Research Article

Bicinchonic Acid Assessment for Improvement of Sperms Mitochondrial Functions by Ginseng Extract and Nanoparticales in Rats

Noor A. Namaa*, Hayder A.N. Al-Zamely

Department of Physiology, Biochemistry and Pharmacology, College of Veterinary Medicine, University of Al-Qadisiyah, Al-Diwaniyah, Iraq.

Received | November 15, 2023; Accepted | January 02, 2024; Published | February 05, 2024

*Correspondence | Noor A. Namaa, Department of Physiology, Biochemistry and Pharmacology, College of Veterinary Medicine, University of Al-Qadisiyah, Al-Diwaniyah, Iraq; Email: vet21.post10@qu.edu.iq.

Citation | Namaa NA, Al-Zamely HAN (2024). Bicinchonic acid assessment for improvement of sperms mitochondrial functions by ginseng extract and nanoparticales in rats.Adv. Anim. Vet. Sci., 12(3):441-450.

DOI | https://dx.doi.org/10.17582/journal.aavs/2024/12.3.441.450

ISSN (Online) | 2307-8316

Copyright: 2024 by the authors. Licensee ResearchersLinks Ltd, England, UK.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

INTRODUCTION

The majority of sperms are composed of significant mitochondrial components. Anifandis and others, 2017. found a link between decreased sperm motility and aberrant sperm mitochondria, proving that mtDNA deletions and other changes can have an impact on sperm functionality and male infertility at the molecular level. Research has showed that, in comparison to control samples, sperm from non- zoo spermic samples has abnormally high levels of certain mtDNA and transcripts that encode mitochondrial proteins that are encoded by the nucleus (Nowicka-Bauer et al., 2018; Ferramosca et al., 2021). Additional significant discovery was the correlation between Measurements of the quantity, vitality and activity of sperm as well as the function of enzymes found in mitochondria in sperm, particularly the mitochondrial ETC complex like succinate dehydrogenase (SDHA) (Zhou et al., 2019).The citrate SDHA The enzymes included in the Krebs cycle that are nuclear-encoded also showed greatest interactions among each of the investigated enzymes in arrangement as said by many readings (Ehinger et al., 2015; Beyramzadeh et al., 2017). Additional research revealed that animals lacking the antioxidant cytochrome with a testis- specific subunit produced significantly less sperm than did normal mice (Sinkler et al., 2017; Sun et al., 2018; Park and Pang, 2021). The effects of these inhibitors on sperm motility are detrimental to male fertility because several studies have shown that several ETC inhibitors may decrease the ability of the sperm mitochondria to move (Davila et al., 2016; Aitken, 2017). Ginseng is among valuable Chinese herbal medicine uses herbs. There are up to nine different varieties of ginseng, with the most common being Asian ginseng (Panax ginseng), American ginseng (Panax quinquefolium) and Japanese ginseng (Panax japonicus) which are all named after specific geographical regions. Ginseng has been shown to have a wide range of physiological effects, including those on the nervous, immunological, and cardiovascular systems. It has also been used to enhance sex performance. In countries in East Asia, ginseng. It has been put to use for several thousand millenia in the form of conventional medicine to heal ailments. It’s got risen one of the herbs that has been utilized the most frequently during the past 30 years (Yu et al., 2017). The distinctive bioactive elements of ginseng are ginsenosides, which are triterpene saponins. But ginsenosides are not the only part of ginseng that has therapeutic properties. The compound’s active component, gonintin, was just recently identified (Shao et al., 2018; Lee et al., 2018; Cho et al., 2019). However, the majority of pharmacological and medical studies on ginseng have focused on ginsenosides. He et al. (2018) claim that Throughout worldwide. Ejaculated sperms incubated with ginseng extract exhibited significantly higher sperm counts than those treated with a vehicle, according to studies (Wiwanitkit, 2005). In contrast to freshly harvested, untreated, thawed sperm, The movement of sperm or integrity of the membrane are greatly improved after therapy by ginsenoside Rg1 (50 g/ ml) (Kim et al., 2013). These findings suggest that the addition of ginseng extract to the cryogen used to preserve sperm could increase fertility. Due to polymeric nanoparticles’ (PNPs) increased stability, effectiveness, and security of administering drugs locally and systemically and promote Increased availability is made possible by efficient passage through membrane barriers, and therapeutic location selection is made possible by a change of the surface characteristics (Mittal et al., 2019).

MATERIALS AND METHODS

Preparation of ginseng extract and NPs

500g of American ginseng herb was purchased from a powder from a pharmacy store in the city of Babylon, Hilla, and the herbal material was worked on at Al-Qadisiyah University, College of Veterinary Medicine. Where the work was done by taking 100g of the powder of the American ginseng herbal substance and placing it in the thumble of the Soxhlet apparatus in the presence of ethanol (70%) and in this way the American ginseng was extracted. This process is repeated from (10-12) times for two days. After completing the process, the material was filtered by filter paper, then the material was placed in a plate and evaporated with the evaporation device for 2-3 days until the extract was dried. After the fumigation process, the herbal material American ginseng, where the extract of the herbal material was prepared, where it was dissolved in BBS phosphate solution, and the material was placed in the Vortex Mixer for five minutes, The American ginseng extract which prepared in the previous step were dissolved in BBS phosphate solution, and the material was placed in the Vortex Mixer for five minutes, and then it was placed in the Intellingnt ultrasonic processor using a device of UP200ht at a temperature of 50 ºC for a period of 30 minutes, a solution was obtained Research to convert the solutions in to Nano sizes .After that stored in 4 ºC until use (Aldulemy et al., 2021).

