Study of Hygienic Quality of Mare’s Milk and Its Use in the Development of Natural Cosmetics

Abstract

Background: The natural cosmetics market is expanding, and milk, valued for its biological properties and low toxicity, is gaining popularity as a cosmetic ingredient due to its moisturizing, anti-inflammatory, and anti-aging effects. Mare’s milk, distinct from cow’s milk, offers superior microbiological quality and potential as a luxury product, though it remains underutilized in Poland’s cosmetics industry. This study examined the hygienic quality of mare’s milk and soaps derived from it. Methods: The study was conducted on a stud farm with twenty-five mares and two stallions of the Sztumski breed, under strict hygiene and feeding standards. Physicochemical and microbiological analyses of mare’s milk and the resulting soaps included assessments of nutrient content, microbiological testing, and challenge tests conducted in accordance with ISO 11930 to evaluate antimicrobial properties and product safety. Results: The milk showed high microbiological quality, low fat (0.64–0.96%) and protein (1.70%) content, and a high lactose level (6.61%). Most soap samples were free of microbial growth, demonstrating their hygienic status and effective production decontamination. Although some preservatives showed limited efficacy against specific microorganisms, three soap samples remained resistant to contamination throughout the 28-day challenge test. Conclusions: Overall, mare’s milk soaps proved safe and stable. Improvement of their formulation could further enhance their stability and competitiveness in the natural cosmetics market.

1. Introduction

The market for cosmetic products described as natural has been growing at a rate of more than 10% per year over the last quarter of a century and occupies an increasingly prominent position in the sales of cosmetics and toiletry products, and in a wide assortment of household chemical products [1,2]. In 2023, the organic cosmetics market in Poland was valued at PLN 224 million (Polish Zloty) [3]. Plant-, animal-, and mineral-derived active substances have been known and used for many years in pharmacy, herbal medicine, homeopathy, and cosmetics [1]. The cosmetics industry is currently seeking ingredients with higher bioactivity and bioavailability for natural cosmetics [4]. One such ingredient of animal origin is milk. According to Kazimierska and Kalinowska-Lis [5], milk and colostrum possess high biological potential and, owing to their natural origin and non-toxicity, they have numerous applications in cosmetics and dermatology.

Natural cosmetics are products containing ingredients of natural origin, exhibiting soothing properties with minimal risk of adverse effects. According to standards defined by organizations such as COSMOS or ECOCERT, these products must comply with specific criteria regarding ingredient sourcing, processing methods, and environmental impact. Increasing consumer interest in ecological and health-related aspects supports the popularity of natural and organic cosmetics as a safe and environmentally friendly alternative [6,7]. In organic cosmetics, the use of synthetic and semi-synthetic substances is prohibited. If such substances are present, the product composition must rely exclusively on organic materials produced in accordance with strict standards. One of the key criteria in the evaluation of organic cosmetics is possession of a certificate issued by internationally recognized certifying bodies (e.g., COSMOS and ECOCERT). Products containing no more than 5% synthetic ingredients may receive certification confirming their organic origin, as defined by organizations such as COSMOS-standard AISBL or ECOCERT [7,8,9,10].

There is also a group of products known as dermocosmetics, classified as active or functional cosmetics. These serve as an intermediate category between traditional cosmetic and pharmaceutical products. They contain active substances and exert cosmetic effects, but do not fall under medicinal product regulations. Although the term “functional cosmetics” lacks an official legal definition in the European Union, it is commonly used in marketing and the scientific literature [9,11].

Milk is rich in proteins, including casein and whey proteins, i.e., α-lactalbumin (α-LA), β-lactoglobulin (β-LG), serum albumin (BSA), and immunoglobulins, and proteins with antimicrobial properties—lactoferrin, lactoperoxidase, and lysozyme [6,7]. This last group is involved in destroying pathogenic microbes, e.g., Escherichia coli, Candida albicans, Clostridium difficile, Shigella flexneri, Streptococcus mutans, and Helicobacter pylori. Milk proteins and the peptides derived from them also exhibit opioid, antioxidant, anti-tumor, and immunomodulatory activity. Milk and milk products were used for cosmetic purposes as early as ancient times. Currently, milk and its components are once again increasingly used as cosmetic ingredients, as they can improve skin condition by reducing acne lesions and blackheads, regulating sebum secretion, alleviating inflammatory changes, and exerting moisturizing, protective, tonic, smoothing, whitening, anti-aging, and irritation-relieving effects [5].

