 Kurdistan Journal of Applied Research (KJAR) Print-ISSN: 2411-7684 | Electronic-ISSN: 2411-7706 Website: Kjar.spu.edu.iq | Email: kjar@spu.edu.iq 61 Proceeding Conference (Not Peer Reviewed): 5th International Conference on the Health and Medical Science : Toward Gathering Iraqi Medical Powers (ICHMS 2021) Effect of Cannabis seeds on Some biomedical parameters in male rats Salah Omer Hamabor Hunar Mustafa Wassman Medical Laboratory Department Medical Lab analysis College of health and Medical Technology Cihan University- Sulaimani Sulaimani Polytechnic University Sulaimani, Iraq Sulaimani,Iraq Email: hunar.mustafa@sulicihan.edu.krd Email:salah.hamabor@spu.edu.iq Karwan Anwar Hassan Department of Biology College of educational Science/ Faculty education University of Garmian Kalar, Iraq Email:Karwan.anwar@sulicihan.edu.krd Article Info ABSTRACT Proceeding Conference (Not Peer Reviewed): 5th International Conference on the Health and Medical Science : Toward Gathering Iraqi Medical Powers (ICHMS 2021) (cannot be used for academic title promotion or for postgraduate students) Cannabis, sometimes known as hemp, is a plant that originated in Central Asia. Cannabis seeds nutritional profile is astounding. Cannabis seed, most digestible, balanced, natural, and complete supply of protein, amino acids, and necessary fats found anyplace in nature when it is in its pristine organic natural condition. Cannabis seeds can be used in a variety of medical fields. Each 100 gm of the seeds used in the study contains 26, 37 and 20 gm of protein, fat and carbohydrate respectively. The present study used two groups of male rats (control and treatment groups). Rats were 11 weeks of average age. 10 mg/kg of body weight of powdered cannabis seeds were administrated orally to treatment group daily for 15 days. Alkaline Phosphatase, glutamic oxaloacetic transaminase and glutamic pyruvic transaminase (ALP, GPT, and GOT), creatinine and uric acid was done for estimating the effect of cannabis seeds on renal and liver function. Enzymes Alkaline phosphatase, glutamic oxaloacetic transaminase and glutamic pyruvic transaminase (ALP, GPT, and GOT) as well as creatinine, uric acid and very low-density lipoprotein (VLDL) were not significantly (p<0.05) different in both groups. Level of Cholesterol, triglyceride and c-reactive protein were significantly (p<0.05) reduced. Serum total protein, albumin and globulin Keywords: cannabis seeds, renal function, liver function, anti-inflammatory effect. | 62 were significantly (p<0.05) elevated in treatment group comparing to control group. The aim of this study is to examine physiological impact of cannabis whole seeds. Identifying the relationship(s) between liver and renal functions and cannabis seeds, as well as the anti- inflammatory effect of cannabis seeds and possible of using cannabis seeds as herbal medicine for health care in male rats. Copyright © 2019 Kurdistan Journal of Applied Research. All rights reserved. 1. INTRODUCTION Approximately 70000 unique plant species had been used at the smallest amount as soon as in the records of conventional medication. World health organization (WHO) has suggested that about 80% of the populace used at the smallest amount one natural medication of their lifestyles for medical purposes. Herbal medicine and plants were used for remedy of the many unique forms of sicknesses consisting of liver, kidney, coronary heart sicknesses, etc.… therefore, scientists around the world have studied unique sorts of herbal plant in phrases of chemical and physiological properties, indications, and their side effects. Cannabis species is one among the herb plants that has been studied by many scientists in vitro round the world [1]. Cannabis sativa, typically called hemp, could be a reasonably cannabis. it's one in every of the oldest cultivated plants, and determinative its specific origin is tough thanks to its long history of cultivation. cannabis is a green leafy plant with distinctive opposing, typically 7-fingered, lance-shaped leaves that's dioecian (male and feminine flowers in numerous plants[2]. Cannabis sativa L. is a popular herbaceous plant from central Asia that has been utilized in traditional medicine, it can be used as a drug (marijuana) or a non-drug (marijuana) (hemp). The former is often used for different purposes such as recreational and medical purposes, whilst the latter is critical to the fiber and food industry [1]. Different preparations of Cannabis sativa have been used to cure a multitude of illnesses as traditional medicine in Asian, including diarrhea, inflammation, headache, nausea, hematochezia and alopecia. Cannabis sativa has anti-inflammatory, analgesic, antipyretic, and antidiarrhea properties.