Characterization of nanoparticales

UV–visible spectroscopy

The absorbance of the produced nanoparticles was measured using UV-visible spectrophotometer at wavelengths ranging from 200 to 800 nm with 1 nm wavelength interval (Agilent-Cary 60, United States).

Scanning electron microscopic analysis (SEM)

The form, surface morphology, Size of the nanoparticles were determined using scanning electron microscopy. To summarize, the produced nanoparticle dispersion was air dried before being loaded into sample containers. The sample was then coated with gold using a sputter, coater in a vacuum, and images were obtained using a SEM at 20 kV and various magnifications.

Infrared (FTIR) fourier transform spectroscopy

The chemical composition was determined using Fourier transform infrared (FTIR) spectroscopy. It is one of the methods used to characterize nanoparticles by gathering information on the maximum values in particular ranges of surface activity, determining the inhomogeneous nature of the absorption and reflection spectra, and interpreting the information using the wavelengths that are present in the spectrum. The actual through sampling is done by Fourier transform (a mathematical operation), examining the vibrational structures of the materials Also, to determine the structure of the solid is done using the FTIR technique (Chen et al., 2015). And FTIR is used Crystallinity and nanoparticle sizes (Kumar and Kumbhat, 2016).

X-ray diffraction ( XRD ) analysis (XRD)

X-ray diffraction is a widely used technique as a useful characterization tool for a study, The chemical and crystalline composition as well as the physical properties of the material are non - destructive. And you use Also to characterize the crystal structure of Alicia. XRD analysis was performed to detect the composition (Zhang et al., 2016) for the crystalline compound.

Study design and sample collection

Sixty Wister rats were randomly selected and subjected for preparatory period; during which, they kept at 23 ± 2ºC, fed on basal laboratory diet and tap water, and exposed to 12/12 light and dark conditions. After one week, the study animals were equally divided into three groups as following:

1. First group (Negative control group): Animals of this group remain without any treatment and received only daily distilled water and give (2ml) from phosphate buffer solution (PBS)
2. Second groups (T1): Animals of this group were treated by Ginseng extract received (500 mg/kg) (2ml) orally by stomach tube for 60 day.
3. Third groups (T2): Animals of this group were treated by Ginseng NPs received (250mg/kg) (2m) orally by stomach tube for 60 days (Kamel et al., 2019).

At the ending of an experimental period, all study animals were a euthanized with chloroform and subjected for collection of testis tissue samples.

Bicinchonic acid assay

The total protein contents in homogenized testicular tissues were measured using the Bicinchoninic Acid Assay Kit (Elabscience, China) according to the manufacturer’s instructions. A standard curve for bovine serum albumin (BSA) was created using BSA dilutions in RIPA lysis Buffer according to manufacturer’s instructions. All samples were diluted using RIPA Lysis Buffer. Duplicate BSA or sample lysates were pipetted into plate. BCA reagents were used and from the mixture were added to each well of a well plate. The well plate was covered with aluminum foil, rocked using a plate shaker, and then incubated. After the incubation time, the absorbance value Optical Density (OD) of each well was measured using a Microplate reader at 562 nm. The linear standard curve of mean absorbance (two values) of BSA was used to measure the concentration of protein in the samples.

Western blotting assay (SDS-PAGE)

Protein Sample Preparation (Extraction of Proteins from Tissue Samples). carefully wash the tissue with pre-cooled PBS Buffer ((0.01 M) to (pH 7.4)) and remove any debris. The tissues should be weighed and broken up before being lysed with (RIPA) Lysis Buffer (E BC R327) at the proper ratio (add (10 L) PMSF and (10 L) Na 3a VO 4 to each (1mL) to RIPA Lysis of Buffer).-According to the tissue weight ratio, homogenization is advised: RIPA Lysis Buffer volume = 3:10. To 0.3 g of tissue the specimen, add 1 milliliter of RIPA Lysis Buffer; the precise volume can be adjusted to suit the needs of the experiment. Shakeandl yseonice for (30) minutes homogeneity. To completely lyse the cells and reduce the sample’s density, sonicate sample for 1 minute (in an iceewater bath) using 2 s of sonicat and 2 s of spacing time. A centrifuge at 4 °C for 10 minutes at 12,000 revolutions per minute (rpm). Using the BCA technique, measure the protein level in the dregs. Electrophoresis based on molecular weight of the target protein prepare an appropriate separation gel. In a reserve well, add 5 L of Precision Plus Protein TM Dual Color Standards Marker (161-0374, Bio-Rad, UK) to confirm target molecular weights and the degree membrane of transfer. With Electrophoresis Buffer (EBCR331) begin with the electrophoresis process.45 minutes of electrophoresis at 100 volts while the samples are in the stack gel followed by 50 minutes at 150 volts when the blue wave enters the gel that separates the samples. Bromophenol blue electro phonetically migrates across the gel in around 1.35 hours. Transfer Membrane (Wet Transfer) Depending on the molecular r weight of the target proteins use the (PVDF) Membranes (EBCR266) with a 0.45 m pore size. The PVDF Membrane must first be activated by soaking in methanol for one minute before being placed in transmembrane Buffer (EBCR333). Additionally, the Transmembrane Buffer must be pre-soaked with fiber mat and filter paper before usage. Put the parts in the following sequence: (negative electrode) black plate, fiber mat, filter paper, and gel PVDF The wet transference tank contains the membrane filtration paper fiber mate white plate(positive electrode) in the following order discharge bubbles, clamp, and place in wet transfer tanks. Depending on the molecular mass of the chosen protein, modify the transmembrane conditions. Make sure that low temperatures are used during the transmembrane procedure. Following the transmembrane, gently remove the (PVDF) Membrane and washed with (TBST) Buffer for a minut. Incubation of Antibodies (Ab):-Soak the PVDF The membrane for 1.5 hours at room temperature in TBST Buffer (EBCR335) containing 5% bovine serum albumin (BSA) as a blocking buffer. Incubate Overnight at 4 °C, soak the (PVDF)* Membrane in the primary antibody working solution. and then give it a gentle shake. The primary antibody should be diluted as directed in the antibody manual (Anti-SDHA Antibody and Beta Actin; 0.5L: 1mL BSA). Using TBST Buffer, wash the PVDF Membrane three times for 15 seconds each. Use a TBST Buffer containing 5% BSA to dilution the secondary anti body’s (EAB 1003, EAB c1001) at a ratio of 1:5000 in accordance with the recommended secondary antibody concentration. Incubate at room temperature for an hour off a shakers (PVDF) Membrane Cleaning on three occasions (15 minutes) each with TBST Buffer. Detection: (EBC R347) excellent chemiluminescent substrate detection kit contains the following components: combine A and B in a (1:1) ratio as a working liquid. Take the (PVDF) Membranes out of the (TBST) Buffer and immerse it with a filter papers in the solution. Place the (PVDF) Membrance within the X-ray cassette, continuously saturate it with the ECL work solution, expel the air bubble, and detect the result using X-ray film. 3. To get the best photograph, adjust the exposure time.