The composition of mare milk differs significantly from that of cow milk and is similar to that of human milk [8,9,10]. It has lower fat content than cow milk but a much higher content of α-linolenic acid (ALA), and lower content of stearic acid (18:0) and saturated fatty acids (SFAs) [11]. The protein content in mare milk (1.5–2.8%) is lower than in cow milk (3.1–3.8%) [12]. The casein content in mare milk (1.07%) is less than half that in cow milk (2.51%) but three times higher than in human milk (0.37%) [13]. This milk contains high levels of lactoferrin (9.9–10.0% of whey proteins), lysozyme (6.6–6.9% of whey proteins), and immunoglobulins (18.7–20.9% of whey proteins). Due to its high content of antibacterial substances such as lactoferrin, lysozyme, lactoperoxidase, and immunoglobulins, mare milk is of higher microbiological quality than raw cow milk [12]. This has been confirmed by Musaev et al. [14], who demonstrated that mare milk has antibacterial and antiviral properties. Czyżak-Runowska et al. [15] tested the quality of raw and refrigerated mare milk and did not detect Salmonella spp. or coliforms. They also confirmed that mare milk can be stored for 72 h at 4 °C without significantly affecting its microbiological quality. Dankow et al. [16] also confirm the high hygienic quality of mare milk. In terms of somatic cell count and total microbial count, it is markedly superior to milk from other farm animals. The quality and chemical composition of mare milk are influenced by multiple factors, both environmental and associated with breed or geographic location. Mare milk has long been used as a supportive agent in the treatment of many diseases. The antibodies and proteins (lysozyme and lactoferrin) it contains help fight infections and reduce inflammatory reactions. They exert a beneficial effect on the human body, playing an important role in the prevention and treatment of many diseases. The increased interest in this type of milk may also be due, among other things, to its vitamin content, which is essential for the proper functioning of the body. It has been used to treat tuberculosis, viral hepatitis type C, psoriasis, gastrointestinal conditions (gastric ulcers, cirrhosis, inflammation of the gallbladder, and pancreatitis), respiratory conditions (bronchitis, whooping cough, and asthma), and migraines [13]. There is currently renewed interest in mare milk and its products [17,18,19,20]. The production and processing of mare milk are developing in France, Italy, Mongolia, China, Kazakhstan, Kyrgyzstan, Greece, Germany, and many other countries [19]. In practical terms, the lack of an organized network for mare milk sales in the European Union is a significant limitation on its widespread availability. In addition, the short shelf life of raw milk requires the use of preservation methods such as freezing, freeze-drying, or spray drying [15,21]. In Poland, the milking of mares to obtain milk for consumption or for pharmaceutical or cosmetic purposes is practically unknown [17].

Due to its unique nutritional properties and low allergenicity, mare milk is a valuable cosmetic product [22]. It is most commonly used in various balms and soaps with antioxidant properties [23]. Due to the unique properties of mare milk and the growing consumer interest in prestige goods, these cosmetics may gain recognition among customers around the world. Therefore, an attempt was made to determine the hygienic status of mare milk and soaps produced from it in terms of microbiological quality and safety of use.