[1, 3, 4]. Due to a variety of factors, includes unlawful cultivation ,diversity of active components, and low quantity of certain of them in plant[5], Cannabis phytochemical study has been restricted, as has broad medicinal usage of Cannabis products. Cannabinoids, such as tetrahydrocannabinol (THC) and cannabidiol (CBD), are psychoactive components of cannabis plants that may be found in the flower, stems, stalks, and leaves, but they are not present in seeds until they are transferred from the flower to the seed's outer shell[6]. Liver and kidney are two active organs in the body. The functions of liver and kidney most evaluate while using any medicine and herbal (natural) medicines. Estimation of normal kidney and liver can be done via their activity, functions and cells. Any damage or injury in the cells have direct effects on kidney and liver functions. Liver function can be estimated via the level of protein, albumin, globulin, cholesterol and triglyceride. One the other hand any necrosis or injury if occur in hepatic cells (liver cells) can be determining by liver enzymes in serum especially serum GPT, ALP and GOT. As it is well known that liver is a site for detoxification, clearing blood from poisoning and drugs. Kidney is another organ can be determining its functions using cholesterol and uric acid. Any drug can be having direct effects on renal and liver functions or lead to cells damage. So that any herbal drugs should estimate its toxicity and its effect on liver, renal cells and functions. Cannabinoids and the endocannabinoid system appear to be essential in the treatment of liver disorders and normal liver function. Cirrhosis (liver scarring) and fatty liver disease are examples of this (FLD)[7]. People with viral liver disease may benefit from cannabinoids to help them cope with traditional therapies like chemo[8]. Following the legalization of cannabis | 63 in certain countries, such as Canada and the United States, there is likely to be a surge in interest in medicinal cannabis, particularly for chronic refractory symptoms and palliative illnesses like those seen in CKD patients[9]. Hempseed is usually referred to be one of the most nutritionally complete food sources due to its high nutritional characteristics. Hempseed may be fed on whole (hulled seed) or deshelled (hempseed kernel), in addition to in processed meals like as oil, flour, and protein powder[10]. Despite the fact that genotypes and environmental variables have revealed a wide range of hempseed compositions, it generally contains 25–35 percent lipids with a unique and precisely balanced fatty acids (FAs) composition; 20–25 percent easily digestible proteins rich in important amino acids; and 20–30 percent carbohydrates, the majority of which is dietary fiber, mostly in the form of dietary fiber, Hemp protein is made up mostly of globulin and albumin, which are distinguished by their high levels of arginine and glutaminic acid. The modulation of organ function and human metabolism was shown to be positive with these two types of hemp protein. Hemp seed protein also contains a number of antioxidative bioactive peptides. Hemp seeds, on the other hand, include antinutritional elements such as phytic acid and trypsin inhibitors[11]. Oil makes around 30 to 35 percent of hempseed, with unsaturated fatty acids accounting for 90 percent of the total. Essential fatty acids, linolic acid, omega-3 fatty acid, and monounsaturated fatty acid dominate hempseed oil. Linolic acid is necessary as a precursor for the formation of dihomo-linolenic acid (DGLA) and arachidonic acid, whereas -linolenic acid is required for omega-3 fatty acid assembly. These fatty acids have been studied extensively for their ability to protect against cardiovascular disorders, Hempseed's outer shell contains the majority of the carbs, while dehulling eliminates three-quarters of the fiber content. Statistics on the characteristics of hempseed fiber are currently lacking. Hempseed is also high in polyphenols, which include hydroxycinnamic acid in particular [2]. Few researches have looked at the impact of cannabis seeds in the diets of animals and humans on physiological, immunological, and oxidative state and health, so that the study aimed to find the effects of cannabis seeds on liver and renal function and also the possible of using cannabis seeds as herbal medicine for health care. 2. METHODS AND MATERIALS Two groups of male rats were designed to be a control group and treatment group. The average age of rats was 245  10 gm. Each group were contained 5 male rats. The animals had been uncovered to a 12hours light-darkish cycle and treated constant with popular protocols. The animals had been kept in