Introduction

The capacity of a person to conceive through regular sexual activity is referred to as fertility. Normal fertility requires the production of enough healthy sperm by the male. Nutrients in almonds may aid in cancer prevention, heart health, bone health, fertility, as well as other areas. Almonds provide enough protein to satisfy one-eighth of a person's daily needs with just one ounce (Arthur et al., 2018). The almond tree is a member of the Rosaceae family. They are classified as either members of the Prunoideae or Amygdaloideae subfamily. Due to the widespread practice of assigning almonds to their own family (Prunaceae or Amygdalaceae), it is now often assumed that prunus evolved from the Spiraeoidae subfamily. The average height of an almond tree is 4-10 meters, as well as its trunk is roughly 30 centimeters in diameter. Young twigs are born a green tint, but as they age and are exposed to more sunlight, they turn purple. The average leaf length is three to five inches. The five-petaled, baby-pink blossoms are delicate and lovely. Once planted, it takes 5–6 years for the tree to achieve maturity, which occurs in the fall (Lee et al., 2017). The capacity to cultivate aesthetically pleasing Almond domestication made it one of the earliest fruit trees to be domesticated. By the early Bronze Age, around 2000 B.C., almonds had been domesticated. To remove toxicity, wild almonds were likely leached or roasted before being gathered for human consumption. Before there were farms and homes. However, cultivated Almonds in their sweet form are not poisonous. According to Diamond, early farmers grew a mutant characterized by a lack of glycoside amygdalin "at first unintentionally in the garbage heaps then more intentionally in their orchards" due to a common genetic mutation (Markiewicz-Zukowska et al., 2022). Two distinct varieties of almonds exist. Bitter almonds as well as sugared almonds. The sweet variety of almonds can be eaten, while the bitter variety is deadly. When compared to its sweet counterpart, the bitter almond is longer and wider. Half of the oil in a sweet almond is solid. Hydrogen cyanide may be extracted from Bitter Almond. The kernel, often known as the almond's meat, the middle shell, as well as the outer green shell make up an almond. The kernel coat, brown skin, or seed coat is a thin, leathery coating. The kernel provides most of the plant's calories. Vitamins, protein, minerals, and fiber can all be found in abundance in almonds. They can be consumed either raw or roasted. They're technically a drupe rather than a nut (Ozcan, 2023). Seed extract, skin, and hull phenolic chemicals have been isolated from almonds over the past few decades. Polyphenols, an abundant micronutrient in the human diet, have been revealed to protect against cancer also cardiovascular disease. In addition to a beneficial association between almonds and male fertility, the health benefits of polyphenols rely on the amount taken as well as their bioavailability (Creedon et al., 2023). Several phenolic compounds isolated from almonds and almond products are discussed, along with their antioxidant characteristics as well as potential applications as dietary antioxidants (Rajaram et al., 2023).

Aim of the Study

The main focus of this investigation is to ascertain the positive impacts of almond seeds found in the Al-Baha area in raising the level of fertility in subfertility rats.

Materials and Methods

Materials

Preparation of Almond Seeds

Almond seeds were obtained from a local market in the Al-Baha area and then properly cleaned, sliced into little pieces, in addition, oven-dried for three days at 50 degrees Celsius, before crushing and and grinding into a powder form.

Cadmium Chloride

was acquired with no modifications from the German chemical firm Merk and put into immediate service.

Experiential Animals

Thirty mature male Sprague Dawley rats weighing (175-170 g) were utilized in this investigation. The Medical Insects Research Institute in Doki, which is located in Cairo, is where we received these rats.

Rats were acclimated to laboratory life for a week by keeping them in wire cages and feeding them standard laboratory food. Rats were fed from specialized food cups to prevent spillage, and water was offered in glass tubes propped up to one side of the cage; both were available ad libitum and were examined regularly.

Biological Experiment

Rats' Normal Diet

The diet based on basil included vitamin mixture (1%), corn starch (69.5%) 10% casein, 10% maize oil, 5% cellulose, 0.25% choline chloride, 0.35% methionine,  and 4% salt mixture (Morsi, 1992).

Test diet baseline components included Fe(C₆H₅O₇) 26H₂O (55 mg), MgSO₄.2H₂O (20₄ mg), CaCO₃ (600 mg), NaCl (334 mg), CaHPO₄.2H₂O (150 mg), K₂HPO₄ (645 mg), ZnCl₂ (0.5 mg), CuSO₄.5H₂O (0.06 mg), MnSO₄.4H₂O (10 mg),  and K_l (1.6 mg) (Hegsted et al., 1941).