2. Materials and Methods

2.1. Animals

The study was carried out at a stud farm housing approximately 25 mares and 2 stallions of the cold-blooded Sztumski breed, included in a genetic resources conservation program. The animals were kept in an outdoor system with a covered shelter and 24 h access to a paddock. Mares with foals were housed individually in stalls, together with their foals, and were paddocked after milking for about five hours a day throughout the year. Straw was used as bedding in the stalls, with fresh straw added daily, and the stalls were cleaned twice a week. All horses had ad libitum access to drinking water and hay year-round. Cast-iron drinkers were installed in the individual stalls, and a water trough was installed in the paddock. Mares and stallions kept outdoors received concentrate feed in the form of approximately 1.5 kg/day of crushed oats with a mineral and vitamin supplement at a dose of 100 g/day. Nursing mares received approximately 16 kg/day of concentrate feed, divided into two portions (morning and evening), along with the mineral and vitamin supplement at a dose of 100 g/day. Lactation lasted approximately eight months from the birth of the foal, and the mares were milked during the final six months.

2.2. Physicochemical Analysis of Milk

The analysis was conducted on milk collected individually from nine mares, in a volume of 40 mL from the total daily milking (mares were milked once a day, two hours after separating the foal from the mother). The milk was transported under refrigerated conditions to the laboratory and analyzed within 24 h. The contents of fat, protein, lactose, and dry matter, as well as the freezing point of the milk, were determined using the MilkoScan FT analyzer (FOSS, Hilleroed, Germany). Somatic cell count (SCC) was assessed with the Fossomatic™ FC (FOSS, Hilleroed, Germany), and the total bacterial count (TBC) was determined using the BactoScan FC bacteria analyzer (FOSS, Hilleroed, Germany).

2.3. Microbiological Analysis of Milk and Soaps, Challenge Tests

The material for the analyses included milk collected during morning milking of the mares and soaps produced from it. The soaps were prepared in cottage industry settings. In order to ensure repeatability and a high level of hygiene during the production of soaps containing mare’s milk, each batch was prepared according to the same, precisely defined recipe, maintaining identical proportions of ingredients. The raw materials were measured with high accuracy using laboratory scales, and the process of combining the components was carried out in vessels designated exclusively for cosmetic production. The soap mass was always mixed using the same equipment—manual stirrers or kitchen mixers dedicated solely to cosmetic manufacture. Both the temperature at which the ingredients were combined and the pH of the final product were monitored for each batch, and the soap mass was poured into molds certified for food contact. All batches matured under identical, strictly controlled conditions in a well-ventilated and clean room.

At every stage of production, great attention was paid to maintaining high hygiene standards. Before beginning each batch, all work surfaces, equipment, and vessels were disinfected with an ethanol solution of at least 70%. Individuals involved in the production process used disposable gloves, protective masks, and aprons to minimize the risk of product contamination. Raw materials were stored in clean, closed containers, and mare’s milk was transported and stored under refrigerated conditions, being used on the same day. Each batch was thoroughly documented—production date, composition, and production conditions were recorded. The results of microbiological tests and physicochemical measurements were archived, enabling ongoing monitoring of both the repeatability and safety of the product. This approach ensured that the production of soaps with mare’s milk was consistent and met high hygienic standards, which is crucial for the reliability of the research results and consumer safety. The soap formulation did not include synthetic preservatives. Preservation was based solely on the physicochemical properties of the soap matrix, low water activity, the presence of natural antimicrobial compounds in mare’s milk (e.g., lysozyme and lactoferrin), and ingredients such as essential oils (e.g., tea tree and lavender), which exhibit mild antimicrobial effects.

Before collecting milk samples, an assessment of the mammary gland was conducted to exclude the presence of inflammatory conditions, and the teat orifice was disinfected using tissues soaked in a disinfectant solution with a broad antibacterial spectrum. Milk was collected manually by expressing it from individual teats into sterile containers, separately for each mare. Microbiological quality control of the raw milk was carried out immediately after transportation to the laboratory. Sterile inoculation loops were used to plate the milk onto agar media enriched with 5% sheep blood to culture pathogens with high growth requirements. MacConkey agar was used as a selective medium for Gram-negative bacteria, Sabouraud agar with chloramphenicol for fungal isolation, and Edwards–Chodkowski selective differential agar for the selective isolation of streptococci. After 24 h incubation, the colonies were assessed macroscopically and microscopically. Colonies were identified using Gram staining, and catalase and coagulase testing were performed using a staphylococcus test (Biomed, Lublin, Poland) and API biochemical tests (BioMerieux Poland Ltd., Warsaw, Poland).

