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VOLUME 04 ISSUE 01 PAGES: 102-112 

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ABSTRACT 

Research was undertaken to examine the amino acid and elemental composition of “Hyposedaf”, a dry extract known 

for its hypotensive properties. Quantitative analyses of both micro and macro elements were conducted using 

Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Additionally, the amino acid content was quantitatively 

analyzed through High-Performance Liquid Chromatography (HPLC). This study identified 20 different amino acids in 

the “Hyposedaf” dry extract, 10 of which are essential. Notable amino acids, such as histidine, cysteine, tyrosine, 

methionine, valine, asparagine, and leucine, amounted to a total of 29.15 mg. Of all detected amino acids, 46.5% were 

classified as non-essential, while 53.5% were essential. The elemental analysis of the “Hyposedaf” dry extract revealed 

61 different elements, including 7 essential elements (Co, Cr, Cu, Fe, Mn, Se, Zn) and 4 conditionally necessary 

elements. Key elements like potassium, calcium, sodium, zinc, iron, magnesium, and phosphorus, found in substantial 

amounts, are crucial for human health. The concentrations of heavy metals and arsenic in the dry extract were within 

safe limits according to the Sanitary-epidemiological rules and norms, complying with the quantitative standards State 

Pharmacopoeia of the Republic of Uzbekistan (Volume 1, Issue 1) and State Pharmacopoeia of the Russian Federation 

XIV (RF) [1,2,3]. The findings indicate that “Hyposedaf” dry extract possesses significant value, offering potential for 

medical application and the development of other medicinal forms. 

  Research Article 

 

“HYPOSEDAF” DRY EXTRACT’S AMINO ACID AND ELEMENTAL 

COMPOSITION STUDY 
 

Submission Date: January 21, 2024, Accepted Date:  January 26, 2024,  

Published Date: January 31, 2024  

Crossref doi: https://doi.org/10.37547/ajbspi/Volume04Issue01-16 

 

 

Safarova Diyora Tolibovna 
Assistant Of The Tashkent Pharmaceutical Institute, Uzbekistan 

 

Maksudova Firuza Khurshidovna 
Tashkent Pharmaceutical Institute, Doctor Of Pharmaceutical Sciences, Associate Professor, Uzbekistan 

 

Karaeva Nargizakhan Yuldash Qizi 
Assistant Of The Tashkent Pharmaceutical Institute, Uzbekistan 

Journal Website: 

https://theusajournals.

com/index.php/ajbspi 

Copyright: Original 

content from this work 

may be used under the 

terms of the creative 

commons attributes 

4.0 licence. 

 

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KEYWORDS 

Dry extract, micro and macro elements, amino acids, HPLC method, mass spectrometry. 

INTRODUCTION

Extensive research has established the critical role of 

minerals and amino acids in the human body, acting as 

fundamental nutrients for synthesizing all proteins. 

Besides contributing to brain function, including 

intellectual activity, motivation, and general mental 

state, amino acids play a vital role in maintaining overall 

well-being. These nutrients are key components of all 

human tissues, such as muscles. Amino acids and 

chemical elements not only exhibit specific 

pharmacological effects but also demonstrate 

synergism with various substances, paving the way for 

the development of combined drugs with multiple 

functional properties derived from plants. Importantly, 

macro and micronutrients, as well as amino acids 

sourced from plants, are more effectively absorbed by 

the human body, owing to their “biological” quantities 

present in plants. Furthermore, amino acids assist in 

the optimal performance of mineral substances within 

the human body. The study of amino acid and mineral 

compositions in medicinal plants and 

phytopreparations is vital for the creation of new 

pharmaceutical drugs [4]. 

Currently, it's essential to understand the trace 

element content in the entire body of medicinal plants, 

including their separate generative and vegetative 

organs - roots, stems, leaves, buds, flowers, fruits, and 

seeds. This knowledge significantly expands the scope 

of their utilization. In plants, bioelements are found in 

forms that are absorbable by the human body, such as 

amino acids, vitamins, proteins, etc., and they are 

present in complex forms. This fact underscores the 

importance of medicinal plants as sources of 

biologically active substances and elements. The 

investigation of the elemental composition of 

medicinal plants and their based preparations is not 

only necessary for determining their value but also for 

preventing toxic and carcinogenic properties due to 

increased levels of heavy metals and arsenic [5,6]. 