Statistical analysis

All data come from at least three independent experiments. Values were represented as per (Mean ± Standard Error (M±SE)). Satiating significance be there calculated using SAS using the one way ANOVA. Differences were considered significant atP< 0.05 (*) and P<0.01 (**) considered significant (Stroupet al., 2018).

RESULTS and Discussion

Characterization of ginseng NPs

The Araliaceae family includes the perennial plant ginseng. In the current study, we efficiently and affordably biosynthesized gold nanoparticles utilizing ginseng aqueous extract. The analysis includes UV_V is spec HR, TEM, XRD, FTIR and SAED. analyses were used to evaluate the produced ginseng gold nanoparticles (GNPs). The surface plasma on resonance peak at (538 nm), is observed was detected by UV-Vis spectroscopic investigation, and it doesn’t fade until 30 days had passed since incubation. The produced gold nano particles have a shape is spherical crystalline nature are varying sizes are confirmed by the result of (HRTEM XRD and SAED). According to FTIR data, the biological elements in ginseng reduced gold ion to create gold nanoparticle.

UV-visible spectroscopy analysis

A double beam UV-visible spectrophotometer (Shimadzu, model 1800) having two matched quartz cells with 1 cm light path length and loaded with UV probe software was used for the recording of spectra and measuring absorbance for method development and validation study.

The wave length of nanoparticles were measured from 200 to 1100 nm for wave length by recording UV-visible spectrum of prepared nanoparticales solution. Maximum absorbance (λ max 1.372) was shown at 245 nm Figure 1.

 

SEM analysis

The morphology of nanoparticles was also confirmed in terms of crystallite size utilizing scanning electron microscopy (SEM). The SEM images shown in Figure 2. Exhibited those particles have spherical shape with a smooth exterior. The average diameter of the obtained NPs evaluated by SEM was 98.24 nm at range from 49 to 132 nm.

 

The analysis of ginseng nanoparticles was conducted using FTIR (fourier transform infrared spectroscopy)

Fourier Transform Infrared Spectroscopy (FTIR) spectroscopy of ginseng exhibited the presence of bending vibration of the aliphatic CH bond at the peaks 781.70 and 2919.18cm-1. While the wave numbers 977.80, 963.09, 1577.34, 1540.23 and 1510.24cm-1, which are may be attributed to the stretching and blending of the double carbon bond of the C=C. As for the peaks at numbers 1627.76, 1602.58, 1281.88 and 1233.44 cm-1, they may be due to the presence of a vibrational bending of C=O and C-O, due to the aromatic amine group, respectively. As OH was found at the wave numbers 3778.20, 3687.97, 3658.73, 2956.30, 2919.18, and 2850.02cm-1, was attributed to the carboxylic groups. According to the above, the FTIR spectrum of pure extract of ginseng showed a number of peaks, including the wave numbers 1627.76 and 1602.58 cm-1 that are attributed to the CH, in addition, there are other groups in the solution that confirm the presence of ginseng molecules associated with, including cm-1, respectively Figure 3.

 

The text describes an x-ray diffraction analysis

The Paul Scherrer equation is utilized for determining the size of powder crystals, expressed as Dp = (0.94 X λ)/ (β X Cos θ). The average crystallite size (Dp), line broadening (β), Bragg angle (θ), and X-Ray wavelength (λ) are all important factors to consider. The present data of XRD Ginseng recorded four main peak 2 Theta at 26.231, 42.048, 54.374 and 77.229 with their FWHM 0.363, 0.777, 0.767 and 0.572 to recorded size at 22, 11 and 15 nm, respectively Figure 4.

Bicinchonic acid assay

The total protein concentration data revealed a substantial increase (p<0.05) in values of both Ginseng extracts. (2.44±0.07) and Ginseng NP (3.02±0.05) groups when compared with the value of control (1.89±0.05) group. However, no significant variationwas found between values of both Ginseng extract and Ginseng NP groups Tables 1, 2 and Figure 5.