The experimental baseline diet included Calcium pantothenic acid (0.40 mg), Vitamin K (0.50 Iu), Vitamin B12 (2.00 g), Vitamin E (10 Iu), Pyridoxine (1.00mg), Thiamin (0.50 mg), Folic acid (0.02 mg), Vitamin D (100 Iu), Niacin (4.00 mg), Para-amino – benzoic acid (0.02 mg), Vitamin A (200 Iu), Inositol (24 mg), Choline chloride (200 mg), (Campbell, 1963).

Diet Experiment

Table 1 illustrates the trial diet, which is ten percent of the basic diet enriched with plant powders.

Table 1. The basic and experimental diets' compositions.

Induced Subfertility for Rats

Cadmium chloride (0.1%) injections at 1ml/kg body weight caused male subfertility in rats (Kini et al., 2009).

Rats

Albino Sprague-Dawley male rats of adulthood, with a body weight (B.Wt) of 150-160 g. The animals were taken from the Animal Laboratory when they were fourteen to sixteen weeks old. The samples were housed in neat, well-maintained plastic cages with stainless steel tops. Before the experiment, rats were fed the basal diet for one week to allow them to adjust. Ad libitum water was available from a small-mouth bottle attached to a metallic tube and a plastic tubing at the mouth. For seven days before we began with the research, the rodents were trained to a schedule of twelve hours of sunlight followed by the same hours of darkness, as was previously mentioned.

Experimental Design

The rats were kept in standard conditions, in wire cages at 25 degrees Celsius. Rats were separated into the following groups:

Group 1: Neutral control group consisting of six healthy rats. The rats in this group were fed the typical rodent diet and given only tap water.

Group 2: Subfertility induced rats (24 rats). These rats subdivided into 4 subgroups for eating up the diets in trials for (four) weeks consistent with the following:

Group 2: Six rats: Positive control group (untreated group).

Group 3: Six rats: Infertility rats fed on basal diet +5% almond seeds powder.

Group 4: Six rats: Infertility rats fed on a basal diet of +10% almond seeds powder.

Group 5: Six rats: Infertility rats fed on basal diet +15% almond seeds powder.

Biological Evaluation

Each day for the 28-day study, the amount of food consumed was noted, and body weight was recorded each week. The feeding efficiency ratio (F.E.R.), the body weight growth (B.W.G. %), and the organ weight were all calculated (Chapman et al., 1959).

Blood Sampling

After the trial, samples from the blood were taken from the participants after they had fasted for twelve hours. After taking blood samples with highly specialized glass tubes using the retro-orbital approach, the blood samples were left to clot at room temperature (37 degrees Celsius) for a period of half an hour. Serum was extracted from blood samples by centrifuging them for 10 minutes at 3000 rpm before glucose testing. To prepare for analysis, we carefully aspirated the residue, transferred it to sterile polypropylene tubes with secure caps, and then froze it at -20 degrees Celsius.

The kidney, liver, heart, and, spleen were collected, rinsed in a salt solution, measured, as well as stored in ten percent formalin as instructed (Drury & Wallington, 1980).

Biological Evaluation

Body weight gain % (BWG%), Feed efficiency ratio (FER), food consumption, and feed efficiency ratio. Using the equation following (Chapman et al., 1959):

Analytical Biochemistry

• • • • Measurement of Liver Enzyme Activity
      • Measurement of (AST) Aspartate Aminotransferase Activity

According to Reitman and Frankel (1957), the aspartate aminotransferase enzyme was measured with a spectrophotometer along with specific kits (BioMerieux).

• • • • Measurement of Serum (ALT) Alanine Aminotransferase Activity

Reitman and Frankel (1957) established a calorimetric method for evaluating alanine aminotransferase enzyme activity.

• • • • Evaluation of Serum (ALP) Alkaline Phosphatase Activity

Based on Roy's method from 1970, the colorimetric measurement of alkaline phosphatase was completed.

• • • • Determination of Follicle-Stimulating Hormone (FSH)

Follicle-stimulating hormone was identified colorimetrically according to the method of (Fahim et al., 1982).

• • • • Determination of Luteinizing Hormone (LH)

Luteinizing hormone was determined colorimetrically following the method of (Fahim et al., 1982).

• • • • Determination of Testosterone Hormone

The levels of the hormone testosterone were measured colorimetrically using the technique of (Pardelles et al., 1985).

Histopathological Examination

After the time of the experiment had passed, the animals were sacrificed, then liver samples were collected, after which they were fixed in neutral formalin at a concentration of ten percent, dried in ethyl alcohol, purified in xylene, and at last embedded in paraffin wax. Hematoxylin staining, in addition, was carried out on slices that were around four and six microns thick (Carleton & Maycock, 1978).

Statistical Analysis

The statistical analysis was performed with a one-way classification as the basis for the computations. Snedecor and Cochran (1967) proposed an analysis of variance (ANOVA) that made use of the Least Significant Difference (LSD).

Results and Discussion

The initial goal of this trial was to estimate the positive effects of almond seeds found in the Al-Baha area in raising the level of fertility in subfertility rats.