Considering the above, the feasibility of using 

medicinal plant raw materials or extracts based on 

them in healthcare practice involves studying their 

amino acid and elemental composition. Amino acids 

are crucial for plant development, occurring during 

photosynthesis and participating in a wide range of 

biochemical reactions that support optimal growth 

and development [7,8,9]. 

Object of the Study: The aim of this study is to 

investigate the amino acid and elemental composition 

of the dry extract of “Hyposedaf”. 

 

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VOLUME 04 ISSUE 01 PAGES: 102-112 

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Experimental Part: The research focused on the dry 

extract of “Hyposedaf”, conditionally named and 

derived from various plant raw materials, including 

peppermint leaves (Folia Menthae piperitae L.), 

Turkistan lion's tail grass (Herba Leonuri turkistanicae), 

dark-red hawthorn fruits (Fructus Crataegi sanguineae 

Pall), and field horsetail grass (Herba Equiseti arvensis). 

This extract complied with the specifications outlined 

in the publications State Pharmacopoeia of the Russian 

Federation and XIV State Pharmacopoeia (RF) [10,11]. 

In the first phase of our research, we concentrated on 

the elemental composition of “Hyposedaf” analyzing 

its macro and microelements through Inductively 

Coupled Plasma Mass Spectrometry (ICP MS) as per 

the methodological guidelines [12]. 

In the second phase, the amino acid composition of the 

“Hyposedaf” dry extract was examined. This involved 

isolating free amino acids and conducting their 

quantitative analysis by comparing the retention times 

and peak areas of a standard sample with those of 

amino acids obtained from phenylthiocarbamyl. 

Results and Discussion: The “Hyposedaf” dry extract 

was analyzed using the highly sensitive Inductively 

Coupled Plasma Mass Spectrometry (ICP-MS) method. 

This technique enabled the simultaneous 

determination of numerous elements in a single 

sample with very low detection limits. The mass-

spectral analysis was developed in Russia and validated 

in Uzbekistan (MBI) UzO`U 0677:2015 (MBI №499-

AEM/MS), facilitating the determination of 61 elements 

(Na, Mg, Al, P, K, Ca, Ti, Mn, Fe, Li, Be, B, Sc, V, Cr, Co, 

Ni, Cu, Zn, Ga, As, Se, Rb, Sr, Y, Zr, Nb, Mo, Ag, Cd, In, 

Sn, Sb, Te, Cs, Ba, La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, 

Er, Tm, Yb, Lu, Hf, Ta, W, Re, Pt, Au, Tl, Pb, Bi, Th, U). 

This method utilized argon inductively coupled plasma 

as an ion source. The test sample, weighing 0.1 g, was 

introduced into the plasma in solution form. Prior to 

measurement, the sample underwent solubilization. 

The resulting solution, containing the test sample, was 

aerosolized with a stream of argon and introduced into 

the plasma. The core temperature of the plasma 

reached up to 9000°C, at which the sample was 

desiccated, and the molecular elements transitioned to 

an atomic state, becoming ionized to form positive 

ions. These positive ions, along with electrons and 

neutral particles, were channeled through cone-

shaped interfaces into the vacuum segment of the 

analyzer, passing through several electric ion lenses 

that filtered out electrons and neutral particles from 

the ions. 

The test sample and a standard sample, each weighing 

0.1 g, were placed in Teflon bags. Decomposition of the 

test, standard, and control samples was performed in 

a system involving Teflon bags with lids (Hot Block, 

Environmental Express). The Hot Block, consisting of 

36 or 40 cells depending on the model, allowed for the 

heating of dissected samples in large volumes. This 

decomposition process utilized a mixture of HF, HClO4, 

and HNO3 acids in an open system to fully dissolve the 

test sample. After removing the bags from the heating 

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plate and allowing them to cool, 3 ml of concentrated 

hydrochloric acid was added to each. The resultant 

solutions were transferred to polyethylene tubes, 

diluted with distilled water, mixed, and subsequently 

analyzed. Elemental measurements in aqueous 

solutions were conducted within a mass range of 5 to 

240. The analysis of each sample, ranging from 36 to 40 

samples, took approximately 2-3 minutes. Following 

each measurement, the spray system was thoroughly 

rinsed with a 2% nitric acid solution and water for 0.5-1 

minute.  