plastic cages and had been allowed adaptation duration of seven days in a good ventilated room with a temperature of 28±2C0. Cannabis seeds were obtained from PK cannabel company in Russia-Moscow. Seeds were crushed using special grinder to be in powder form. 10 mg/kg body weight of powdered cannabis seeds were suspended in 1 ml of distal water and gave to treatment group orally for 15 days used special gavage tube. The animals had been anesthetized using special jar containing cotton saturated with chloroform and diethyl- ether in ratio 1:1 and blood sample were collected using cardiac puncture method[12]. Sera were stored at -20 C0. Biochemical tests were done using cobas c 311 (Roche diagnostics, Reagent were used originally from Roche company, Germany-2018) for both groups. Tests were included liver function test: Serum GOT, serum GPT and serum ALP. Renal function: creatinine (SCr) and uric acid (SUA). Serum globulin, albumin, total protein, cholesterol, VLDL and triglyceride (TG) levels also were estimated. C- reactive protein (CRP) levels were done using titration method by using cobas c 311 (Roche diagnostics, Reagent were used originally from Roche company, Germany-2018). The reagents used for GPT were contain TRIS buffer, L- alanine, albumin (bovine), LDH (microorganisms), stabilizers; preservative, 2-Oxoglutarate, NADH in different concentrations according to the manufactory. ALP reagent compost of 2-amino-2-methyl-1-propanol, magnesium acetate, zinc sulfate, N-(2-hydroxyethyl) -ethylenediamine tri-acetic acid, p-nitrophenyl phosphate. Materials used in GOT kits were: TRIS buffer, L-aspartate, MDH (microorganism), LDH (microorganisms), albumin (bovine), NADH and 2-oxoglutarate. Serum total protein kits contain Sodium hydroxide, potassium sodium tartrate, potassium iodide, copper sulfate. Reagent used for estimation of serum albumin was contain Buffer; preservatives; surfactants, BCP, buffer; | 64 preservatives; surfactant and the materials used in kits for serum globulin materials were TRIS/HCl buffer, NaCl, EDTA; preservative, latex particles coated with polyclonal anti-human β2-microglobulin antibody (rabbit). PIPES buffer, Mg2+, sodium cholate, 4-aminophenazone, phenol, fatty alcohol polyglycol ether, cholesterol esterase (Pseudomonas spec.), cholesterol oxidase (E. coli), peroxidase (horseradish) were used in serum cholesterol reagent. Materials used to estimate serum TG concentration were PIPES buffer, Mg2, sodium cholate, ATP, 4-aminophenazone, 4-chlorophenol, lipoprotein lipase (Pseudomonas spec.), glycerol kinase (Bacillus stearothermophilus), glycerol phosphate oxidase (E. coli) and peroxidase (horseradish). For VLDL the Dilution buffer: potassium dihydrogen-phosphate, dipotassium hydrogen-phosphate, potassium chloride, sodium aside, magnesium sulfate heptahydrate, sodium phosphor-tungstate n-hydrate Lipoprotein-lipase, cholesterol esterase, diaphorase, nicotinamide adenine dinucleotide, tetrazolium salt, glycerol dehydrogenase, cholesterol dehydrogenase used in reagent. CRP reagent was contained HEPES buffer, Anti-human CRP antibody (goat) and Latex-conjugate. The statistically analyzed data was given as the mean SD of two groups, with the means compared using IBM SPSS 26 software. At P0.05, the differences were statistically significant [13]. 3. RESULTS The results show the impress of Cannabis seeds on a number of biomedical or bio-chemical markers using male Albino rats. Table 1 showed the effect of Cannabis seeds on serum total protein, serum albumin and serum globulin of male rats, though, treatment group showed alteration significantly (P<0.05). The serum GPT, GOT and ALT in both groups showed no significant (P<0.05) difference when we compared with the control group as showed in table 2. Effect of cannabis seeds on the level of serum creatinine and serum uric acid in control and treatment group was not significantly (P<0.05) different (table 3). The cholesterol and triglyceride levels showed significant (P<0.05) decrease in treatment group when compared with the control group while VLDL showed no significantly (P<0.05) changed in treatment and control groups (table 4). The influence of Cannabis seeds for C- reactive protein in male rats showed reduction in treated group significantly (P<0.05) (table 5). Table 1: Effects of cannabis seeds on the level of serum total protein, serum albumin and serum globulin in (gm/dL) in control and treatment group (mean  SD). Groups Total protein Albumin Globulin Control 6.56 0.16 3.52 0.14 2.66 0.57 Treatment 7.47 0.33* 4.49 0.40* 4.25 0.28 * * The levels are significantly higher (P0.05) in treatment group compare to control group. Table 2: Effects of cannabis seeds on the level of serum GPT, serum GOT