Investigation of minerals, testosterone, and transaminases in the semen and serum of fertile and infertile men belongs to different age groups

doi:10.4236/abb.2012.35074

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Advances in Bioscience and Biotechnology, 2012, 3, 574-579 ABB

http://dx.doi.org/10.4236/abb.2012.35074 Published Online September 2012 (http://www.SciRP.org/journal/abb/)

Investigation of minerals, testosterone, and transaminases

in the semen and serum of fertile and infertile men belongs

to different age groups

Zahed Mahmood1*, Muhammad Riaz1, Imtiaz Mahmood Tahir1, Abdul Shakoor1, Zia Ur Rahman2,

Muhammad Usman Qamar Saeed1

1Department of Chemistry and Biochemistry, University of Agriculture, Faisalabad, Pakistan

2Department of Physiology and Pharmacology, University of Agriculture, Faisalabad, Pakistan

Email: *[email protected]

Received 24 June 2012; revised 28 July 2012; accepted 10 August 2012

ABSTRACT

The present study was aimed to assess the potential of

infertility to induce the adverse effects with reference

to testosterone, Triiodothyronine (T3), Thyroxine (T4),

Alanine Aminotransferase (ALT), Aspartate Ami-

notransferase (AST), zinc, copper and iron. All the

samples were divided into four groups according to

age and disorder (Group 1, 10 infertile men of 25 - 40

years; Group 2, 10 fertile men of 25 - 40 years; Group

3, 10 infertile men of 41 - 60 years and Group 4, 10

fertile men of 41 - 60 years). Semen and blood sam-

ples were analyzed by atomic absorption spectropho-

tometry to determine minerals while, Testosterone, T3

and T4 were determined by enzyme immunoassay

kits. ALT and AST were determined using standard

kit assay method. The levels of testosterone and T3

and AST in the fertile semen of 41 - 60 years age

group were increased significantly (P ≤ 0.001) as

compared to that of fertile semen of 25 - 40 years age

group. While, the level of T4 in the fertile semen of 41

- 60 years age group was decreased significantly (P ≤

0.001) as compared to that of fertile semen of 25 - 40

years age group. In case of fertile serum, only the

level of testosterone was significantly decreased (P ≤

0.05) in the 41 - 60 years age group as compared to 25

- 40 years age group. The levels of testosterone and

Cu in the infertile serum of 41 - 60 years age group

were decreased significantly (P ≤ 0.001). While, the

levels of T3, T4, ALT and Fe in the infertile serum of

41 - 60 years age group were increased significantly (P

≤ 0.05) as compared to that of infertile serum of 25 -

40 years age group.

Keywords: Semen; Reproduction; Infertility; Minerals;

Testosterone; Transaminases

1. INTRODUCTION

Infertility primarily refers to the biological inability of a

person to contribute to conception. According to World

Health Organization [1], infertility is a period of two

years without conception. After one year of infertility

many couples seek a medical opinion because modern

artificial reproduction techniques like intra-cytoplasmic

sperm injection (ICSI) can help couples to overcome

infertility [2]. Sub fertility is the failure to conceive after

1 year of regular, unprotected intercourse with the same

partner. Sub fertility is caused by sperm defects or dys-

function [3].

Testosterone is a principal male sex hormone and is

primarily secreted in the testes of males. In men, testos-

terone plays a crucial role in the development of male

reproductive tissues such as the testis and prostate as

well as promoting secondary sexual characteristics such

as increased muscle, bone mass and the growth of body

hair [4]. The thyroid hormones, T3, and T4 are tyrosine

based hormones formed by the thyroid gland mainly re-

sponsible for regulation of metabolism [5]. ALT and AST

are enzymes located in liver cells that leak out into the

general circulation during the injury of liver cells [6].

Among the metal trace elements Zinc is one of the

most interesting nutritional traces in the reproductive

system. In man it is a cofactor of more than 200 met-

allo-enzymes and plays an important role in the normal

testicular development, spermatogenesis, and sperm mo-

tility [7]. Copper is an essential trace element required in

the diet because it is the metal cofactor for a variety of

enzymes (amine oxidase, superoxide dismutase, cyto-

chrome oxidase and tyrosinase). However, excess copper

can cause problems because of its capacity to oxidize

proteins and lipids, bind to nucleic acids and enhancing

the production of free radicals. Therefore, it is important

to maintain the amount of copper in the body within

normal limits. Copper reduces sperm motility by reduc-

*Corresponding author.