 

 

Western blotting assay

The results of the current study showed the level of SDHA protein in Ginseng extract group and Nano group compared to non treated control group. SDHA protein increased significantly in both treated group in compare with control group. Moreover, theGinseng nanoparticls goup showed highly significant increased (3.029 ± 0.058): P ˂ 0.0001) in level of SDHA compared with Ginseng extract group(2.443 ± 0.079): P ˂ 0.0001), respectively (Figure 6A, B, C). The image showing the level

 

Table 1: Total protein content in homogenized tissue in Ginseng Nanoparticles and other group.

Standard concentrations of BSA1 (mg/mL)			2 Reading data (replication) absorbance unite (nm)		Samples codes		2 Reading data (replication) Absorbance unite (nm		Final concentration of total protein (µg/µL)
2			1.135	1.191	C1		3.129	3.331	16.9
1.5			0.904	0.96	C2		3.303	3.346	15.3
1			0.672	0.733	C3		1.838	1.948	25.1
0.75			0.501	0.546	E1		3.225	3.111	20.8
0.5			0.431	0.527	E2		2.921	3.095	21.4
0.25			0.298	0.361	E3		3.151	3.234	17.4
o.125			0.192	0.246	N1		3.156	3.247	13.8
0.025			0.157	0.183	N2		3.107	3.026	19.6
5		0.121		0.15	N3	3.001		3.281	18.1

 

1Bovine serum albumin standard concentrations. 2Absorbance reading using Micro plate reader at 562 nm wavelength.

 

 

Table 2: SDHA/B-Actin concentration in ginseng Nanoparticles and other group.

Group	Conc- (Mean ±SE)
Control	1.89±0.05A
Ginseng Extract	2.44±0.07B
Ginseng Nanoparticles	3.02±0.05C
P Value	<0.0001
LSD(P<0.05)	0.192

 

Different letters between any two means denote to the significant difference at P<0.05

 

of SDHA protein in extract group and Nano group compared to non treated control group. SDHA protein increased significantly in both treated group in compare with control group. (B) The image showing the level of B-Actin as a housekeeping gene that presents the equelizing of pipetting error and non effecting of cell homeostasis during treated process for all animals of experiment. (C) The bar chart showing the columns of means, which are based on band intensity analysis data for each group.

UV-visible spectroscopy analysis

For the method development and validation study, a double beam UV-visible spectrophotometer (Shimadzu, model 1800) with two matched quartz cells with 1 cm light path length and loaded with UV probe software was used to record spectra and measure absorbance. recording spectra and measuring absorbance. Wave length of nanoparticles was measured (Gibis et al., 2014).

SEM analysis

The morphology of nanoparticles was also confirmed in terms of crystallite size utilizing scanning electron microscopy (SEM). exhibited those particles have spherical shape with a smooth exterior (Olatunad et al., 2020).

The analysis of ginseng nanoparticles was conducted using FTIR (Fourier Transform Infrared Spectroscopy)

The FTIR spectra for ginseng showed appear new the study uses FTIR results to predict the chemical and biological activities of biomolecules, providing insights for effective plant product use. Exhibited the characteristic bands at 1384.04 and 1224.17 cm-1 indicating the presence of N-H stretching aliphatic primary amine group, at 2959.01 cm-1 presence of strong broad N-H stretching amine salt. Moreover, the carboxylic acid O-H was confirmed at 1384.04 cm-1. While strong - C-O for primary alcohol and sulfoxide was recorded at 1056.99 and 1033.85 respectively. The presence of esters (S-OR) group at 703.77 cm-1 (Pimentel-Moral et al., 2019).

The text describes an x-ray diffraction analysis

In common, the material in crystalline state appears The XRD analysis revealed the crystalline nature of ginseng extracts within the liposome, indicating better solubility in aqueous medium. The ginseng extracts were found to be interposed between phospholipid bilayer chains, enhancing flow ability (Rasmussen et al., 2020).

Bicinchonic acid assay

It should be highlighted that the detection of- producing proteins of normal spermiogenesis, suggesting a potential target pathway for further investigation. In seminal plasma, the cytoskeleton and intracellular proteins have been seen in earlier investigations (Pilch and Mann, 2006; Batruch et al., 2012). These proteins may come from spermatogenesis, epididymal fluid, or prostasomes, secretory vesicles seen in seminal plasma (Gerena et al., 1998; D’Amours et al., 2012; Pilch and Mann, 2006). Which have been demonstrated to transport particular proteins to spermatozoa in the male reproductive tract and so modify sperm activity, are particularly abundant in the epididymal fluid. Ginseng appears to enhance sperm quality in guinea pigs exposed to dioxin, according to recent research (Hwang et al., 2004). The results of numerous clinical experiments to increase sperm quality and quantity have been mixed (Chaney, 1979). In the current study, ginseng-treated rats had considerably higher epididymis sperm counts than the rats in the control group. The use of ginseng also increased sperm motility, although it had little effect on testicular or body weight. This shows that ginseng might be suggested as a therapy for male infertility linked to spermatogenesis. Expanding the therapy options for male factors in infertility might benefit from a mechanistic knowledge of how ginseng promotes spermatogenesis (Park et al., 2007). According to reports, oxidative stress contributes to infertility in people with UMI (Doshi et a., 2015; Mayorga-Torres, 2014) and has an impact on spermatozoa function. According to reports, sperm protein expression levels are changed when under ongoing oxidative damage examined by (Agarwal et al., 2014) Canonical pathways like Cellular compromise, cell death, and survival are all interconnected processes. We have previously shown that infertile males has a significant reactive oxygen species content have variable expression of proteins related with oxidative stress, protein changes, and energy consumption and control (Sharma and Kiran, 2013). Additionally, in spermatozoa under oxidative stress, the proteins linked to the antioxidant system, such as peroxiredoxin, are under expressed (Sharma and Kiran, 2013; Hamada et al., 2012). The process of fertilization depends heavily on sperm proteins that are specifically relevant to reproductive function. The study on infertile males with normozoospermia revealed that several proteins required for fertilization were under expressed (reviewed by Bracke et al., 2018). In patients with in vitro fertilization failure, Li et al. (2018). The study has established a connection between the fertilization process and (ACRBP (acrosin- binding protein)) and (ZPBP1 (zona pellucida- binding protein)) (Liu et al., 2018).

