K.; Abe, K.; Kimata, Y.; Kitahama, K.; Yamashita, E. and Terada, H. 2001. Efficient radical trapping at the surface and inside the phospholipid membrane is responsible for highly potent antiperoxidative activity of the carotenoid astaxanthin. Biochimica et Biophysica Acta (BBA) - Biomembranes 1512:251-258. https://doi.org/10.1016/S0005-2736 (01)00326-1 Goto S. Kogure K. Abe K. Kimata Y. Kitahama K. Yamashita E. Terada H. 2001 Efficient radical trapping at the surface and inside the phospholipid membrane is responsible for highly potent antiperoxidative activity of the carotenoid astaxanthin Biochimica et Biophysica Acta (BBA) - Biomembranes 1512 251 258 https://doi.org/10.1016/S0005-2736 (01)00326-1 Hamilton, R. M. G. 1982. Methods and factors that affect the measurement of egg shell quality. Poultry Science 61:2022-2039. https://doi.org/10.3382/ps.0612022 Hamilton R. M. G. 1982 Methods and factors that affect the measurement of egg shell quality Poultry Science 61 2022 2039 https://doi.org/10.3382/ps.0612022 Hayat, Z.; Cherian, G.; Pasha, T. N.; Khattak, F. M. and Jabbar, M. A. 2010. Oxidative stability and lipid components of eggs from flax-fed hens: Effect of dietary antioxidants and storage. Poultry Science 89:1285-1292. https://doi.org/10.3382/ps.2009-00256 Hayat Z. Cherian G. Pasha T. N. Khattak F. M. Jabbar M. A. 2010 Oxidative stability and lipid components of eggs from flax-fed hens: Effect of dietary antioxidants and storage Poultry Science 89 1285 1292 https://doi.org/10.3382/ps.2009-00256 Hudon, J. 1994. Biotechnological applications of research on animal pigmentation. Biotechnology Advances 12:49-69. https://doi.org/10.1016/0734-9750 (94)90290-9 Hudon J 1994 Biotechnological applications of research on animal pigmentation Biotechnology Advances 12 49 69 https://doi.org/10.1016/0734-9750 (94)90290-9 Jardini, F. A. and Mancini Filho, J. 2007. Avaliação da atividade antioxidante em diferentes extratos da polpa e sementes da romã ( Punica granatum, L.). Revista Brasileira de Ciências Farmacêuticas 43:137-147. https://doi.org/10.1590/S1516-93322007000100017 Jardini F. A. Mancini J. Filho 2007 Avaliação da atividade antioxidante em diferentes extratos da polpa e sementes da romã ( Punica granatum, L.) Revista Brasileira de Ciências Farmacêuticas 43 137 147 https://doi.org/10.1590/S1516-93322007000100017 Jones, D. R.; Tharrington, J. B.; Curtis, P. A.; Anderson, K. E.; Keener, K. M. and Jones, F. T. 2002. Effects of cryogenic cooling of shell eggs on egg quality. Poultry Science 81:727-733. https://doi.org/10.1093/ps/81.5.727 Jones D. R. Tharrington J. B. Curtis P. A. Anderson K. E. Keener K. M. Jones F. T. 2002 Effects of cryogenic cooling of shell eggs on egg quality Poultry Science 81 727 733 https://doi.org/10.1093/ps/81.5.727 Kostic, D.; White, W. S. and Olsen, J. A. 1995. Intestinal absorption, serum clearance, and interactions between lutein and B-carotene when administered to human adults in separate or combined oral doses. American Journal of Clinical Nutrition 62:604-610. https://doi.org/10.1093/ajcn/62.3.604 Kostic D. White W. S. Olsen J. A. 1995 Intestinal absorption, serum clearance, and interactions between lutein and B-carotene when administered to human adults in separate or combined oral doses American Journal of Clinical Nutrition 62 604 610 https://doi.org/10.1093/ajcn/62.3.604 Lana, G. R. Q. 2000. Avicultura. Livraria e Editora Rural Ltda., Campinas. Lana G. R. Q. 2000 Avicultura Livraria e Editora Rural Ltda Campinas Lana, S. R. V.; Lana, G. R. Q.; Silva, L. C. L.; Salvador, E. L.; Leão, A. P. A.; Lana, A. M. Q. and Barros Jr, R. F. 2018. Effect of temperature