OPEN ACCESS

Z. Mahmood et al. / Advances in Bioscience and Biotechnology 3 (2012) 574-579 575

ing oxidative processes and glucose consumption [8].

Iron is a vital component of a group of heme proteins

that function in oxygen transport or as enzymes in redox

systems. A small amount of iron is enclosed in several

non heme metalloenzymes. The chief complexes coordi-

nating iron with the cell are heme and heme containing

proteins, hemosiderin and ferritin [9,10]. The aim of this

study was to assess the potential causes of infertility and

old age to induce the adverse effects with reference to

testosterone, T3, T4, ALT, AST, zinc, copper and iron.

2. MATERIALS AND METHODS

2.1. Sample Collection

Semen and blood samples were collected from each in-

dividual patient of each group. Semen samples were col-

lected by masturbation in a clean specimen container

after sexual abstinence for 3 - 5 days followed by lique-

faction at 37˚C. The semen samples were evaluated ac-

cording to World Health Organization recommendations

(ejaculate volume, pH, time to liquefaction, sperm con-

centration, motility and morphology). The remaining se-

men samples were centrifuged at 3000 × g for 10 min to

obtain the seminal plasma. The separated seminal plasma

was stored at –80˚C until further analysis for the detec-

tion of testosterone, Zn, Cu, Fe, T3, T4, ALT and AST

level. Blood samples were centrifuged at 4000 × g for 15

min after complete coagulation and supernatant serum

was collected in Eppendorf tubes and frozen until further

analysis for the detection of testosterone, Zn, Cu, Fe, T3,

T4, ALT and AST level.

2.2. Measurement of Minerals

Zinc, iron and copper in the samples were measured by

atomic absorption spectrophotometer. The samples were

aspirated to the instrument and reading was recorded at

the Monochromator wavelength 213.9, 285.2 and 342.7

nm for zinc, iron and copper respectively.

2.3. Measurement of Testosterone

Testosterone was determined with the help of DRG Tes-

tosterone Enzyme Immunoassay Kit.

2.4. Measurement of T3 & T4

Triiodothyronine (T3) and Thyroxine (T4) were deter-

mined by using J D Biotech T3 and J D Biotech T4 En-

zyme Immunoassay Kit respectively.

2.5. Measurement of ALT

Human Gasellschaft fur Biochemica and diagnostica

mbH (EC 3.1.3.1) kit was used to determine the level of

ALT as described by Saher et al. [11].

2.6. Measurement of AST

The AST was evaluated by adopting the methodology as

described by Reitman and Frankel [12].

2.7. Statistical Analysis

The results were expressed as mean ± SEM for all con-

tinuous variables. Mean ± SEM have been prepared in

Microsoft® Excel Windows Version 2007. The data ob-

tained were analyzed by applying student t-test using

Minitab 15 Windows Version to assess the significant

differences between all selected age groups.

3. RESULTS AND DISCUSSION

The levels of testosterone and T3 and AST in the fertile

semen of 41 - 60 years age group were increased signifi-

cantly (P ≤ 0.001) as compared to that of fertile semen of

25 - 40 years age group. While, the level of T4 in the

fertile semen of 41 - 60 years age group was decreased

significantly (P ≤ 0.001) as compared to that of fertile

semen of 25 - 40 years age group. There was no signifi-

cant difference among the levels of ALT, Zn, Cu and Fe

between the fertile semen of 25 - 40 years age group and

41 - 60 years age group (Figure 1).

In case of fertile serum, only the level of testosterone

was significantly decreased (P ≤ 0.05) in the 41 - 60

years age group as compared to 25 - 40 years age group.

While, the levels of T3, T4, ALT, AST, Zn, Cu and Fe

had no significant difference between the fertile serums

of 25 - 40 years age group and 41 - 60 years age group

(Figure 2).

In the infertile semen of 41 - 60 years age group, the

levels of testosterone, T4, Zn and Cu were decreased

significantly (P ≤ 0.001, P ≤ 0.05) as compared to that of

infertile semen of 25 - 40 years age group. While, the

levels of T3, ALT and AST were increased significantly

(P ≤ 0.001, P ≤ 0.05) in the infertile semen of 41 - 60

years age group as compared to that of infertile semen of

25 - 40 years age group (Figure 3).

The levels of testosterone and Cu in the infertile serum

of 41 - 60 years age group were decreased significantly (P

≤ 0.001) as compared to that of fertile semen of 25 - 40

years age group. While, the levels of T3, T4, ALT and Fe

in the infertile serum of 41 - 60 years age group were in-

creased significantly (P ≤ 0.001, P ≤ 0.05) as compared to

that of infertile serum of 25 - 40 years age group (Figure

4).