Western blotting assay

By controlling redox proteins in elderly rats Korean red ginseng, also known as Panax ginseng Meyer, is a type of ginseng native to Korea has been shown to improve testis efficiency Sperm maturation is a crucial process in the reproductive cycle. The purpose of this study was to see Korean red ginseng water extract (KRG-WE) regulates changes in enzymatic and non-enzymatic antioxidants in aged rats’ testes, affecting spermatogenesis, sex hormone receptors, and enzymes controlling oxidation, histone deacetylation, and growth-related activities. Additionally, the enzyme-modified P. ginseng (Pectinase-treated) reduced the oxidative stress caused by hydrogen peroxide in GC-2 sperm cells, as well as altered expression of testicular genes in elderly rats (Kopalli et al., 2015). Ginseng nanoparticles demonstrated a noticeable improvement in mitochondrial enzymes through increased intestinal absorption of extract as well as highly cellular uptake by tissue, prolonged circulation with sustained released the statement is currently inactive. compound of extract, and synergistic interaction between extract to potentiate the body immune system with improved normal function. On the other hand, immunohistochemical analysis of SDHA in Ginseng semen corroborated the notion of a considerable rise in SDHA gene expression and protein expression of mitochondria in rats given red ginseng. The NPs group found 95% of semen in the epididymis lumen, and SDHA was significantly expressed in every epididymal epithelial cell in the study by Wang et al. (2020). Compared to the group of Ginseng extract semen, where SDHA appears to be involved in approximately. It has been discovered that Spermato motility and, consequently, fertilization potential are connected with Spermato mitochondrial activity. Additionally, a number of studies have shown in which the mitochondrial tablet directs a wide range of waving proteins to the average-piece and may thus be essential for spermatozoa function (Lu et al., 2017; Khalid et al., 2020; Gallo et al., 2021). The study found that Ginseng extract and Ginseng NPs significantly increased SDHA protein levels, as confirmed by the western blotting assay results. Italiano et al. (2012) noted a decrease in SDHA protein expression in tumor cases compared to non-tumor instances, stating that this was to be expected given that tumor-related mutations can result in shortened SDHA proteins. The SDHA gene encodes the primary catalytic subunit, and germline mutations in SDHA are linked to neurological conditions like early- onset encephalopathy, ocular atrophy, ataxia, and myopathy (Ghaoui and Sue, 2018; Di Donato, 2009). Ginsenosid is the primary active component of ginseng. Ginsenoside Rb1 controls ROS generation and levels in mitochondria to control mitochondrial activity (Li et al., 2012; Zhou et al., 2019). According to research, Lycium barbarum polysaccharides greatly enhance mitochondrial activity, plasma membrane integrity, and sperm survival (Zheng et al., 2022). By blocking the antioxidant action of the caspase pathways, angelica polysaccharides decrease apoptosis and ROS, safeguard mitochondrial integrity, and enhance energy metabolism (Zhuang et al., 2020). The primary active component isolated from Epimedium, Icaridin is frequently used to treat male erection problems. By promoting cell proliferation and obstructing mitochondria-dependent apoptotic pathways, as well as by exhibiting protective actions against testicular dysfunction, it cures sperm abnormalities in rats (He et al., 2021). According to Yin et al. (2010) psyllium polysaccharide may be helpful as a natural antioxidant. According to Shin et al. (2011) Cuscuta sinensis is a TCM herb with liver and kidney tonic qualities, can enhance kidney function. According to Loh et al. (2010) the active ingredient in Leonurus japonicus leonurin can suppress the synthesis of mitochondrial ROS and restore mitochondrial function and redox state. Curculigo orchioides improves sexual dysfunction brought on by excessive sugar levels and supports Yang; it also increases sperm parameters and count in rats (Thakur et al., 2012). TCMs frequently have several components, multiple potencies, multiple targets and multiple action pathway.

CONCLUSIONS and Recommendations

By utilizing the Western Blotting and Bicinchonic Acid tests, this work offered fresh insights into the mitochondrial expression of SDHA proteins. Both ginseng extract and ginseng NP treatment indicated a considerable protein expression, with ginseng NP use in particular showing no adverse effects during the course of the trial. However, isolating the active Ginseng components and creating specialized NPs based on one of these components can result in novel treatments with distinctive properties. It is also important to continue researching the enhancement of ginseng NPs and proper control of genes associated to steroidogenesis and mitochondrial biogenesis.

ACKNOWLEDGMENTS

The author is grateful to The Head and all staffs of the Department of Physiology, Biochemistry and Pharmacology (College of Veterinary Medicine, Al-Qadisiyah University) for all facilities and helping in this work.

NOVELTY STATEMENT

The novelty of the study is pocus on physiologically activity of Ginseng Nanoparticales in improvement of rats sperms mitochondrial function which means possibility for improvement of sperms functions and then increase animal reproduction.

AUTHOR’S CONTRIBUTION

These authors each contributed equally.

Ethical approval

The current study was conducted with the permission of the Departments of Physiology, Biochemistry, and Pharmacology. College of Veterinary Medicine, Al-Qadisiyah University (Al-Qadisiyah, Iraq).

