BIOCHEMICAL CHARACTERIZATION OF PROTEIN CONTENT IN Pleurotus ostreatus CULTIVATED UNDER LABORATORY CONDITIONS

ABSTRACT

Pleurotus ostreatus is one of the mushroom species with high nutritional value, rich in proteins, amino acids, vitamins, and minerals. In this study, an effective method for cultivating Pleurotus ostreatus under laboratory conditions was developed, and its protein content was analyzed using biochemical methods. The amount of water-soluble protein in the mushroom samples was determined by the Lowry method, while total protein content was determined using the Kjeldahl method. Additionally, protein fractions were analyzed by high-performance liquid chromatography (HPLC). The results showed that total protein content ranged from 28.95% to 31.19%, while water-soluble protein accounted for 8.1%. HPLC analysis revealed various protein and peptide fractions in the mushroom, with more than 90 peaks recorded. The obtained results confirm that Pleurotus ostreatus is a protein source with high biological value and indicate its potential applications in the food and biotechnology sectors.

АННОТАЦИЯ

Pleurotus ostreatus является одним из видов грибов с высокой пищевой ценностью, богатых белками, аминокислотами, витаминами и минеральными веществами. В данном исследовании была разработана эффективная методика культивирования Pleurotus ostreatus в лабораторных условиях, а также проведен биохимический анализ содержания белка. Количество водорастворимого белка в образцах грибов определяли методом Лоури, тогда как общий белок определяли методом Кьельдаля. Кроме того, фракционный состав белков был исследован с использованием высокоэффективной жидкостной хроматографии (ВЭЖХ). Полученные результаты показали, что общее содержание белка варьировало в пределах от 28,95% до 31,19%, тогда как доля водорастворимого белка составляла около 8,1%. Анализ ВЭЖХ выявил наличие различных белковых и пептидных фракций в образцах гриба, при этом было зарегистрировано более 90 хроматографических пиков. Полученные данные подтверждают, что Pleurotus ostreatus является ценным источником белка с высокой биологической ценностью и свидетельствуют о перспективности его применения в пищевой промышленности и биотехнологии. В частности, результаты могут быть использованы при разработке функциональных продуктов питания и биотехнологических процессов.

Keywords: Pleurotus ostreatus, protein analysis, Kjeldahl method, Lowry method, HPLC, biotechnology.

Ключевые слова: Pleurotus ostreatus, анализ белка, метод Кьельдаля, метод Лоури, ВЭЖХ, биотехнология.

Introduction. Currently, food security issues and the search for alternative protein sources are among the important directions of global scientific research. Due to the increasing population and shortage of traditional protein sources, the need to explore new biological sources is growing [1]. In this context, mushrooms, especially Pleurotus ostreatus, are considered a promising biological resource with high nutritional value. Pleurotus ostreatus is one of the easily cultivable, fast-growing, and highly productive mushrooms, containing essential amino acids, vitamins (B group), and minerals [2]. Studies have shown that the protein content in this mushroom exceeds that of many plant sources [3]. The Lowry and Kjeldahl methods were jointly used to determine the protein content in Pleurotus ostreatus because each method has advantages in measuring different aspects. The Lowry method is a sensitive and rapid colorimetric method for detecting water-soluble proteins, allowing the evaluation of protein fractions’ bioactivity and metabolic availability [4]. Meanwhile, the Kjeldahl method, based on total nitrogen content, accounts for all protein components, including both soluble and insoluble proteins, and reliably determines total protein content [5]. Therefore, the combined use of these methods allows a qualitative and quantitative assessment of proteins in mushrooms, enhancing the scientific reliability of the results. Furthermore, high-performance liquid chromatography (HPLC) enables the determination of protein and peptide fractions’ composition and relative amounts [6]. Thus, the aim of this study was to cultivate Pleurotus ostreatus under conditions suitable for modern biotechnology laboratories and to isolate its proteins in pure form without additives. Due to the high protein content, Pleurotus ostreatus is widely cultivated today. However, isolating its proteins in pure form, without additives and efficiently, remains an important scientific and technological challenge. Therefore, this study aimed to develop methods for laboratory cultivation and high-purity protein isolation. This approach serves to comprehensively study Pleurotus ostreatus as a protein source and expand its applications in the food industry and biotechnology.

Materials and Methods

Pleurotus ostreatus was cultivated under laboratory conditions on a sterile substrate. The substrate was prepared from wheat straw (Triticum aestivum) plant residues and sterilized in an autoclave at 121°C for 30 minutes. After sterilization, mushroom mycelium was added to the substrate and incubated at 25°C. The mushroom was grown for 21 days, and the resulting biomass was collected for analysis. Only one Pleurotus ostreatus sample was used in the study, as the aim was to cultivate this mushroom under laboratory conditions and determine its protein content in pure form. Comparisons with other mushrooms were not included, making a single sample sufficient. Protein Analysis: Water-soluble protein content was determined using the Lowry method, and a calibration curve was prepared based on albumin standard solution. Measurements were taken using a spectrophotometer at an optical density of 750 nm. Total protein content was determined by the Kjeldahl method, and nitrogen content was calculated using the following formula:

Figure 1. Calibration curve for protein determination by the Lowry method

HPLC Analysis: Protein fractions were analyzed by HPLC using a C18 column. The mobile phase consisted of a mixture of water and organic solvents, and detection was performed using a UV detector. The combined application of Lowry and Kjeldahl methods enabled a qualitative and quantitative assessment of proteins.

Figure 2. HPLC chromatogram of protein and peptide fractions in Pleurotus ostreatus

Results and Discussion.

Total protein content in Pleurotus ostreatus determined by the Kjeldahl method ranged from 28.95% to 31.19%. According to the Lowry method, water-soluble protein content was 8.1%. HPLC analysis revealed numerous protein and peptide fractions, with over 90 peaks recorded. The highest intensity peaks were observed at retention times of 11.192 min (11.51%), 13.952 min (11.25%), and 35.732 min (7.09%). The obtained results were consistent with other studies. For instance, research by Sharma et al. reported that protein content in Pleurotus ostreatus ranged from 25–30%, which aligns with our findings [3]. Similarly, studies by Barros et al. highlighted the high content of bioactive compounds in this mushroom [1]. The protein composition of Pleurotus ostreatus confirms its high biological value, enabling its use in the production of functional food products.

Conclusion. The study results indicate that Pleurotus ostreatus can be effectively cultivated under laboratory conditions. Biochemical analyses confirmed a high protein content, with total protein ranging from 28.95% to 31.19% and water-soluble protein at 8.1%. HPLC analysis verified the presence of various protein and peptide fractions. These results demonstrate that Pleurotus ostreatus is a protein source with high biological value, with significant potential applications in the food industry and biotechnology.

The main novelty of this study is that we successfully cultivated a single Pleurotus ostreatus sample under laboratory conditions and determined its protein in pure form without additives. Until now, other researchers have not been able to isolate proteins from Pleurotus ostreatus under laboratory conditions with such purity and quality. This approach allows the future production of high-purity proteins from mushrooms, their use in modern biotechnology, and the identification of various carbohydrate fractions. Moreover, the study opens new prospects for producing Pleurotus ostreatus as a high-biological-value protein source and enhancing its functional properties.

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