The analysis of the elemental composition of the 

“Hyposedaf” dry extract is presented in Table 1. 

Table 1. 

The elemental composition of the “Hyposedaf” dry extract 

Element Quantity, mkg/g Element Quantity, mkg/g 

Makroelementlar Gallic, Ga 0,170 

Potassium, K 65 000 Yttrium, Y 0,150 

Calcium, Ca 10 000 Neodymium, Nd 0,130 

Magnesium, Mg 9 100 Antimony, Sb 0,110 

Millielements Tin, Sn 0,076 

Tory, Th 0,071 

Phosphorus, P 4000 Zirconium, Zr 0,067 

Sodium, Na 1300 Platinum, Pt <0,05 

Aluminum, Al 460  Gold, Au <0,05 

Iron, Fe 280 Praseodym, Pr 0,04 

Manganese, Mn 53,0 Samari, Sm 0,028 

Zinc, Zn 47,0 Nanoelements 

Strontium, Sr 47,0 Gadolinium, Gd 0,018 

Rubidium, Rb 39,0 Tantalum, Ta <0,01 

Boron, B 32,0 Thallium, Tl <0,01 

Microelements European, Eu 0,01 

Titan, Ti 13,0 Dysprosium, Dy 0,01 

Copper, Cu 6,90 Ermiy, Er 0,01 

Nickel, Ni 4,10 Ittermiy, Yb 0,01 

Lithium, Li 2,10 Terbiy, Tb <0,01 

Molybdenum, Mo 1,40 Tuly, Tm <0,01 

Chrom, Cr 1,20 Gafni, Hf <0,01 

Silver, Ag 1,10 Berelli, Be 0,006 

Selenium, Se 0,50 Golmi, Ho 0,006 

Cerium, Ce 0,320 Indium, In 0,001 

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The subsequent section details the quantitative 

composition of elements in the dry extract, measured 

in micrograms per gram (as classified by Polyanskaya 

I.S., 2005): 

- Macroelements (К – 65 000, Ca – 10 000, Mg-9100);  

- Millielements (Р – 4000, Na – 4300, Al – 460, Fe – 280, 

Mn – 53, Zn – 47, Sr-47, Rb-39, В – 32);  

- Microelements (Ti-13, Cu 6.9, Ni-4.10,Li-2.10, Mo-1.40, 

Cr-1.20,Ag-1.10, Se-0.50, Ce-0.32, Ga-0.170, Y-0.150, Nd-

0.130, Sb-0.110, Sn-0.076, Th-0.071, Zr-0.067, Pt-<0.05, 

Au-<0.05, Pr-0.04, Sm-0.028 

- Nanoelements (Gd-0.18, Ta-<0.01, Tl-<0.01, Eu-0.01, Dy-

0.01, Er-0.01, Yb-0.01, Tb-<0.01, Tm-<0.01, Hf-<0.01, Be-

0.006, Ho-0.006, In-0.001) 

The results indicate that the “Hyposedaf” dry extract 

has a high biological value, containing substantial 

amounts of elements crucial for human life, such as 

potassium, calcium, phosphorus, magnesium, sodium, 

aluminum, iron, etc. Literature suggests that 

potassium compounds are vital for muscle contraction 

and brain function, and their deficiency can lead to 

fatigue and overall body weakness. 

Magnesium is an essential element found in all body 

tissues, crucial for the normal functioning of cells. It 

holds particular importance for internal organs, 

especially the heart and blood vessels. 

Calcium compounds present in the analyzed dry 

extract are key microelements for human life and 

health. In medical applications, they serve as sedatives, 

hemostatics, treatments for allergic diseases, and 

blood pressure regulators. Calcium is key in 

strengthening the skeletal system, forming tooth 

enamel, and plays significant roles in the immune 

system, skin, hair, nail health, and reproductive 

function. It is also instrumental in enhancing metabolic 

processes within the body. 