Before testing the soaps made from mare’s milk, their microbiological quality was assessed by determining the numbers of individual groups of microorganisms. The analyses were performed using the plate method and the automated TEMPO system (BioMerieux). Prior to the analysis, each soap sample was homogenized, and then 10 g of the material was weighed out and placed in sterile bottles containing Ringer’s solution with Tween 80. A series of ten-fold dilutions of the samples was prepared in Ringer’s solution and plated on Petri dishes with appropriate microbiological media. The material was analyzed for the following:

• Total number of aerobic mesophilic bacteria (on enriched agar), incubation for 48 h at 30 °C (BTL Sp. z o.o., Warsaw, Poland), according to PN-ISO 7889:2007 [24]
• Total number of fungi (Sabouraud agar), incubation for 5–7 days at 30 °C (BTL Sp. z o.o., Warsaw, Poland)
• Number of Escherichia coli (mFC medium), incubation for 18–24 h at 44 °C (BTL Sp. z o.o., Warsaw, Poland)
• Number of Clostridium perfringens (TSC agar with iron sulfide), incubation for 48 h at 37 °C (BioMerieux Poland Ltd., Warsaw, Poland)
• Presence of Salmonella spp. (SS and XLD agar), after pre-enrichment in buffered peptone water and selective enrichment in Rappaport–Vassiliadis medium (BTL Sp. z o.o., Warsaw, Poland) for 24 h at 37 °C. Final identification was carried out using API tests (BioMerieux Poland Ltd., Warsaw, Poland) and polyvalent sera (Biomed S.A.)

Samples were plated in triplicate. After incubation, colonies were counted using an automatic Scan 300 colony counter (Interscience Laboratories, Saint-Nom-la-Bretèche, France).

Seven soaps were selected for challenge tests. The tests were performed according to ISO 11930 [25] ‘Cosmetics—Microbiology—Evaluation of the antimicrobial protection of a cosmetic product’. Each soap sample was then contaminated with reference strains of microorganisms from the ATCC collection. Strains of Staphylococcus aureus, Pseudomonas aeruginosa, Candida albicans, and Aspergillus niger were homogenized in an inoculum prepared from physiological saline with Tween 80 to obtain a suspension with an optical density of 0.5 on the McFarland scale, corresponding to a cell concentration of 10⁶ CFU per ml of sample. The suspension was added to 20 g of soap samples placed in sterile containers. The contaminated samples were stored in a thermostat at 25 °C in the dark. Samples for further analysis were collected on day 0 (the day of inoculation) and again on days 7, 14, and 21 of incubation. They were plated on sterile microbiological TSA medium for bacteria and Sabouraud agar for fungi to determine the occurrence and growth intensity of the microorganisms. The composition of the soaps is given in

Table 1. Composition of the soaps tested in the study.

Soap No.	Basic Composition
1	Mare milk, coconut oil, cocoa butter, hemp oil, grape seed oil, karanja oil, Olive oil, tomato sauce, peloid, lavender fragrance
2	Mare milk, rice oil, coconut oil, castor oil, peloid
3	Mare milk, sweet almond oil, olive oil, cocoa butter, beeswax
4	Mare milk, coconut oil, sweet almond oil, castor oil, lavender and orange fragrance
5	Mare milk, coconut oil, pumice
6	Mare milk, rice oil, coconut oil, shea butter, birch bark tar, tea tree fragrance
7	Mare milk, stearic acid, cocoa butter, coconut oil, castor oil, bentonite clay

.

All data regarding the chemical composition, hygienic quality, and freezing point of mare’s milk presented in

Table 2. Physicochemical composition, hygienic quality, and freezing point of mare milk.