and serum ALT in (U/L) in control and treatment group (mean  SD). Groups SGPT SGOT SALT Control 37.03  3.32 117.73 11.57 244.33 60.01 Treatment 39.9  8.73NS 120.05 30.55NS 237.5 44.20NS NS: No significant different between control and treatment groups. Table 3: Effects of cannabis seeds on the level of serum creatinine and serum uric acid in (mg/dL) in control and treatment group (mean  SD). Groups Creatinine Uric acid Control 0.33  0.005 2.36  0.25 | 65 Treatment 0.37  0.031NS 2.38  0.35NS NS: No significant different between control and treatment groups. Table 4: Effects of cannabis seeds on the level of serum creatinine and serum uric acid in (mg/dL) in control and treatment group (mean  SD). Groups Cholesterol Triglyceride VLDL Control 73.03  3.20 123.8  14.89 25  3 Treatment 56.92  5.74* 71.85  10.40* 26  10.55NS * The concentrations are significantly lower (P0.05) in treatment group compare to control group. NS: No significant different between control and treatment groups. Table 5: Effects of cannabis seeds on the level of serum C- Reactive protein (U/L) in control and treatment group (mean  SD). Groups C-Reactive protein Control 0.93  0.13 Treatment 0.17  0.08* * The concentrations are significantly lower (P0.05) in treated group when compare to the control group. 4. DISCUSSION The liver has plain effect at the activities for various organs withinside the frame system. due to its key characteristic in metabolism (xenobiotic) and its gateway function withinside the frame system, it's miles vulnerable to xenobiotic-brought on damage [14]. In metabolism, detoxification, and biotransformation, the liver plays a critical function. As a result, changes in liver biomarkers and function tests may be utilized the extent assesses about harm or destroy as a result of intake cannabis seeds prior to biopsy [15]. Total protein in serum made up the summation of albumin and globulin, any change in the concentration of these fraction have effect on the concentration of total protein. The substantial (p<0.05) rise in Albumin, Globulin, and Total protein concentrations suggested that the powdered cannabis seeds boosted protein synthesis and/or mobilization (table 1). Masses of serum proteins, as well as provider proteins, enzymes, complement, and immunoglobulins, make up the globulin fraction. The majority of them are made by the liver, with the exception of immunoglobulins, which are made in plasma cells. More specifically, the observed rise in globulin level might be attributable to the cannabis seeds ability to generate antibodies. [16]. The greatest concentration of protein in the plasma is albumin. Albumin accounts for more than half of the total protein in serum. Within the intravascular compartment, about 30 to 40 percent of the body's total albumin pool is located. Can carries a large number of molecules throughout the body. The distribution of bilirubin, hormones, metals, vitamins, and medicines is aided by albumin. It plays a vital role in metabolism of fat through fatty acids binding and keeping them in a soluble state within the plasma. It prevents the blood fluid from seeping into tissues [17]. Part of the liver's general protein production is albumin. Chronic liver illness, such as cirrhosis or nephrotic syndrome, causes albumin levels to drop [18]. As a result, the remarked rise of albumin suggests that seeds (cannabis) might assist calcium in the bloodstream control the flow of water from the bloodstream into human tissue and may have a hepatoprotective impact [19]. This is also confirmed by the ratio of albumin to globulin result also substantial rise in protein content [20]. Some of tests assess how well the liver performs its daily functions of making protein and removing bilirubin, a blood waste. Other tests measure the amount of enzymes released by liver cells in response to injury or disease. Enzyme serum glutamic pyruvic transaminase (SGPT) is known to rise in the presence of liver illness and has been used to assess [21, 22]. Serum | 66 glutamic oxaloacetic transaminase (SGOT) is mostly found in gill cells, kidney cells, muscle cells, and liver parenchyma cells [23]. The mitochondrial and cytoplasmic forms of SGOT exist as a one-of-a-kind isoenzyme bureaucracy that may be genetically unique. SGOT is most prevalent in the coronary heart as compared to other body tissues such as the liver, striated muscle, and kidney. Elevated mitochondrial SGOT noticeable in widespread tissue necrosis throughout cardiac infarct and likewise in chronic liver illnesses like hepatic tissue degeneration and necrosis [14]. Alkaline phosphatase is found in the cell membrane and endoplasmic reticulum, or the liver biliary ducts cell lining, bone and placental tissue [24]. No