The high concentration of sodium compounds in the 

dry extract, combined with potassium, aids in 

maintaining acid-base balance, osmotic concentration 

of blood, activation of numerous enzymes, and 

contributes to the formation of membrane potential. 

Therefore, the “Hyposedaf” dry extract can be 

regarded as a valuable source of both macro and 

microelements. 

According to normative documents, the content of 

heavy metals and arsenic in plants and their derived 

medicinal preparations is in compliance with the 

standards of XIV State Pharmacopoeia and Sanitary-

epidemiological rules and norms. It has been verified 

that these concentrations do not exceed established 

limits. 

The obtained results are presented in Table 2. 

 

Table 2 

Amount of heavy metals and arsenic in dry extract of “Hyposedaf” mkg/g 

 

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Element Quantity according to 
the XIV State 

Pharmacopoeia  

Quantity as requested 
by the World Health 

Organization 

Determined amount 

Lead, Pb 6,0 10,0 0,750 

Cadmium, Cd 1,0 0,3 0,012 

Mercury, Hg 0,1 - not identified  

Arsenic, As 0,5 1,0 0,10 

 

In the subsequent stage of the research, we focused 

on analyzing the amino acid composition of the 

“Hyposedaf” dry extract. Initially, the process involved 

isolating free amino acids. For this, 1 ml of the test 

sample was combined with an equal volume (1 ml) of 

20% trichloroacetic acid. After a duration of 10 minutes, 

the resultant precipitate was separated through 

centrifugation at a speed of 8000 revolutions per 

minute for 15 minutes. Subsequently, 0.1 ml of the 

clarified liquid was isolated and subjected to 

lyophilization. The hydrolyzate was then evaporated, 

and the dry residue was dissolved in a triethylamine-

acetonitrile-water mixture (ratio 1:7:1) and dried again. 

The amino acid phenylthiocarbamyl derivatives (FTC) 

were synthesized using the phenylisothiocyanate 

reaction, following the methodology of Steven A. and 

Koen Daniel. The identification of these amino acid 

derivatives was carried out via high-performance liquid 

chromatography (HPLC). Employing an Agilent 

Technologies 1200 HPLC column detector (75x4.6 

mm), we utilized solution A: 0.14M CH3COONa with 

0.05% TEA at pH 6.4, and solution B: CH3CN. The flow 

rate was maintained at 1.2 ml/min, with detection at 

269 nm. The gradient percentage per minute was 

established as follows: 1-6%/0-2.5 min; 6-30%/2.51-40 

min; 30-60%/40.1-45 min; 60-60%/45.1-50 min; 60-

0%/50.1-55 min [13]. 

The chromatographic analysis of both the standard 

amino acid sample of the “Hyposedaf” dry extract and 

the amino acids present in the extract under study are 

depicted in Figures 1 and 2. 

Table 3 presents the quantitative composition and 

chemical structure classification of the amino acids. 

According to the data in Table 3, the “Hyposedaf” dry 

extract contains 20 amino acids, including 10 essential 

amino acids, which highlight its substantial quantity 

and pharmacological significance. The total amino acid 

content is 29.151 µg/g, of which 15.605 µg/g are 

essential amino acids, and the remaining 15.605 µg/g 

are non-essential. 

 

 

 

 

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1.Fig. Standard sample chromatogram of a mixture of amino acids 

 

2.Fig. Chromatogram of dry extract of “Hyposedaf”  

In the “Hyposedaf” dry extract, histidine stands out as 

a crucial essential amino acid, present at a 

concentration of 4.856877 µg/g. It is a key component 

of hemoglobin — the vital protein in red blood cells 

responsible for carrying oxygen from the lungs to body 

cells, and for transporting carbon dioxide in the 

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opposite direction. A deficiency in histidine can impede 

the body’s ability to produce the necessary quantity of 

hemoglobin, potentially leading to anemia. Histidine 

plays an important role in the functioning of the 

gastrointestinal tract, as well as in liver and kidney 

operations. Furthermore, the body converts histidine 

into histamine, a compound integral to the nervous 

system. Histamine is involved in synthesizing various 

hormones and is essential for regulating the heart's 

rhythm [14]. 