and storage time on the quality of eggs from commercial laying hen. Archivos de Zootecnia 67:93-98. Lana S. R. V. Lana G. R. Q. Silva L. C. L. Salvador E. L. Leão A. P. A. Lana A. M. Q. Barros R. F. Jr 2018 Effect of temperature and storage time on the quality of eggs from commercial laying hen Archivos de Zootecnia 67 93 98 Leandro, N. S. M.; Deus, H. A. B.; Stringhini, J. H.; Café, M. B.; Andrade, M. A. and Carvalho, F. B. 2005. Aspectos de qualidade interna e externa de ovos comercializados em diferentes estabelecimentos na região de Goiânia. Ciência Animal Brasileira 6:71-78. Leandro N. S. M. Deus H. A. B. Stringhini J. H. Café M. B. Andrade M. A. Carvalho F. B. 2005 Aspectos de qualidade interna e externa de ovos comercializados em diferentes estabelecimentos na região de Goiânia Ciência Animal Brasileira 6 71 78 Leeson, S. and Caston, L. 2004. Enrichment of eggs with lutein. Poultry Science 83:1709-1712. https://doi.org/10.1093/ps/83.10.1709 Leeson S. Caston L. 2004 Enrichment of eggs with lutein Poultry Science 83 1709 1712 https://doi.org/10.1093/ps/83.10.1709 Leeson, S.; Caston, L. and Namkung, H. 2007. Effect of dietary lutein and flax on performance, egg composition and liver status of laying hens. Canadian Journal of Animal Science 87:365-372. https://doi.org/10.4141/A06-043 Leeson S. Caston L. Namkung H. 2007 Effect of dietary lutein and flax on performance, egg composition and liver status of laying hens Canadian Journal of Animal Science 87 365 372 https://doi.org/10.4141/A06-043 Li-Chan, E.; Powrie, W. D. and Nakai, S. 1995. The chemistry of eggs and egg products. p.109-160. In: Egg science and technology. 4th ed. Stadelman, W. J. and Cotterill, O. J., eds. Food Products Press, New York. Li-Chan E. Powrie W. D. Nakai S. 1995 The chemistry of eggs and egg products 109 160 Egg science and technology 4th Stadelman W. J. Cotterill O. J. eds Food Products Press New York Maiani, G.; Periago Castón, M. J.; Catasta, G.; Toti, E.; Cambrodón, I. G.; Bysted, A.; Granado-Lorencio, F.; Olmedilla-Alonso, B.; Knuthsen, P.; Valoti, M.; Böhm, V.; Mayer-Miebach, E.; Behsnilian, D. and Schlemmer, U. 2009. Carotenoids: Actual knowledge of food sources, intakes, stability and bioavailability and their protective role in humans. Molecular Nutrition & Food Research 53:S194-S218. https://doi.org/10.1002/mnfr.200800053 Maiani G. Periago Castón M. J. Catasta G. Toti E. Cambrodón I. G. Bysted A. Granado-Lorencio F. Olmedilla-Alonso B. Knuthsen P. Valoti M. Böhm V. Mayer-Miebach E. Behsnilian D. Schlemmer U. 2009 Carotenoids: Actual knowledge of food sources, intakes, stability and bioavailability and their protective role in humans Molecular Nutrition & Food Research 53 S194 S218 https://doi.org/10.1002/mnfr.200800053 Magnuson, A. D.; Sun, T.; Yin, R.; Liu, G.; Tolba, S.; Shinde. S. and Lei, X. G. 2018. Supplemental microalgal astaxanthin produced coordinated changes in intrinsic antioxidant systems of layer hens exposed to heat stress. Algal Research 33:84-90. https://doi.org/10.1016/j.algal.2018.04.031 Magnuson A. D. Sun T. Yin R. Liu G. Tolba S. Shinde. S. Lei X. G. 2018 Supplemental microalgal astaxanthin produced coordinated changes in intrinsic antioxidant systems of layer hens exposed to heat stress Algal Research 33 84 90 https://doi.org/10.1016/j.algal.2018.04.031 Nabi, F.; Arain, M. A.; Rajput, N.; Alagawany, M.; Soomro, J.; Umer, M.; Soomro, F.; Wang, Z.; Ye, R. and Liu, J. 2020. Health benefits of carotenoids and potential application in poultry industry: A review. Journal of Animal Physiology and Animal Nutrition 104:1809-1818. https://doi.org/10.1111/jpn.13375 Nabi F. Arain