Biological Changes

Influence of Almond Seeds on the Feed Efficiency Ratio, Body Weight Gain (g), and Feed Intake (g/d) in Subfertility Rats

Statistics offered in Table 2 indicate the mean values of (FER) in addition to (FI) (g/d) for the negative control, (BWG) (g/d), positive control, and other diverse groups of subfertility rats fed on different levels of almond seeds. It could be observed that a decrease in mean values of body weight gain and feed efficiency ratio of the control (+) group took place which were (0.11±0.034g and 0.004±0.001) respectively, while (FI) value was increased (30.87±2.93 g/d). Percent increase of control (-) for BWG and FER were (609.09% and 850%) respectively, with values being; (0.78±0.089g and 0.35±0.005) respectively, but a 33.95% decrease in (FI) occurred being; (20.39±1.08 (g/d).

Table 2. Shows the Influence of almond seeds on the feed efficiency ratio, BWG (g), and feed intake (g/d) in subfertility rats.

Effect of Almond Seeds Powder on Liver Enzymes (AST, ALT, ALP, and AST/ALT) (U/L) in Hepatic Rats

Data from Table 3 showed a significant rise in aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase activity, and AST/ALT for control positive groups which were (105.08±1.59 U/L, 50.93±2.47 U/L, 269.55±1.67 U/L and 2.06±0.082) respectively, compared to control negative group which were (76.23±1.28 U/L, 43.0±1.99 U/L, 171.5±2.32 U/L and 1.773±0.01) respectively.

Due to feeding on almond seeds powder (5%, 10%, and 15%) the significant decreases of (AST, ALT, ALP also aspartate aminotransferase/alanine aminotransferase) for rats were evident compared with the control (+ve)- group. Maximum decrease of (AST, ALT, and ALP) recorded to G4 (10% almond seeds) respectively. While for AST/ALT ratio, the lowest value was recorded for G3 (5% almond seeds) which was (1.25±0.019) but revealed nonsignificant differences between G5 (15% almond seeds) for AST/ alanine aminotransferase ratio which were (1.586±0.012).

Table 3. Effect of almond seeds powder on liver enzymes (AST, ALT, ALP, and aspartate aminotransferase/alanine aminotransferase) (U/L) in subfertility rats

Effect of Almond Seeds Powder on Sexual Hormones of Subfertility Rats

1. Testosterone Hormone (ng/ml)

Data in Table 4 illustrate the mean value of (testosterone) hormone (ng/ml) of subfertility rats fed on several diets. It could be observed that the mean value of (Testosterone) of the control () group was higher than control (+) group, being 2.35±0.01 and 0.38±0.05 respectively, designated significant variance with percent of increase +518.4% when compared to the control (+) group. All infertility rats fed on various diets revealed significant increases in mean values as compared to the control (+) group. The values were 0.86±0.02, 0.39±0.01 and, and 0.73±0.01 (ng/ml) for (5, 10 and and 15% almond seeds powder). The percent of increases were +126.3, +2.6, and +92.1 for groups 3, 4, and 5 respectively. Group 3 (5% almond seeds) recorded a better treatment of testosterone hormone as compared to the control (+) group.

2. Follicle Stimulating (FSH) Hormone (MLU/ ml)

The mean value of (FSH) of the control (-) group was higher than control (+) group, being 0.11±0.03 and0.27±0.01 respectively, Table 4 suggested significant alteration with a percent of increase +62.96% when compared to the control (+) group. Groups 3, 4, and 5 for (5, 10, and 15% almond seeds powder) revealed significant decreases in mean values as compared to the control (+) group. The measurements were 0.18±0.05 and 0.12±0.01 (MLU/ml). The percent of increases were +80.00, +20, and +50 for groups 3, 4, and 5 respectively. Group 5 (15% almond seeds) documented the best treatment of FSH hormone

3. Luteinising (LH) Hormone

The average level of luteinizing hormone (LH) hormone (MIU/ml) in subfertility rats fed the various diets is displayed in Table 4. It was discovered that there was a significant distinction between the control (+) as well as control (-) groups, with the control (-) group having a lower mean value of (LH) than the control (+) group (0.11±0.009 vs. 0.17±0.002), indicating a reduction of 35.30 percent. All subfertility rats fed on various diets revealed significant decreases in mean values as compared to the control (+) group. The values were 0.14±0.005, 0.115±0.0009, and 0.11±0.003 (MIU/ml) for (5, 10 and 15% almond seeds powder) respectively. The percent of decreases were -17.7, -32.4 and-35.3 for groups 3, 4, and 5 respectively. Group 5 (15% almond seeds) was found to be the better treatment of LH hormone.

Table 4. Effect of almond seeds powder on sexual hormones (Testosterone hormone, (FSH) hormone, and (LH) hormone) in subfertility rats.