Egg collection was undertaken during a 2-week period, between 1 and 25 February 2022. Eggs were collected from areas where the largest producers of commercially sold eggs in Romania are located. The areas from where the eggs were collected are shown in Table 1 and Fig. 1. A total of 50 samples were collected, 10 from each housing system. Forty eggs were collected from commercial producers and 10 from villages where people raise their hens in backyards for household consumption of produced eggs. All hens were fed with commercial feed with equal mineral content, composed of 2800 kcal/kg metabolisable energy, 18% crude protein, 0.72% cysteine+methionine, 0.9% lysine, 3.5% calcium and 0.35% phosphorus. Hens were also given mineral supplements on a regular basis. In addition, free-range, organic and backyard chickens had access to outdoor foraging areas. Backyard hens frequently received food leftovers from household meals with no supplementary vitamins and minerals. Organic and free-range poultry had access to yards with natural grass and soil for 5 to 6 hours per day. Backyard chickens had access to the yard and surrounding areas for more than 8 hours per day.

Figure 1.

Map of Romania with areas of collection. (TUBS, CC BY-SA 3.0).

The collected eggs were cracked and the egg white was separated from the yolk into ceramic containers. The samples were heated in ventilated ovens at 104 °C until no further weight loss was registered. All samples were evenly ground and homogenised.

The glassware used for analysis was firstly washed with detergent and rinsed, then filled with previously-prepared 6N nitric acid (HNO₃), left for 12 hours and, finally, rinsed with double-distilled water.

Using a digital analytical balance, five grams of each sample were measured and added to a glass beaker containing a 10 ml solution composed of 10 volumetric parts of concentrated HNO₃ (65%) and 3 volumetric parts of concentrated sulphuric acid (H₂SO₄). The beakers were left for 30 minutes during which time the initial reaction between the egg and the acid mixture abated. Next, the beakers were heated to 90 °C for 45 minutes, then the temperature was increased to 140 °C. During the boiling process, the volume in each container was maintained above 3 ml by the addition of concentrated HNO₃. The wet digestion process was complete at the time when the solution inside the beakers turned lighter and the released steam became white-coloured. For metal ions fixation, a mixture of 0.5% hydrochloric acid (HCl) and 2% HNO₃ was added to each solution. After filtration through ceramic filters, the samples were collected in sterile containers and double-distilled water was added until 25 ml final volume per sample was reached.

A blank sample was prepared in a similar way without the addition of the egg component.

The analytical investigations were performed with a high-resolution radial viewing ICP-OES system - HORIBA JY ULTIMA 2 (Jobin Yvon, Longjumeau, France). The stock solutions of the elements of interest were prepared by using Merck mono – element standard solutions, traceable to SRM from NIST 1000 mg/l Certipur.

The ICP-OES working conditions are described in Table 2.

The collected data were subject to one-way variance analysis followed by the Tukey-HSD test for testing the effects of the husbandry system on the mineral content of eggs. Differences with p < 0.05 were considered statistically significant. All statistical analyses were performed using SPSS for Mac OS (IBM® SPSS® Statistics, version 23, IBM Corp.)

Table 3 identifies the detected values for the essential microelements. The higher values for essential microelements are closely dependent on the origin of the eggs – those from backyard, organic and free-range systems showed higher concentrations of essential elements, giving higher nutritional value to the eggs.

Considered an essential element in the human diet, chromium functions in maintaining normal glucose tolerance primarily by regulating insulin action. It has been described that the presence of optimal amounts of biologically-active chromium resulted in much lower amounts of insulin being required. Glucose intolerance, related to insufficient dietary chromium, appears to be widespread (Anderson 1992). Although the mechanism of this action has not been clearly described, it has been proposed that chromium binds to an oligopeptide to form chromodulin, a low-molecular-weight, chromium-binding substance that binds to and activates the insulin receptor to promote insulin action (Ross et al. 2014).

In accordance with the US Institute of Medicine (2001), the recommended daily intake for chromium for an adult is 25–35 mcg/day (US Institute of Medicine, Food and Nutrition Board 2001). With the average content of chromium per egg of approx. 2 mcg/serving, a single egg provides 1% of the daily recommended dosage of the mineral.

In 2014, the European Food Safety Authority (EFSA) published a scientific opinion concluding that there is a lack of scientific evidence for chromium being an essential element and, therefore, setting chromium intake recommendations would be inappropriate (EFSA 2014).