Conflict of interest

The authors have declared no conflict of interest.

REFERENCES

Agarwal S, Lucca D, Serum A, Trebbia F (2014). Inconsistent regulators: Evidence from banking. Quart. J. Econ., 129(2): 889-938. https://doi.org/10.1093/qje/qju003

Aitken RJ (2017). Reactive oxygen species as mediators of sperm capacitation and pathological damage. Mol. Reprod. Dev., 84(10): 1039-1052. https://doi.org/10.1002/mrd.22871

Aldulemy QLM, Abdul-Razak MMA (2021). Effect of nano seaweed extract on tillaring pattrin growth and yield of barley varieties. Int. J. Aquat. Sci., 12(2): 5385-5400.

Anifandis G, Amiridis G, Dafopoulos K, Daponte A, Dovolou E, Gavriil E, Psarra AMG (2017). The in vitro impact of the herbicide roundup on human sperm motility and sperm mitochondria. Toxics, 6(1): 2. https://doi.org/10.3390/toxics6010002

Batruch I, Smith CR, Mullen BJ, Gruber E, Lo KC, Diamonds EP, Jarvis KA (2012). Analysis of seminal plasma from patients with non-obstructive azoospermia and identification of candidate biomarkers of male infertility. J. Proteome Res., 11(3): 1503-1511. https://doi.org/10.1021/pr200812p

Beyramzadeh M, Dikmen ZG, Erturk NK, Tuncer ZS, Akbiyik F (2017). Placental respiratory chain complex activities in high-risk pregnancies. J. Maternal-Fetal Neonatal Med., 30(24): 2911-2917. https://doi.org/10.1080/14767058.2016.1268594

Bracke A, Pesters K, Punjabi U, Hoogewijs D, Dewilde S (2018). A search for molecular mechanisms underlying male idiopathic infertility. Reprod. Biomed. Online, 36(3): 327-339. https://doi.org/10.1016/j.rbmo.2017.12.005

Chaney EM (1979). Supermadre: Women in politics in Latin America. University of Texas Press.

Chen T, He T, Benesty M, Khotilovich V, Tang Y, Cho H, Zhou T (2015). Xgboost extreme gradient boosting. R Package Version 0.4-2., 1(4): 1-4.

Cho YJ, Kim IH, Cho YJ (2019). Numerical analysis of the grand circulation process of Mang-Bang beach-centered on the shoreline change from 2017. 4. 26 to 2018. 4. 20. J. Korean Soc. Coastal Ocean Eng., 31(3): 101-114. https://doi.org/10.9765/KSCOE.2019.31.3.101

D’amours G, Kibar Z, Mahoney G, Feint R, Tiny F, Deselects V, Limier E (2012). Whole-genome array CGH identifies pathogenic copy number variations in fetuses with major malformations and a normal karyotype. Clin. Genet., 81(2): 128-141. https://doi.org/10.1111/j.1399-0004.2011.01687.x

Davila CC, Reinhart CF, Bemis JL (2016). Modeling Boston A workflow for the efficient generation and maintenance of urban building energy models from existing geospatial datasets. Energy, 117: 237-250. https://doi.org/10.1016/j.energy.2016.10.057

Di Donato F (2009). La sienna e la rete louse public Della region nell’età del web (Vol. 93). Firenze university press. https://doi.org/10.26530/OAPEN_341473

Doshi P, Adsule P, Banerjee K, Oulkar D (2015). Phenolic compounds, antioxidant activity and insulinotropic effect of extracts prepared from grape (Vitis vinifera L) byproducts. J. Food Sci. Technol., 52: 181-190. https://doi.org/10.1007/s13197-013-0991-1

Dwivedi C, Sahu R, Tiwari SP, Satapathy T, Roy A (2014). Role of liposome in novel drug delivery system. J. Drug Delivery Ther., 4: 116-129. https://doi.org/10.22270/jddt.v4i2.768

Ehinger BV, König P, Ossandón JP (2015). Predictions of visual content across eye movements and their modulation by inferred information. J. Neurosci., 35(19): 7403-7413. https://doi.org/10.1523/JNEUROSCI.5114-14.2015

Ferramosca A, Lorenzetti S, Di Giacomo M, Murrieri F, Coppola L, Zara V (2021). Herbicides glyphosate and glufosinate ammonium negatively affect human sperm mitochondria respiration efficiency. Reprod. Toxicol., 99: 48-55. https://doi.org/10.1016/j.reprotox.2020.11.011

Gallo MB, Aghagoli G, Lavine K, Yang L, Siff E J, Chiang SS, Michelow IC (2021). Predictors of COVID-19 severity: a literature review. Reviews in medical virology, 31(1): 1-10.https://doi.org/10.1002/rmv.2146

Gerena RL, Irikura D, Urade Y, Eguchi N, Chapman DA, Killian GJ (1998). Identification of a fertility-associated protein in bull seminal plasma as lipocalin-type prostaglandin D synthase. Biol. Reprod., 58(3): 826-833. https://doi.org/10.1095/biolreprod58.3.826

Ghaoui R, Sue CM (2018). Movement disorders in mitochondrial disease. J. Neurol., 265: 1230-1240. https://doi.org/10.1007/s00415-017-8722-6

Gibis M, Zeeb B, Weiss J (2014). Formation characterization and stability of encapsulated hibiscus extract in multilayered liposomes. Food Hydrocol., 38: 28-39. https://doi.org/10.1016/j.foodhyd.2013.11.014