Mare No.	Fat (%)		Protein (%)		Lactose (%)		Dry Matter (%)		Freezing Point (°C)		SCC (ln)		TBC (ln)
	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD	($\overline{\mathbf{x}}$)	SD
1	0.64 a	0.08	1.63	0.13	6.80	0.33	9.77	0.34	−0.54	0.01	3.60	3.96	2.05	1.84
2	0.75	0.08	1.65	0.13	6.70	0.26	9.80	0.28	−0.54	0.01	2.15	1.48	1.65	1.64
3	0.78 b	0.11	1.75	0.08	6.37	0.09	9.60	0.14	−0.54	0.01	2.43	2.34	1.28	1.55
4	0.90	0.38	1.80	0.19	6.56	0.08	9.97	0.59	−0.54	0.01	2.03	1.90	1.03	1.11
5	0.80	0.12	1.75	0.04	6.54	0.14	9.78	0.17	−0.53	0.01	1.87	1.20	1.56	1.57
6	0.77	0.09	1.66	0.12	6.62	0.10	9.75	0.30	−0.54	0.01	2.15	1.86	2.38	3.01
7	0.72	0.07	1.63	0.12	6.71	0.29	9.76	0.35	−0.53	0.01	1.30	0.53	2.59	2.63
8	0.96	0.42	1.77	0.18	6.50	0.02	9.93	0.58	−0.54	0.01	1.79	1.30	1.97	2.21
9	0.64	0.06	1.69	0.04	6.67	0.09	9.70	0.05	−0.53	0.02	1.79	2.26	2.36	3.07
Mean	0.77	0.22	1.70	0.13	6.61	0.21	9.79	0.34	−0.53	0.01	10.81	19.72	7.26	11.28

Abbreviations: a, b—means with different superscripts within a row are significantly different at p < 0.05; SD—standard deviation; SCC—somatic cell count; TBC—total bacterial count.

were subjected to statistical analysis using Statistica software (StatSoft, Krakow, Poland), version 13.3. For each parameter (dry matter, fat, protein, lactose, freezing point, somatic cell count, and total bacterial count), mean values ($\overline{\mathbf{x}}$) and standard deviations (SDs) were calculated. Somatic cell count and TBC were transformed using the natural logarithm (ln).

Before performing analysis of variance (ANOVA), the assumption of homogeneity of variances was verified using Levene’s test. Confirmation of this assumption enabled the use of one-way ANOVA. In cases where significant overall differences were found (p < 0.05), Tukey’s post hoc test was conducted to identify statistically significant differences between samples from individual mares.

Differences were considered statistically significant at p < 0.05. All calculations were performed in accordance with standard procedures for reporting laboratory research results.

3. Results

The dry matter content in milk from the Sztumski mares did not exceed 10%, ranging from 9.70% to 9.93%. The fat content did not exceed 1%, ranging from 0.64% to 0.96%. The average protein content in the milk from the mares was 1.70% (ranging from 1.63% to 1.77%). The average lactose concentration was 6.61% (6.37–6.80%). The freezing point of the milk averaged −0.5 °C, with a range from −0.53 °C to −0.54 °C. The somatic cell count (SCC) ranged from 1.30 to 3.60/mL, and the total bacterial count (TBC) from 1.03 to 2.59/mL. Only the fat content in mare’s milk showed statistically significant differences (p < 0.05), observed between mare no. 1 and mare no. 3. Values are presented in columns, and different superscripts (a, b) denote statistically significant differences. No additional grouping was identified (Table 2).

The results pertaining to the microbiological quality of mare milk and the soaps produced from it indicate very low levels of microbial contamination in the samples. In four milk samples (nos. 3, 5, 6, and 9), no microbial growth was observed, while the remaining samples contained single colonies of streptococci, such as Streptococcus agalactiae (sample no. 1) or coagulase-negative streptococci (samples 2, 7, and 8). An exception was sample no. 8, in which Staphylococcus aureus was also detected (

Table 3. Results of microbiological testing of mare milk.

Milk Sample No.	Result of Microbiological Testing
1	Single colonies of streptococci—Streptococcus agalactiae
2	Single colonies of coagulase-negative Streptococcus (CNS)
3	ng
4	Single colonies of Streptococcus faecalis
5	ng
6	ng
7	Single colonies of coagulase-negative Streptococcus (CNS)
8	Single colonies of coagulase-negative Streptococcus (CNS) and Staphylococcus aureus
9	ng

Abbreviations: ng—no growth.

