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13БИОЛОГИЧЕСКИЕ НАУКИ
UDC 663.8+616.155.16
Partsei Kh. Yu.
Assistant of the Department of Biological and Medical Chemistry
named after G. O. Babenka Ivano-Frankivsk National Medical University Ivano-Frankivsk, Ukraine Erstenyuk H. M. Doctor of Biological Sciences, Professor Department of Biological and Medical Chemistry named after G.O. Babenka
Ivano-Frankivsk National Medical University Ivano-Frankivsk, Ukraine Kindrat I.P.
Candidate of Biological Science, PhD, Assistant of the Department of Biological and Medical Chemistry named after G.O. Babenka
Ivano-Frankivsk National Medical University Ivano-Frankivsk, Ukraine
EFFECT OF ENERGY DRINK ON LIGAND FORMS OF HEMOGLOBIN
Парцей Христина Юрьевна
Ассистент кафедры биологической и медицинской химии имени Г.А. Бабенко Ивано-Франковского национального медицинского университета
Ивано-Франковск, Украина Эрстенюк Анна Михайловна Доктор биологических наук, профессор кафедры биологической и медицинской химии имени Г.А. Бабенко Ивано-Франковского национального медицинского университета
Ивано-Франковск, Украина Киндрат Ирина Петровна Кандидат биологических наук, PhD, ассистент кафедры биологической и медицинской химии имени Г. А. Бабенко Ивано-Франковского национального медицинского университета
Ивано-Франковск, Украина
ВЛИЯНИЕ ЭНЕРГОНАПИТКА НА ЛИГАНДНЫЕ ФОРМЫ ГЕМОГЛОБИНА
DOI: 10.31618/ESSA.2782-1994.2022.3.77.246
Summary. The article is devoted to the study of the effect of energy drink on ligand forms of hemoglobin in experimental animals. According to our results, the consumption of energy drink causes significant changes in both the total level of hemoglobin and its ligand forms. In particular, we have established a reduction of oxyhemoglobin and accumulation of dyshemoglobin: methemoglobin, sulfhemoglobin and carboxyhemoglobin, which in turn affects the oxygen homeostasis of the body, the development of tissue hypoxia and can cause structural and functional disorders in the body under such conditions.
Аннотация. Статья посвящена изучению влияния энергонапитка на лигандные формы гемоглобина экспериментальных животных. Как свидетельствуют полученные нами результаты, потребление энергетического напитка приводит к существенным изменениям как общего уровня гемоглобина, так и его лигандных форм. В частности, нами установлено снижение уровня оксигемоглобина и накопление дисгемоглобинов: метгемоглобина, сульфгемоглобина и карбоксигемоглобина, что, в свою очередь, влияет на кислородный гомеостаз организма, развитие тканевой гипоксии и может служить причиной структурно-функционального нарушений в организме при таких условиях.
Key words: energy drink, laboratory rats, hemoglobin, oxyhemoglobin, carboxyhemoglobin, sulfhemoglobin, methemoglobin.
Ключевые слова: энергетический напиток, лабораторные крысы, гемоглобин, оксигемоглобин, карбоксигемоглобин, сульфгемоглобин, метгемоглобин.
INTRODUCTION. The consumption of energy drinks, which are characterized by high content of caffeine, taurine and carbohydrates with guarana, ginseng, B vitamins among adolescents, is growing every year, as they are designed to stimulate the central
nervous system (CNS) and energy metabolism [6,8,13]. Children and adolescents who do not consume caffeine may be prone to intoxication due to lack of pharmacological tolerance [9,14]. Of no less concern
are the combined effects of caffeine with other components of the energy drink in the adult population.
Hemoglobin plays an important role in the body's adaptation to toxic effects, as it provides oxygen transport function and participates in the regulation of acid-base balance [11]. Xenobiotics, by binding to hemoglobin, disrupt the stability of the structure of not only this hemoprotein but also erythrocytes, block the transport of oxygen to the tissues and cause hypoxia. [7].
Based on this, it is important to study the level of total hemoglobin and its ligand forms, in particular: oxy-, carboxy-, sulf- and methemoglobin under the conditions of energy drink consumption.
MATERIALS AND METHODS. The study was conducted using male Wistar rats, which were kept in the vivarium under appropriate lighting conditions, temperature, humidity and standard diet. All animals had free access to feed (based on daily requirements) and water (based on 20 ml of water per rat per day). The amount of standard feed consumed for laboratory animals was determined by its residue in the feeder. Control over the growth and development of animals was performed by weighing them at the beginning and at the end of the experiments. The experimental animals were divided into four groups: 1st group - received drinking water (control group); 2nd group - received an energy drink for a month and the collection of material was carried out on the 10th day at the end of the experiment; 3rd group - received an energy drink for a month and the collection of material was carried out on the 20th day at the end of the experiment; 4th group -received an energy drink for a month and the collection
of material was carried out on the 30th day at the end of the experiment.