Table 3 

Analysis of amino acids of dry extract of “Hyposedaf” 

№ Amino acid Amount of amino 
acids, mkg/g 

Aliphatic amino acids 

Monoaminocarbons 

1. Alanine 0,077593 

2. Glysine 0,8991 

3. Valin 2,49772 

4. Isoleucine 0,342585 

5. Leucine 1,502734 

Oxysaminocarbons 

6. Serine 0,845982 

7. Threonine 0,310199 

Monoaminodicarbons 

8. Aspartic acid 0,321104 

9. Glutamic acid 0,31053 

Monoaminocarbon amides 

10. Asparagine 1,800129 

11. Glutamine 0,909695 

Diaminocarbons 

12. Lysine 0,194589 

13. Arginine 0,472063 

Sulfur preservative 

14. Cysteine 4,444809 

15. Methionine 2,508621 

Aromatic amino acids  

16. Phenylalanine 0,341222 

17. Tyrosine 3,820187 

18. Tryptophan  0,517421 

Heterocyclic amino acids  

19.  Proline 2,178288 

20. Histidine 4,856877 

Amount of non-essential amnio acids  13,544 

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Amount of exchangeable amino acids 15,605 

Total amount of amino acids 29,15145 

Methionine, at a concentration of 2.508621 µg/g, ranks 

second in abundance in the “Hyposedaf” dry extract. 

This essential sulfur-containing amino acid is known for 

its multiple unique functions [15]. Crucially involved in 

protein synthesis, methionine's fat-soluble properties 

also aid in preventing fatty liver disease. Furthermore, 

it serves as a foundational component for several 

hormones, including adrenaline, choline, and 

melatonin. Cysteine, present at 4.444809 µg/g, is 

another sulfur-bearing amino acid. Classified as semi-

essential or conditionally essential, cysteine is unique 

among the twenty amino acids for containing a thiol 

group (-HS) capable of undergoing both oxidation and 

reduction reactions. The oxidation process transforms 

cysteine into cystine, and it also contributes 

significantly to metabolic processes and the formation 

of peptides and proteins [16]. 

Tyrosine, found at 3.820187 µg/g, plays roles in 

reducing appetite, decreasing fat accumulation, 

stimulating melanin production, and enhancing the 

functionality of the adrenal, thyroid, and pituitary 

glands [17]. 

CONCLUSION 

In conclusion, the “Hyposedaf” dry extract contains 61 

micro and macro elements. This includes 7 essential 

elements (Co, Cr, Cu, Fe, Mn, Se, Zn) and 4 conditionally 

necessary ones. Key elements like potassium, calcium, 

sodium, sulfur, iron, magnesium, and phosphorus are 

crucial for human health and are prominently featured 

in the extract. The concentrations of heavy metals and 

arsenic in the extract were found to be within 

acceptable limits as defined by the Sanitary-

epidemiological rules and norms of State 

pharmacopoeia of the Republic of Uzbeksitan (Volume 

1, Issue 1) and XIV State Pharmacopoeia (RF). 

Moreover, the amino acid analysis of the dry extract 

revealed the presence of 20 amino acids, including 10 

essential ones. The total amino acid content is 29.151 

µg/g, with essential amino acids accounting for 15.605 

µg/g and non-essential ones making up the remaining 

15.605 µg/g. These findings underscore the 

“Hyposedaf” dry extract’s richness in amino acids, 

macro, and microelements, affirming its significant role 

in the development of other effective pharmaceutical 

formulations. 

REFERENCES 

1. 2.3.2.1078-01. Hygienic requirements for the 

safety and nutritional value of food products. 

Sanitary and epidemiological rules and 

standards: [implemented 01.09.2002]. 

Moscow: Ministry of Health of Russia, 2002, 

p.266. 

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2. State Pharmacopoeia of the Republic of 

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6. Moricawa T., Ninomiya K., Akaki J., Kakihara N., 

Kuramoto H., Matsumoto Y., Hayakawa T., 

Muraoka O., Wang L.-B., Wu L.-J., Nakamura S., 

Yoshikawa M., Matsuda H. Dipeptidyl 

peptidase-IV inhibitory activity of dimeric 

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Helichrysum arenarium // Journal of Natural 

Medicines. 2015; 69 (4): 494–506. DOI: 

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