).

The results of the microbiological testing of the mare milk soaps were varied. The soaps produced from mare milk generally exhibited excellent microbiological quality. Most samples showed no microbial growth, confirming effective decontamination and preservation during processing. However, in four samples (nos. 2, 4, 5, and 7), single colonies of Bacillus spp. were observed, indicating minor environmental contamination likely related to the ubiquity of these bacteria (

Table 4. Results of testing for microbiological contamination of soaps made from mare milk.

Soap	Result of Microbiological Testing
1	ng
2	Single colonies of Bacillus spp.
3	ng
4	Single colonies of Bacillus spp.
5	Single colonies of Bacillus spp.
6	ng
7	Single colonies of Bacillus spp.

Abbreviations: ng—no growth.

).

The challenge tests conducted to evaluate the resistance of the soaps to microbial contamination indicated varied results. Four soap samples (nos. 1, 3, 4, and 5) demonstrated complete inhibition of microbial growth over the 28-day observation period, confirming their high antimicrobial stability. In samples 2 and 7, growth of Aspergillus niger and Staphylococcus aureus was observed (

Table 5. Challenge tests of soaps made from mare milk.

Soap	Result of Microbiological Testing
	Day 7	Day 14	Day 28
1	ng	ng	ng
2	Single colonies of Aspergillus niger	Growth of Aspergillus niger	Growth of Aspergillus niger
3	ng	ng	ng
4	ng	ng	ng
5	ng	ng	ng
6	Single colonies of Staphylococcus aureus	Abundant growth of Staphylococcus aureus	Abundant growth of Staphylococcus aureus
7	Single colonies of Staphylococcus aureus	Abundant growth of Staphylococcus aureus	Abundant growth of Staphylococcus aureus

Abbreviations: ng—no growth.

).

4. Discussion

The results obtained in this study indicate that milk from Sztumski mares is characterized by favorable physicochemical parameters, which are comparable to or even slightly better than those reported for other breeds covered by conservation programs. The average dry matter content (9.79%) was higher than in milk from Sokolski mares (9.26%) and Polish Half-Bred mares (9.05%) [17]. Similarly, the fat content (0.77%) exceeded the values noted for Sokolski mares (0.33%) and hot-blooded mares (0.40%). In contrast, the average protein content (1.70%) was slightly lower than that in Sokolski (1.90%) and hot-blooded (1.86%) mares, while the lactose concentration was similar to that of Sokolski mares (6.70%) and higher than in hot-blooded mares (6.48%) [17].

Comparable studies on native Kyrgyz horses by Mazhitova and Kulmyrzaev [26] reported even higher levels of milk constituents (dry matter up to 11.76%, protein up to 2.39%, and fat up to 1.90%). These variations underscore the influence of breed and environmental factors on milk composition. Cais-Sokolińska et al. [21], studying Sokolski mares, observed a progressive increase in the freezing point during lactation (from −0.573 °C to −0.556 °C), which was slightly lower than the values noted in the present study. They also reported an SCC of 27,200/mL and a TBC of 9600/mL. In comparison, the milk analyzed in this study had a lower average SCC (10,800/mL) and a similar TBC (11,300/mL), reflecting its high hygienic quality.

Regarding microbiological quality, most milk samples showed either ng or only isolated colonies of streptococci. The detection of Streptococcus agalactiae and coagulase-negative streptococci (CNS) in individual samples suggests occasional minor contamination, which may occur despite the antibacterial properties attributed to mare milk. The presence of Staphylococcus aureus in one sample highlights the need for maintaining strict hygiene during milking, although the low overall microbial counts indicate that such incidents were sporadic. Similar conclusions were drawn by Hazeleger and Beumer [27], who found fecal bacteria in mare milk in the Netherlands and Belgium but did not detect Salmonella, Listeria, Campylobacter, or typical mastitis pathogens.