M. A. Rajput N. Alagawany M. Soomro J. Umer M. Soomro F. Wang Z. Ye R. Liu J. 2020 Health benefits of carotenoids and potential application in poultry industry: A review Journal of Animal Physiology and Animal Nutrition 104 1809 1818 https://doi.org/10.1111/jpn.13375 Olson, J. B.; Ward, N. E. and Koutsos, E. A. 2008. Lycopene incorporation into egg yolk and effects on laying hen immune function. Poultry Science 87:2573-2580. https://doi.org/10.3382/ps.2008-00072 Olson J. B. Ward N. E. Koutsos E. A. 2008 Lycopene incorporation into egg yolk and effects on laying hen immune function Poultry Science 87 2573 2580 https://doi.org/10.3382/ps.2008-00072 Palozza, P.; Barone, E.; Mancuso, C. and Picci, N. 2008. The protective role of carotenoids against 7-keto-cholesterol formation in solution. Molecular and Cellular Biochemistry 309:61-68. https://doi.org/10.1007/s11010-007-9643-y Palozza P. Barone E. Mancuso C. Picci N. 2008 The protective role of carotenoids against 7-keto-cholesterol formation in solution Molecular and Cellular Biochemistry 309 61 68 https://doi.org/10.1007/s11010-007-9643-y Parker, R. S. 1996. Absorption, metabolism, and transport of carotenoids. The FASEB Journal 10:542-551. https://doi.org/10.1096/fasebj.10.5.8621054 Parker R. S. 1996 Absorption, metabolism, and transport of carotenoids The FASEB Journal 10 542 551 https://doi.org/10.1096/fasebj.10.5.8621054 Panaite, T. D.; Nour, V.; Saracila, M.; Turcu, R. P.; Untea, A. E. and Vlaicu, P. A. 2021. Effects of linseed meal and carotenoids from different sources on egg characteristics, yolk fatty acid and carotenoid profile and lipid peroxidation. Foods 10:1246. https://doi.org/10.3390/foods10061246 Panaite T. D. Nour V. Saracila M. Turcu R. P. Untea A. E. Vlaicu P. A. 2021 Effects of linseed meal and carotenoids from different sources on egg characteristics, yolk fatty acid and carotenoid profile and lipid peroxidation Foods 10 1246 https://doi.org/10.3390/foods10061246 Pita, M. C. G.; Piber Neto, E.; Nakaoka, L. M. and Mendonça Junior, C. X. 2004. Efeito da adição de ácidos graxos insaturados e de vitamina E à dieta de galinhas e seu reflexo na composição lipídica e incorporação de a-tocoferol na gema do ovo. Brazilian Journal of Veterinary Research and Animal Science 41:25-31. Pita M. C. G. Piber E. Neto Nakaoka L. M. Mendonça C. X. Junior 2004 Efeito da adição de ácidos graxos insaturados e de vitamina E à dieta de galinhas e seu reflexo na composição lipídica e incorporação de a-tocoferol na gema do ovo Brazilian Journal of Veterinary Research and Animal Science 41 25 31 Puglisi, M. J. and Fernandez, M. L. 2022. The health benefits of egg protein. Nutrients 14:2904. https://doi.org/10.3390/nu14142904 Puglisi M. J. Fernandez M. L. 2022 The health benefits of egg protein Nutrients 14 2904 https://doi.org/10.3390/nu14142904 Rafed, R.; Abedi, M. H. and Mushfiq, S. R. 2024. Nutritional value of eggs in human diet. Journal for Research in Applied Sciences and Biotechnology 3:172-176. https://doi.org/10.55544/jrasb.3.1.28 Rafed R. Abedi M. H. Mushfiq S. R. 2024 Nutritional value of eggs in human diet Journal for Research in Applied Sciences and Biotechnology 3 172 176 https://doi.org/10.55544/jrasb.3.1.28 Ramanatham, L. and Das, N. P. 1992. Studies on the control of lipid oxidation in ground fish by some polyphenolic natural products. Journal of Agricultural and Food Chemistry 40:17-21. https://doi.org/10.1021/JF00013A004 Ramanatham L. Das N. P. 1992 Studies on the control of lipid oxidation in ground fish by some polyphenolic natural products Journal of Agricultural and Food Chemistry 40 17 21 https://doi.org/10.1021/JF00013A004 