Hamada A, Esteves SC, Nizza M, Agarwal A (2012). Unexplained male infertility. diagnosis and management. Int. Braz. J. Urol., 38: 576-594. https://doi.org/10.1590/S1677-55382012000500002

He W, Meng T, He X, Ge S (2018). Unified iterative learning control for flexible structures with input constraints. Automatica, 96: 326-336. https://doi.org/10.1016/j.automatica.2018.06.051

He W, Zhang ZJ, Li W (2021). Information technology solutions, challenges, and suggestions for tackling the COVID-19 pandemic. Int. J. Inf. Manage., 57: 102287. https://doi.org/10.1016/j.ijinfomgt.2020.102287

Hwang MH, Jang NJ, Hyun SH, Kim IS (2004). Anaerobic bio-hydrogen production from ethanol fermentation the role of pH. J. Biotechnol., 111(3): 297-309. https://doi.org/10.1016/j.jbiotec.2004.04.024

Italiano A, Sung YS, Zhang L, Singer S, Maki RG, Coindre JM, Antonescu CR (2012). High prevalence of CIC fusion with double-homeobox (DUX4) transcription factors in EWSR1-negative undifferentiated small blue round cell sarcomas. Genes Chromos. Cancer, 51(3): 207-218. https://doi.org/10.1002/gcc.20945

Kamel H, Longstreth Jr WT, Tirschwell DL, Kronmal RA, Broderick JP, Palesch YY, Elkind MS (2019). The AtRial cardiopathy and antithrombotic drugs in prevention After cryptogenic stroke randomized trial. Rationale and methods. Int. J. Stroke, 14(2): 207-214. https://doi.org/10.1177/1747493018799981

Kim SM, Hsu A, Lee YH, Dresselhaus M, Palacios T, Kim KK and Kong J (2013). The effect of copper pre-cleaning on graphene synthesis. Nanotechnology, 24(36): 365602. https://doi.org/10.1088/0957-4484/24/36/365602

Khalid Z, Yousaf, MA, Khan ATA, Shakoori FR, Munir M, Shakoori AR (2020). Debunking Myths about COVID-19, paranoiac misconceptions, recent developments and its current stance. Pakistan J. Zool., 52: 2377-2387. https://dx.doi.org/10.17582/journal.pjz/20200608144416

Kopalli SR, Hwang SY, Won YJ, Kim SW, Cha KM, Han CK, Kim SK (2015). Korean red ginseng extract rejuvenates testicular ineffectiveness and sperm maturation process in aged rats by regulating redox proteins and oxidative defense mechanisms. Exp. Gerontol., 69: 94-102. https://doi.org/10.1016/j.exger.2015.05.004

Kumar N, Kumbhat S (2016). Essentials in nanoscience and nanotechnology. John Wiley and Sons. https://doi.org/10.1002/9781119096122

Lee JX, Grey M, Ha S, Kunz T, Jain S, Ye Y, Karen Liu C (2018). Dart dynamic animation and robotics toolkit. J. Open Source Software, 3(22): 500. https://doi.org/10.21105/joss.00500

Li J, Shen B, Hong Z, Lin B, Gao B, Chen Y (2012). A facile approach to synthesize novel oxygen-doped g-C 3 N 4 with superior visible-light photoreactivity. Chem. Commun., 48(98): 12017-12019. https://doi.org/10.1039/c2cc35862j

Li J, Tan J, Martino M M, Lui K O (2018). Regulatory T-cells: potential regulator of tissue repair and regeneration. Frontiers in immunology, 9: 585. https://doi.org/10.3389/fimmu.2018.00585

Liu C, Zoph B, Neumann M, Shlens J, Hua W, Li LJ, Murphy K (2018). Progressive neural architecture search. Proc. Eur. Conf. Comp. Vision, pp. 19-34. https://link.springer.com/conference/eccv, https://doi.org/10.1007/978-3-030-01246-5_2

Loh KP, Bao Q, Ang PK, Yang J (2010). The chemistry of graphene. J. Mater. Chem., 20(12): 2277-2289. https://doi.org/10.1039/b920539j

Lu L, Guo X, Zhao J (2017). A unified nonlocal strain gradient model for nanobeams and the importance of higher order terms. Int. J. Eng. Sci., 119: 265-277. https://doi.org/10.1016/j.ijengsci.2017.06.024

Mayorga Torres T (2014). Using DInSAR as a tool to detect unstable terrain areas in an Andes region in Ecuador (South America). In EGU General Assembly Conference Abstracts., p. 16203.

Michelob IC (2021). Predictors of COVID-19 severity a literature review. Rev. Med. Virol., 31(1): 1-10.

Mittal S, Khan MA, Romero D, Wuest T (2019). Smart manufacturing: Characteristics, technologies and enabling factors. Proceedings of the institution of mechanical engineers, Part B. J. Eng. Manuf., 233(5): 1342-1361. https://doi.org/10.1177/0954405417736547

Nowicka-Bauer K, Lepczynski A, Ozgo M, Kamieniczna M, Fraczek M, Stanski L, Kurpisz MK (2018). Sperm mitochondrial dysfunction and oxidative stress as possible reasons for isolated asthenozoospermia. J. Physiol. Pharmacol., 69(3):

Olatundeo O, Benjakul S, Vongkamjan K, Amnuaikit T( 2020). Influence of stabilising agents on the properties of liposomal encapsulated ethanolic coconut husk extract. Int. J. Food Sci. Technol., 55: 702-711. https://doi.org/10.1111/ijfs.14339

Park DH, Lee J, Han I (2007). The effect of on-line consumer reviews on consumer purchasing intention: The moderating role of involvement. Int. J. Electron. Commerce, 11(4): 125-148. https://doi.org/10.2753/JEC1086-4415110405