Study of hemogram parameters: determination of total hemoglobin concentration and absolute erythrocyte content was performed using a hematology analyzer «MYTHIC 18». Determination of the content of oxyhemoglobin, methemoglobin, sulfhemoglobin and carboxyhemoglobin was performed according to the methods described by Sukhomlinov B.F. [3].
All animal experiments were carried out in compliance with the requirements of the European Convention for the Protection of Vertebrate Animals used for Experimental and Scientific Purposes (Strasbourg, 1986), Law of Ukraine № 3447-IV "On Protection of Animals from Cruelty", adopted by the Parliament in February 21, 2006 in a new wording. According to the Article 26 of the Rules for the Treatment of Animals Used in Scientific Experiments, Testing, Educational Process, Production of Biological Products, as well as the recommendations of the First National Congress of Ukraine on Bioethics (Kyiv, Ukraine, 2001).
The results obtained were subjected to statistical analysis by conventional methods [1] using the Student's /-test (Statstica 8).
RESULTS AND DISCUSSION. As a result of our researche, we found that in the blood of rats that consumed energy drink during the month on the 10th and 20th day after the experiment there was a decrease in erythrocytes by 17 % and 14 %, respectively, and an increase on the 30th day by 7 % compared to control animals (Fig.1).
£ (N
9,0 8,0 7,0 6,0 5,0
S 4,0
3,0 2,0 1,0 0,0
#
# T
M control 0 10 day 0 20 day □ 30 day
RBC
Fig. 1. Absolute erythrocyte content under the influence of energy drink. Note: * - p<0.001, # - p<0.05 - reliability compared to the control group of animals.
A decrease in the number of erythrocytes on the 10th and 20th day after the end of the experiment may indicate a violation of hematopoietic processes in these
animals or hemolysis of erythrocytes or impaired hemoglobin synthesis.
At the same time, studies of total hemoglobin in the blood of rats consuming energy drink (Fig. 2)
showed an increase on the 10th and 30th day after the end of the experiment by 8 % and 6 %, respectively, and a decrease on 19 % on the 20th day compared with an intact group of animals. The reasons for the decrease in hemoglobin can be considered as a violation of biosynthesis and enhanced breakdown of this hemoprotein. The increase of this indicator may be the
result of an adaptive response of animals to the consumption of energy drink. Based on the obtained results and based on data from the scientific literature [5], it is important to study the ligand forms of hemoglobin to understand the biochemical mechanisms of adaptation to the influence of various factors.
160,0 140,0 120,0 100,0
i 80,0 60,0
40,0
20,0
0,0
#
m
#
M control El 10 day m 20 day □ 30 day
HGB
Fig. 2. The level of total hemoglobin under the influence of energy drink. Note: * - p<0.001, # - p<0.05 - reliability compared to the control group of animals.
To assess the supply of oxygen to tissues, the indicator of the dynamics of changes in the content of both total hemoglobin and oxyhemoglobin (HbÜ2) is informative. Studies have shown that the level of HbÜ2 decreased in the experimental groups by 1.5, 1.6, and 1.8 times, respectively, compared with the control group (Fig.3).
The decrease in oxyhemoglobin may be due to a number of factors: changes in the morphofunctional state of erythrocytes, which is confirmed by analysis of acid erythrograms of rats, which we presented earlier
[2]; violation of the structure of hemoglobin and the affinity of hemoglobin for oxygen. The combination of such factors has a direct impact on the level of the active form of hemoglobin, which in turn affects the oxygen homeostasis of the body. The level of HbO2 is largely determined by the presence of dyshemoglobin, such as methemoglobin, sulfhemoglobin, carboxyhemoglobin, which prevents the saturation of hemoglobin with oxygen, reducing the level of oxyhemoglobin in transported blood and promote tissue hypoxia [7].
140,0
120,0
100,0
*
80,0
60,0
40,0
20,0
0,0
* T
Ш control EH 10 day 0 20 day □ 30 day
HbO2
Fig. 3. The level of oxyhemoglobin under the influence of energy drink. Note: * - p<0.001 - reliability compared to the control group of animals.
The results of the study of the content of the experimental groups in 3.9, 4.2 and 3.8 times,
methemoglobin under the influence of energy drink consumption showed an increase in this derivative in
respectively, on the 10th, 20th and 30th day compared with the intact group (Fig.4).