significant different between control and treatment male rats indicate the powdered cannabis seeds at 10 mg/kg body weight for 15 days are safe for liver, muscle and kidney cells (table 2). Serum creatinine and uric acid levels are traditional indicator for renal function [25]. Normally serum creatinine levels can also be used to gauge how fast the kidneys filter blood (glomerular filtration rate). Because blood creatinine levels vary so much from person to person, the glomerular filtration rate may offer a more realistic picture of renal function[26]. Serum creatinine is a popular measure for identifying changes in glomerular filtration rate (GFR) and chronic kidney disease (CKD) stage, with increased serum creatinine indicating renal disease and reduced renal function[27]. In the same way, CKD patients frequently have increased serum uric acid levels. It's a straightforward biochemical diagnostic for kidney function that's either impaired or pathological[27]. Because renal clearance of serum uric acid is generally decreased after kidney injury or failure, renal function is the major confounder in studies investigating the connection between the serum uric acid and CKD progression[28]. The glomerular filtration rate is calculated using a formula that takes into consideration the serum creatinine counts as well as other variables including age and gender[29, 30]. Taking into consideration in this study the rats were in the same average ages and were male. The result shows there are slight elevating in the level of creatinine and decreasing the level of uric acid, but the changes were not significant in both parameters. That means there are no effect on renal functions. (table 3). The result shows decreasing in the level of serum cholesterol and triglyceride (TG) significantly, but VLDL was changed not significantly (p<0.05) in treatment group (table 4). Both cholesterol and triglycerides (TG) serve important physiological functions in keeping the body healthy[31]. Cholesterol and TG are two indicators that may be used to assess cardiovascular risks since high cholesterol and TG levels can cause health problems, particularly those connected to the heart[32]. Many researches reveal that triglycerides are more and more turning into diagnosed as a hazard issue for cardiovascular disorder, emphasizing the pressing want for scientific trials to decide if reducing blood triglyceride tiers decreases disorder hazard. [33, 34]. VLDL is produced in the liver and is in charge of transporting triglycerides to body cells, which are required for cellular activities. VLDL is made up less of fat and more of protein when triglycerides are transported to cells, leaving cholesterol on the molecule. VLDL's protein composition varies from that of chylomicrons in case the main protein structure is full- duration apo B (apo B100) rather than the truncated apo B48 version. VLDLs, like chylomicrons, are triglyceride-removing substrates for lipoprotein lipase. According to our findings, there is no influence on VLDL's regular functioning[35]. Decreasing cholesterol and triglycerides gives good clue that cannabis seeds can be used to decrease cardiovascular risks, but not significant change in the level of VLDL may due to short duration of taking powdered cannabis seeds which was 15 days. C-reactive protein (CRP) for treatment group was significantly decreased (p<0.05) (table 5). This may give us an idea that cannabis seeds act as anti-inflammation in the body. Liver produces CRP in response to a number of inflammatory cytokines[36]. CRP levels rise quickly in reaction to trauma, inflammation, and infection, and then fall just as quickly after the problem is resolved[37]. As a result, the reduction in CRP levels in the treatment group indicates that the cannabis seeds' anti-inflammatory properties are effective. 5. CONCLUSION | 67 It might also additionally consequently be too hasty to end that the drug become in charge of liver or hepatorenal toxicity as discovered withinside the serum enzymes. More research is exceedingly vital which can be capable of recognize absolutely the various outcomes and mechanisms of cannabis on liver and renal functions. Using 10 mg/ kg of body weight of cannabis whole seeds are safe for liver and kidney, orally intake for 15 days. Cannabis seeds have incredible effects on decreasing cardiovascular risk. Also have good roles in protein metabolism and synthesize as well as albumin and have role in strengthen the immune system via elevation of globulin level. CRP is a marker used for general inflammations in the body. Decreasing the level of CRP, indicate the anti-inflammatory effect of cannabis seeds. 6. 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