4. DISCUSSION

The lower level of testosterone in the fertile serum of 41

- 60 years age group as compared to that of fertile serum

of 25 - 40 years age group indicates that testosterone

level is decreased in older age resulting in the decreased

Z. Mahmood et al. / Advances in Bioscience and Biotechnology 3 (2012) 574-579

576

Figure 1. Biochemical parameters evaluation in fertile semen of age group 2 & 4.

Figure 2. Biochemical parameters evaluation in fertile serum of age group 2 & 4.

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Z. Mahmood et al. / Advances in Bioscience and Biotechnology 3 (2012) 574-579 577

Figure 3. Biochemical parameters evaluation in the infertile semen of age group 1 & 3.

Figure 4. Biochemical parameters evaluation in the infertile serum of age group 1 & 3.

Z. Mahmood et al. / Advances in Bioscience and Biotechnology 3 (2012) 574-579

578

sperm production. In consequence, the testosterone con-

centration is increased in the fertile semen of 41 - 60

years age group as compared to that of fertile semen of

25 - 40 years age group to increase the sperm production.

Significantly lower level of testosterone in the infertile

serum of 41 - 60 years age group represents that testos-

terone deficiency is more pronounced in older age. This

may be due to atrophy of glandular tissue of pituitary or

testes resulting in significantly lower level of testoster-

one in the infertile semen of 41 - 60 years age group.

This may also be due to prostatic inflammation and ob-

struction or blockage of the male reproductive tract.

Hence testis not functioning properly and testosterone

production is affected. Zinc deficiency in infertile semen

of 41 - 60 years age group also causes lower testosterone

production as zinc is involved in steroidogenesis, tes-

ticular development and testosterone synthesis [3]. The

similar results were found by Khan et al. [13]. They ob-

served the decrease in testosterone concentration in the

infertile individuals. Omrani et al. [14] and Weedin et al.

[15] also found the decreased testosterone level in the

infertile men.

OPEN ACCESS

[3] Ebisch, I.M.W., Thomas, C.M.G., Peters, W.H.M., Braat,

D.D.M. and Steegers-Theunissen, R.P.M. (2007) The im-

portance of folate, zinc and antioxidants in the patho-

genesis and prevention of subfertility. Human Reproduc-

tion Update, 13, 163-174.

Significantly lower level of zinc in the infertile semen

of 41 - 60 years age group in the present study might be

due to excessive ROS production. After ejaculation the

abnormal spermatozoa are sources of oxidants which

bind with the zinc and reduce its concentration in semi-

nal plasma. Hence, zinc is considered to be an important

antioxidant protecting the spermatozoa from oxidative

stress. Zinc deficiency decreases testicular weight and

causes shrinkage of the seminiferous tubules. Zinc defi-

ciency also induces atrophy of the seminiferous tubules

and causes failure of the spermatogenesis. Low seminal

zinc levels have been correlated with decreased fertility

potential [16]. Ali et al. [17] observed the low zinc level

in the infertile subjects and found that low zinc level

affected the semen parameters. Colagar et al. [18] found

that poor Zn nutrition may be an important risk factor for

low quality of sperm and idiopathic male infertility.

Significantly lower level of Copper in the infertile se-

men and serum of 41 - 60 years age group may be due to

liver toxicity as it is absorbed by the liver, bound to al-

bumin and excreted into the bile. It is also involved in

protein synthesis during certain enzymatic reactions act-

ing as a cofactor. Nutritional deficiency may also lower

its level in infertile men. Abdul-Rasheed [19] reported a

significant decrease in seminal plasma copper levels in

azoospermic patients. In the present research work, sig-

nificant increase of iron in the infertile serum might be

due to oxidative stress in infertile men. Aydemir et al.

[20] observed the increased iron level in the infertile men.

Eghbali et al. [10] reported that seminal plasma iron

content is related with the motility and viability of the

spermatozoa after ejaculation. Presence of iron in the

seminal plasma will assist spermatozoa to sustain their

functions as long as it is needed for semen fertility.

5. ACKNOWLEDGEMENTS

The authors express gratitude to the lab staff of Physiology and Phar-

macology Department, Faculty of Veterinary Sciences and Staff of Bio-

medical Lab, University of Agriculture Faisalabad, Pakistan. On pro-

viding lab facilities authors are also grateful to the Mian Muhammad

trust Hospital Faisalabad and Biocare Labs, Aziz Fatima Hospital Fais-

alabad, Pakistan.

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