Rao, A. V. and Agarwal, S. 2000. Role of antioxidant lycopene in cancer and heart disease. Journal of the American College of Nutrition 19:563-569. https://doi.org/10.1080/07315724.2000.10718953 Rao A. V. Agarwal S. 2000 Role of antioxidant lycopene in cancer and heart disease Journal of the American College of Nutrition 19 563 569 https://doi.org/10.1080/07315724.2000.10718953 Rostagno, H. S.; Albino, L. F. T.; Hannas, M. I.; Donzele, J. L.; Sakomura, N. K.; Perazzo, F. G.; Saraiva, A.; Teixeira, M. L.; Rodrigues, P. B.; Oliveira, R. F.; Barreto, S. L. T. and Brito, C. O. 2017. Tabelas brasileiras para aves e suínos: composição de alimentos e exigências nutricionais. 4.ed. UFV, Departamento de Zootecnia, Viçosa, MG. Rostagno H. S. Albino L. F. T. Hannas M. I. Donzele J. L. Sakomura N. K. Perazzo F. G. Saraiva A. Teixeira M. L. Rodrigues P. B. Oliveira R. F. Barreto S. L. T. Brito C. O. 2017 Tabelas brasileiras para aves e suínos: composição de alimentos e exigências nutricionais 4 UFV, Departamento de Zootecnia Viçosa, MG Salter-Venzon, D.; Kazlova, V.; Izzy Ford, S.; Intra, J.; Klosner, A. E. and Gellenbeck, K. W. 2017. Evidence for decreased interaction and improved carotenoid bioavailability by sequential delivery of a supplement. Food Science & Nutrition 5:424-433. https://doi.org/10.1002/fsn3.409 Salter-Venzon D. Kazlova V. Izzy Ford S. Intra J. Klosner A. E. Gellenbeck K. W. 2017 Evidence for decreased interaction and improved carotenoid bioavailability by sequential delivery of a supplement Food Science & Nutrition 5 424 433 https://doi.org/10.1002/fsn3.409 Santos, M. S. V.; Espíndola, G. B.; Lôbo, R. N. B.; Freitas, E. R.; Guerra, J. L. L. and Santos, A. B. E. 2009. Efeito da temperatura e estocagem em ovos. Food Science and Technology 29:513-517. https://doi.org/10.1590/S0101-20612009000300009 Santos M. S. V. Espíndola G. B. Lôbo R. N. B. Freitas E. R. Guerra J. L. L. Santos A. B. E. 2009 Efeito da temperatura e estocagem em ovos Food Science and Technology 29 513 517 https://doi.org/10.1590/S0101-20612009000300009 Sarantópoulos, C. I. G. L.; Oliveira, L. M. and Canavesi, E. 2001. Requisitos de conservação de alimentos em embalagens flexíveis. CETEA/ITAL, Campinas. Sarantópoulos C. I. G. L. Oliveira L. M. Canavesi E. 2001 Requisitos de conservação de alimentos em embalagens flexíveis CETEA/ITAL Campinas Shang, X. G.; Wang, F. L.; Li, D. F.; Yin, D. J. and Li, J. Y. 2004. Effects of dietary conjugated linoleic acid on the productivity of laying hens and egg quality during refrigerated storage. Poultry Science 83:1688-1695. https://doi.org/10.1093/ps/83.10.1688 Shang X. G. Wang F. L. Li D. F. Yin D. J. Li J. Y. 2004 Effects of dietary conjugated linoleic acid on the productivity of laying hens and egg quality during refrigerated storage Poultry Science 83 1688 1695 https://doi.org/10.1093/ps/83.10.1688 Shi, J.; Le Maguer, M. and Bryan, M. 2002. Lycopene from tomatoes. p.135-163. In: Shi, J.; Le Maguer, M. and Mazza, G. (eds.). Functional foods: Biochemical and processing aspects. v.2. Functional Foods and Nutraceuticals Series. CRC Press. Shi J. Le Maguer M. Bryan M. 2002 Lycopene from tomatoes 135 163 Shi J. Le Maguer M. Mazza G. eds Functional foods: Biochemical and processing aspects 2 Functional Foods and Nutraceuticals Series CRC Press Silversides, F. G. and Budgell, K. 2004. The relationships among measures of egg albumen height, pH, and whipping volume. Poultry Science 83:1619-1623. https://doi.org/10.1093/ps/83.10.1619 Silversides F. G. Budgell K. 2004 The relationships among measures of egg albumen height, pH, and whipping volume Poultry Science 83 1619 1623 