Park YJ, Pang, MG (2021). Mitochondrial functionality in male fertility from spermatogenesis to fertilization. Antioxidants, 10(1): 98. https://doi.org/10.3390/antiox10010098

Pilch B, Mann M (2006). Large-scale and high-confidence proteomic analysis of human seminal plasma. Genome Biol., 7(5): 1-10. https://doi.org/10.1186/gb-2006-7-5-r40

Pimentel-Moral S, Teixeira M, Fernandes A, Borrás-Linaresi, Arráez-Román D, Martínez-Ferez A, Segura-Carretero A, Soutoe B (2019). Polyphenols-enriched Hibiscus sabdariffa extract-loaded nanostructured lipid carriers (NLC): Optimization by multi-response surface methodology. J. Drug Delivery Sci. Technol., 49: 660-667. https://doi.org/10.1016/j.jddst.2018.12.023

Rasmussen MK, Pedersen JN, Marie R (2020). Size and surface charge characterization of nanoparticles with a salt gradient. Nat. Commun., 11: 1-8. https://doi.org/10.1038/s41467-020-15889-3

Shao C, Ciampaglia GL, Varol O, Yang KC, Flammini A, Menczer F (2018). The spread of low-credibility content by social bots. Nat. Commun., 9(1): 1-9. https://doi.org/10.1038/s41467-018-06930-7

Sharma A, Kiran R (2013). Corporate social responsibility: Driving forces and challenges. Int. J. Bus. Res. Dev., 2(1). https://doi.org/10.24102/ijbrd.v2i1.182

Shin SC, Kim SH, You H, Kim B, Kim AC, Lee KA, Lee WJ (2011). Drosophila microbiome modulates host developmental and metabolic homeostasis via insulin signaling. Science, 334(6056): 670-674. https://doi.org/10.1126/science.1212782

Sinkler CA, Kalpage H, Shay J, Lee I, Malek MH, Grossman LI, Hüttemann M (2017). Tissue-and condition-specific isoforms of mammalian cytochrome c oxidase subunits: from function to human disease. Oxid. Med. Cell. Long., https://doi.org/10.1155/2017/1534056

Stroup W, Milliken G A, Claassen, E A, Wolfinger RD(2018). SAS for mixed models: Introduction and basic applications. SAS Institute.

Sun Y, Zheng L, Yang Y, Tian Q, Wang S (2018). Beyond part models. Person retrieval with refined part pooling (and a strong convolutional baseline). In: Proceedings of the European conference on computer vision (ECCV). Pp. 480-496. https://link.springer.com/conference/eccv, https://doi.org/10.1007/978-3-030-01225-0_30

Thakur JK, Han J, Pokharel SK, Parsons RL (2012). Performance of geocell-reinforced recycled asphalt pavement (RAP) bases over weak subgrade under cyclic plate loading. Geotext. Geomembranes, 35: 14-24. https://doi.org/10.1016/j.geotexmem.2012.06.004

Wang C, Pan R, Wan X, Tan Y, Xu L, McIntyre RS, Ho C (2020). A longitudinal study on the mental health of general population during the COVID-19 epidemic in China. Brain, Behav. Immune., 87: 40-48. https://doi.org/10.1016/j.bbi.2020.04.028

Wiwanitkit V (2005). A review of human sparganosis in Thailand. Int. J. Infect. Dis., 9(6): 312-316. https://doi.org/10.1016/j.ijid.2004.08.003

Wojcik GL, Graff M, Nishimura KK, Tao R, Haessler J, Gignoux CR, Carlson CS (2019). Genetic analyses of diverse populations improve discovery for complex traits. Nature, 570(7762): 514-518. https://doi.org/10.1038/s41586-019-1310-4

Yin YH, Sun N, North TH, Hu SS (2010). Hook formation and mechanical properties in AZ31 friction stir spot welds. J. Mat. Proc. Technol., 210(14): 2062-2070. https://doi.org/10.1016/j.jmatprotec.2010.07.029

Yu Y, Jia T, Chen X (2017). The how and where of plant micro RNAs. New Phytol., 216(4): 1002-1017. https://doi.org/10.1111/nph.14834

Zhang J, Terrones M, Park CR, Mukherjee R, Monthioux M, Koratkar N, Bianco A (2016). Carbon science in 2016 Status challenges and perspectives. Carbon, 98(70): 708-732. https://doi.org/10.1016/j.carbon.2015.11.060

Zheng R, Zhang S, Zeng H, Wang S, Sun K, Chen R, He J (2022). Cancer incidence and mortality in China 2016. J. Natl. Cancer Center, 2(1): 1-9. https://doi.org/10.1016/j.jncc.2022.02.002

Zhou Z, Liu X, Sun K, Lin C, Ma J, He M, Ouyang W (2019). Persulfate-based advanced oxidation processes (AOPs) for organic-contaminated soil remediation A review. Chem. Eng. J., 372: 836-851. https://doi.org/10.1016/j.cej.2019.04.213

Zhu N, Zhang D, Wang W, Li X, Yang B, Song J, Tan W (2020). A novel coronavirus from patients with pneumonia in China, 2019. N. Engl. J. Med., 382(8): 727-733. https://doi.org/10.1056/NEJMoa2001017

Zhuang F, Qi Z, Duan K, Xi D, Zhu Y, Zhu H, He, Q. (2020). A comprehensive survey on transfer learning. Proceed. IEEE, 109(1): 43-76. https://doi.org/10.1109/JPROC.2020.3004555