4,0 3,5 3,0 2,5
1,5 1,0 °,5 0,0
ш control
□ 10 day 0 20 day
□ 30 day
MetHb
*
Fig. 4. Methemoglobin level under the influence of energy drink. Note: * - p<0.001 - reliability compared to the control group of animals.
The formation of methemoglobin in erythrocytes is a constant oxidative process, which is the result of exposure to hemoglobin of various highly reactive molecules (oxygen free radicals) formed during normal cellular metabolism [10].
Based on the obtained data, it is possible to make assumptions about the activation of the protective function of methemoglobin under the conditions of energy drink consumption, as well as the high intensity of oxidative processes in erythrocytes, which are accompanied by the accumulation of superoxide anion and hydrogen peroxide as a result of degradation of hemoglobin and peroxidative modifications of lipids.
Carboxyhemoglobin (HbCO) is a complex formed in erythrocytes under the influence of carbon monoxide. At the same time there is a violation of the ability of oxygen to bind to hemoglobin, the development of hypoxia, which in turn leads to disruption of cellular metabolism and causes the development of various pathological conditions [11].
Figure 5 presents the results of the study of HbCO levels, which indicate a significant increase in this indicator on the 20th and 30th day in 1.8 and 1.7 times, respectively, compared with the control group.
10,0 9,0 8,0 7,0 6,0
Ii 5,0
4,0 3,0 2,0 1,0 0,0
Ш control EU 10 day 0 20 day 0 30 day
HbCO
Fig. 5. Carboxyhemoglobin level under the influence of energy drink. Note: * - р<0.001 - reliability compared to the control group of animals.
Sulfhemoglobin (SHb) is an important ligand of hemoglobin, which is formed due to the irreversible oxidation of hemoglobin, by breaking the methine bridge in the structure of heme and the release of iron ions (Fe2+ Ta Fe3+) [4]. The study of this form of hemoglobin, under the conditions of energy drink consumption, is important because it will determine the level of hemoglobin damage.
The results of the study of the level of sulfhemoglobin in rats that consumed energy drink during the month, on the 10th, 20th and 30th day after the end of the experiment showed an increase in 1.4, 1.5 and 1.37 times, respectively, compared with control group (Fig. 6).
0,8 0,7 0,6 0,5 1*0,4 0,3 0,2 0,1 0,0
Ш control El 10 day 0 20 day 0 30 day
SHb
Fig. 6. The level of sulfhemoglobin under the influence of energy drink. Note: * - p<0.001 - reliability compared to the control group of animals.
High concentrations of sulfhemoglobin in erythrocytes can lead to the accumulation of free iron ions, which in turn act as powerful initiators of free radical processes in cells.
CONCLUSIONS. In accordance with our results, the consumption of energy drink causes significant changes in both total hemoglobin and ligand forms. In particular, we have established a reduction of oxyhemoglobin and accumulation of dyshemoglobin: methemoglobin, sulfhemoglobin and
carboxyhemoglobin, which in turn affects the oxygen homeostasis of the body, the development of tissue hypoxia and may cause structural and functional disorders in the body under such conditions.
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Скиба O.I.
Тернопшьський нацюнальний медичний yuieepcumem iMem 1.Я.Горбачевського,
Украша, 46001, м. Тернопшь, майдан Волi
Грубтко В.В.
Тернотльський нацюнальний педагогiчний утверситет iMeHi Володимира Гнатюка,
Украша, 46027, м. Тернопшь, вул. М. Кривоноса, 2
Гуменюк В.В.
Тернотльський нацiональний педагогiчний утверситет iменi Володимира Гнатюка,
Украша, 46027, м. Тернопшь, вул. М. Кривоноса, 2
ПРОБЛЕМА ВОДОЗАБ1РНИХ ДЖЕРЕЛ ЯК НАСЛ1ДОК ЕВТРОФ1КАЦП I МАЛОВОДДЯ ТА ЗАБРУДНЕННЯ КОМУНАЛЬНОГО ТА СШЬСЬКОГОСПОДАРСЬКОГО ПОХОДЖЕННЯ
O.I. Skyba
I. Horbachevsky Ternopil National Medical University, Ukraine, 46001, Ternopil, Voli Square V.V. Hrubinko
Ternopil Volodymyr Hnatiuk National Pedagogical University, Ukraine, 46027, Ternopil, Maksyma Kryvonosa St., 2
V. V.Humeniuk
Ternopil Volodymyr Hnatiuk National Pedagogical University, Ukraine, 46027, Ternopil, Maksyma Kryvonosa St., 2