https://doi.org/10.1093/ps/83.10.1619 Spada, F. P.; Brazaca, S. G. C.; Coelho, A. D.; Savino, V. J. M.; França, L. C.; Correr, E.; Martins, E.; Fisher, F. S. and Lemes, D. E. A. 2012. Adição de carotenóides naturais e artificiais na alimentação de galinhas poedeiras: efeitos na qualidade de ovos frescos e armazenados. Ciência Rural 42:346-353. https://doi.org/10.1590/S0103-84782012000200025 Spada F. P. Brazaca S. G. C. Coelho A. D. Savino V. J. M. França L. C. Correr E. Martins E. Fisher F. S. Lemes D. E. A. 2012 Adição de carotenóides naturais e artificiais na alimentação de galinhas poedeiras: efeitos na qualidade de ovos frescos e armazenados Ciência Rural 42 346 353 https://doi.org/10.1590/S0103-84782012000200025 Tainika, B.; Abdallah, N.; Damaziak, K.; Waithaka Ng'ang'a, Z.; Shah, T. and Wójcik, W. 2024. Egg storage conditions and manipulations during storage: effect on egg quality traits, embryonic development, hatchability and chick quality of broiler hatching eggs. World's Poultry Science Journal 80:75-107. https://doi.org/10.1080/00439339.2023.2252785 Tainika B. Abdallah N. Damaziak K. Waithaka Ng'ang'a Z. Shah T. Wójcik W. 2024 Egg storage conditions and manipulations during storage: effect on egg quality traits, embryonic development, hatchability and chick quality of broiler hatching eggs World's Poultry Science Journal 80 75 107 https://doi.org/10.1080/00439339.2023.2252785 Vyncke, B. W. 1970. Direct determination of the thiobarbituric acid value in trichloracetic acid extracts of fish as a measure of oxidative rancidity. Fette, Seifen, Anstrichmittel 72:1084-1087. https://doi.org/10.1002/lipi.19700721218 Vyncke B. W. 1970 Direct determination of the thiobarbituric acid value in trichloracetic acid extracts of fish as a measure of oxidative rancidity Fette, Seifen, Anstrichmittel 72 1084 1087 https://doi.org/10.1002/lipi.19700721218 Xavier, I. M. C.; Cançado, S. V.; Figueiredo, T. C.; Lara, L. J. C.; Lana, A. M. Q.; Souza, M. R. and Baião, N. C. 2008. Qualidade de ovos de consumo submetidos a diferentes condições de armazenamento. Arquivo Brasileiro de Medicina Veterinária e Zootecnia 60:953-959. https://doi.org/10.1590/S0102-09352008000400026 Xavier I. M. C. Cançado S. V. Figueiredo T. C. Lara L. J. C. Lana A. M. Q. Souza M. R. Baião N. C. 2008 Qualidade de ovos de consumo submetidos a diferentes condições de armazenamento Arquivo Brasileiro de Medicina Veterinária e Zootecnia 60 953 959 https://doi.org/10.1590/S0102-09352008000400026 Yang, Y. X.; Kim, Y. J.; Jin, Z.; Lohakare, J. D.; Kim, C. H.; Ohh, S. H.; Lee, S. H.; Choi, J. Y. and Chae, B. J. 2006. Effects of dietary supplementation of astaxanthin on production performance, egg quality in layers and meat quality in finishing pigs.Asian-Australasian Journal of Animal Science 19:1019-1025. https://doi.org/10.5713/ajas.2006.1019 Yang Y. X. Kim Y. J. Jin Z. Lohakare J. D. Kim C. H. Ohh S. H. Lee S. H. Choi J. Y. Chae B. J. 2006 Effects of dietary supplementation of astaxanthin on production performance, egg quality in layers and meat quality in finishing pigs Asian-Australasian Journal of Animal Science 19 1019 1025 https://doi.org/10.5713/ajas.2006.1019 Zaheer, K. 2015. An updated review on chicken eggs: production, consumption, management aspects and nutritional benefits to human health. Food and Nutrition Sciences 6:1208-1220. https://doi.org/10.4236/fns.2015.613127 Zaheer K 2015 An updated review on chicken eggs: production, consumption, management aspects and nutritional benefits to human health Food and Nutrition Sciences 6 1208 1220 https://doi.org/10.4236/fns.2015.613127

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