Apart from its nutritional value, mare milk has been regarded since ancient times in parts of the former Soviet Union and East Asia as a type of natural medicine due to its reputed health-promoting properties [28]. While some studies have highlighted its antimicrobial and anti-inflammatory activity, it is difficult to find clinical trials confirming these benefits. Nevertheless, recent reports suggest that the consumption of mare milk may help alleviate intestinal disorders, cow milk intolerance, or skin conditions such as eczema and psoriasis [29].

Nurliyani [30] demonstrated that, due to the presence of antimicrobial compounds, mare milk can be stored for 16 h at room temperature without an increase in the total bacterial count (remaining at 1.4 × 10⁶ cfu/mL). This is consistent with the generally high microbiological stability observed in the present study. The literature also indicates that mare milk is rich in lysozyme and lactoferrin, which contribute to limiting bacterial growth [12].

Little is known about mastitis in mares, as udder infections occur only sporadically. Mete [31] found that Streptococcus equi subsp. zooepidemicus (54.6%) and Escherichia coli (27.4%) were the most common pathogens isolated from quarter milk, while Kozdrowski et al. [32] demonstrated the involvement of S. equi subsp. zooepidemicus and S. equi subsp. equi in spontaneous autoamputation of a diseased section of the mammary gland. In the present study, fungi were not isolated, and the most frequently detected bacteria were CNS, confirming the generally low risk of mastitis and high microbiological quality.

The soaps produced from mare milk generally exhibited excellent microbiological quality. Most samples showed ng, demonstrating effective decontamination and preservation during processing. The detection of Bacillus spp. in three samples is likely linked to environmental contamination, as these bacteria are ubiquitous in soil and air, and low-level presence is not typically a health concern. The results of the challenge tests revealed differences in the resistance of the soaps to intentional microbial contamination. Four soap samples (nos. 1, 3, 4, and 5) showed complete inhibition of microbial growth over 28 days, confirming their antimicrobial stability. However, the growth of Aspergillus niger and Staphylococcus aureus in samples 2 and 7 suggests that the natural preservation system used in these formulations may have limited efficacy against certain microorganisms or under specific storage conditions. Notably, the soaps were formulated without synthetic preservatives, relying instead on the antimicrobial properties of mare’s milk (e.g., lactoferrin and lysozyme) and essential oils such as lavender and tea tree oil. This highlights the need for improved stabilization strategies in future formulations, possibly through the inclusion of certified natural preservatives with broader antimicrobial activity. This underlines the importance of further optimizing preservation systems, especially when formulating natural products based on mare’s milk.

The observations made here align with the findings of Jastrzębska et al. [7], who emphasized that, due to its nutritional and anti-inflammatory properties, mare milk is increasingly used in cosmetics, where it helps moisturize deeper skin layers and exerts antioxidant activity, thus slowing aging processes [22,23]. The overall low contamination levels in the soaps produced in this study, along with their demonstrated ability to inhibit microbial growth in challenge tests, support the view that such products are safe and of high hygienic quality.

5. Conclusions

The conducted study confirmed the high hygienic quality of Sztumski mare’s milk, characterized by a stable composition and low somatic cell and bacterial counts. Although occasional presence of pathogens such as Streptococcus agalactiae, coagulase-negative staphylococci (CNS), and Staphylococcus aureus was observed, the overall microbiological profile supports the suitability of this milk for both consumption and cosmetic applications. Soaps containing mare’s milk demonstrated microbiological stability in most samples; however, the detection of Aspergillus niger and Staphylococcus aureus in some products, along with limited antimicrobial efficacy observed in challenge tests, indicates the need for further optimization of soap formulation and preservation methods. These findings highlight the potential of mare’s milk as a valuable ingredient in skincare products, while emphasizing the importance of establishing optimal production and storage conditions to ensure product safety. Future research should focus on detailed evaluation of the functional properties of mare milk soaps, including their skincare and antimicrobial effects, and developing strategies to enhance the durability and microbiological safety of finished cosmetic products based on mare’s milk.