As seen in Table 3, the medium value for chromium content is nearly double in the backyard, organic and free-range eggs in comparison with those from intensive farming: 0.05546 mg/kg vs. 0.02166 mg/kg.

In its role as an essential mineral, copper has an important role as a co-factor in some enzymes, related to the energy production in the body, synthesis of neurotransmitters and connective tissue, metabolism of iron and activation of neuropeptides (US Institute of Medicine, Food and Nutrition Board 2001; Li et al. 2005).

In accordance with the US Institute of Medicine (2001), the recommended daily intake of copper for an adult is 900–1300 mcg/day (US Institute of Medicine Food and Nutrition Board 2001). One egg contains approx. 10 mcg/serving, thus providing 1.1% of the recommended daily intake of copper.

In our results, the eggs from intensive systems have lower medium concentrations of copper 0.7100 mg/kg compared to 1.1749 mg/kg of those from ecological housing systems.

Iron is a highly important element in the human body. As part of the blood haemoglobin, it is required for oxygen transport and oxidative metabolism. It is also essential in the processes of cellular growth. Lack of nutritional iron can result in iron-deficient anaemia – a disorder leading to reduced performance and increased morbidity.

The current recommended daily intake for adults is between 10 and 15 mg/day (US Institute of Medicine, Food and Nutrition Board 2001). With their iron concentration, eggs could play an important role in human nutrition as a source of dietary iron. With their low energy values, eggs could be an easy choice for these requiring high iron and low-calorie nutrition. One egg provides approx. 10% of the daily intake of iron and less than 40% of the daily calories intake (for 3000 kcal diet).

In our study, the average iron content in ecological eggs was more than five times higher in comparison with the other housing systems: 5.1993 mg/kg vs. 0.8753 mg/kg. This is the result of the access that some of the hens have to iron-rich vegetation (Urtica dioica) and iron-rich soils.

Manganese is an essential element with a trace presence in the human body. It acts as a co-factor in many enzymes through whose actions, manganese is involved in amino acid, cholesterol, glucose and carbohydrate metabolism and immune response (US Institute of Medicine Food and Nutrition Board 2001). With its lower concentration in different housing systems products, we cannot consider eggs as an important nutritional source for the mineral. Meeting the recommended daily intake of between 1.8 and 2.6 mg/day will require a high number of consumed eggs (US Institute of Medicine, Food and Nutrition Board 2001).

Our study showed that most of the eggs are with manganese concentrations below the concentrations detectable by ICP-OES.

One of the most important elements in the human body, zinc is involved in the catalytic activities of hundreds of enzymes, plays a role in DNA and protein synthesis and assists with the proper functioning of the immune system (US Institute of Medicine Food and Nutrition Board 2001).

With recommended daily average intake for adults between 8 and 12 mg/day, eggs could be an easy choice for providing supplementary zinc with low calorie intake (US Institute of Medicine, Food and Nutrition Board 2001). From our study, the detected concentration of the element would provide approx. 5% of the daily intake from a single egg.

The zinc concentration in both types of housing systems had similar concentrations, with its being a little higher in the ecological systems.

In addition to the essential microelements, we tested the yolk samples for the presence of some heavy metals. These metals have no biological functions and are considered contaminants in animal forages (EC 2002). Table 4 shows the levels of Al, Cd, Ni and Pb in the yolk obtained from different housing systems.

Besides the essential and heavy metals, we obtained results for some other elements, as shown in Table 5.

Boron is an element naturally found in grains and mostly in leafy vegetables and greens. With an estimated daily requirement of 1–1.5 mg/day for an adult, eggs of any origin can be used as a good dietary source for the element (WHO 1996).

Higher concentrations of barium in ecologically-produced eggs result from the consumption of green vegetables by the hens when they have access to open fields. For backyard hens, the level of the element can be explained with the addition of lettuce and carrots to their diet.

A higher concentration of strontium in backyard eggs can be explained by richer concentrations of the element in the soils from the areas where the hens were housed. It is evident from other studies that the presence of strontium in eggs has the opposite correlation with calcium – calcium is lower in concentration in the eggs with higher strontium presence (Doberenz et al. 1969).

Magnesium is required for maintaining numerous body processes – muscle and nerve functions, heart rate, blood sugar and DNA production. With a daily recommended dosage of 310–420 mg per adult, eggs are a good source for a healthy diet (US Institute of Medicine, Food and Nutrition Board 1997). Our results demonstrated very slight differences between the eggs from all tested housing systems.