NUTRITIONAL STUDIES ON PROMISING PLANTS FOR NEPHROPATHY TREATMENT OF MALE ALBINO RATS

INTRODUCTION

Chen et al.,(2006)claimed that almond provides with a nutrient-dense source of vitamin E, manganese, magnesium, copper, phosphorus, fibre, riboflavin, monounsaturated fatty acids and protein. Although almost 50% of almond weight is fat, incremental intakes of 7 g /day of this tree nut reduced low-density lipoprotein (LDL) cholesterol concentration by 1%, especially within the context of diets recommended by the National Cholesterol Education Program. Habitual almond consumption does not lead to weight gain, and their inclusion in low-calorie diets appears to promote more weight loss than a comparable carbohydrate-based low-calorie diet. Also, almonds have a low glycemic index and do not adversely impact insulin sensitivity. Almonds are an excellent source of bioavailable α-tocopherol,

Gray et al.,(2000) announced that Sambucus nigra (elder) has been documented as a traditional treatment of diabetes.. The results, demonstrated the presence of insulin-releasing and insulin-like activity in the traditional antidiabetic plant, Sambucus nigra.

Khan et al.,(2012)  said that Juniperus excelsa Bieb. is used in folk medicine for lowering blood pressure (BP). Its BP-lowering effect, endothelium-dependent and endothelium-independent vasodilator effects, and cardio-modulatory effect was stadied. The crude extract of J. excelsa (Je.Cr) induced a dose-dependent (10-300 mg/kg) fall in the arterial BP of anesthetized rats. In isolated rabbit aorta, Je.Cr (0.01-5.0 mg/mL) inhibited high K(+) (80 mM)- and phenylephrine (1 μM)-induced contractions, like that caused by verapamil and papaverine. In guinea pig atria, Je.Cr initially caused mild cardiac stimulation, followed by inhibition, like that exhibited by papaverine. These results reveal that cardiovascular effects of J. excelsa are mediated possibly through a combination of Ca(++) antagonism, nitric oxide-modulating mechanism, and phosphodiesterase inhibitory mechanism, which explain its medicinal use in hypertension.

Fellers et al.,(1990) reported that Grapefruit is an excellent source of many nutrients and phytochemicals that contribute to a healthy diet. Grapefruit is a good source of vitamin C,  contains the fiber pectin, and the pink and red hues contain the beneficial antioxidant lycopene. Studies have shown grapefruit helps lower cholesterol,and there is evidence that the seeds have antioxidant properties. Grapefruit forms a core part of the "grapefruit diet", which was with low glycemic index ,being able to help the body's metabolism burn fat. The grapefruit albedo needs to be investigated for amelioration of  nephropathy. The same conld be suggested for almond nut, elder and Juniperus excelsa.

MATERIALS AND METHODS

1- The used plants and their preparation:

The plants used in this study were:

• Almond  nut  (Prunus dulcis)

• Sambucus herbs(SP.L.)

• Juniperus seeds (SP.L.)

• Grapefruit albedo (Citrus paradise)                                                                                                                                 

The plants ,nuts&grapefruit obtained from supermarket and the special shop and then dried, milled and added to diet.   

״Biological experiment״

2-Basal diet composition of tested rats:

The basal diet consisted of casein (12 % ), corn oil (10%) , choline chloride (0.25%) , vitamin mixture (1%), cellulose (5%) , corn starch (up to 100%) and salt mixture (4%), according to ( Campbell, 1963), and the composition of salt and vitamin mixture were according to (Hegested et al., 1941and Campbell, 1963).

3-Preparation of Rats with Impaired Kidney:

Impaired kidney conld be induced in normal healthy male albino rats by intra-peritoneal injection of gentamicin(aminoglycosides antibiotics) obtained by Memphis Co. form Pharm .Chem.Itd.Cairo.A.R.E.at 10 mg/kg/day for 6days in which the nephrotoxicity,one of the adverse reaction of gentamicin takes place.

4-Experimental design and animal groups:

Thirty five (35) adult male albino rats, Sprague - Dawley strain, weighting (150 ±10 g) were used. All rats were fed on basal standard diet for seven consecutive days as adaptation period. Then, rats were distributed into 7 groups each of  5 rats in which means of rats weight for all groups were nearly equal. All the groups of rats were housed in wire cages and fed on the experimental diet for 4 weeks according to the following groups:

• Group (1): This group was fed on basal diet only as a negative control (healthy rats) (c-ve) (n=5).
• Group (2): This group was fed on basal diet only as a positive control (rats with impaired kidney by gentamicin) (c+ve) (n=5).
• Group (3): This nephritis group was fed on basal diet containing 5% Almond  nut.
• Group (4): This nephritis group was fed on basal diet containing 5% Sambucus herbs.
• Group (5): This nephritis group was fed on basal diet containing 5% Juniperus seeds.
• Group (6): This nephritis group was fed on basal diet containing 5% Grapefruit albedo.
• Group (7): This nephropathy group was fed on basal diet containing 5% mixture of plants powder

5-Biological evaluation:

During the experimental period (28days),the diet consumed  recorded every day, and body weigh recorded every week. The body weight gain (BWG g),feed efficiency ratio (FER),and also  organs weights determinated according to Chapman et al., (1959).

6-Urine and serum:

At the end of experiment period, the urine of each rats was collected for 24 hours and it was analyzed for determination the following parametera: Total protein, sodium and potassium.

Also, blood samples were collected after 12 hours fasting at the end of experiment in which the rats were scarified under ether anesthsia. Blood samples were received into clean dry centrifuge tubes and left to clot at room temperature, then centrifuged for 10 minutes at 3000 rpm to separate the serum. Serum was carefully aspirated and transferred into clean cuvette tubes and stored frozen at-20˚Cfor analysis. All serum samples were analyzed for determination of following parameters: urea, uric acid , creatinine, sodium & potassium, gluatmic oxaloacetic transaminase (GOT),and glutamic pyruvic transaminase (GOT), alkaline phosphatase (ALP) proiten , total cholesterol, triglycerides, high density lipoprotein and glucose.

At the same time the organs: Heart. kidney, lungs, liver, and spleen were removed ,cleaned, weighted, and stored frozen in formallin solution 10% for histopathological testing according to method mentioned by Drury and wallington(1980).

7-Analytical Methods:

.Determination of urea:

Urea was determination was according to the enzymatic method of Patton and Crouch(1977).

.Determination of uric acid:

           Uric acid was determination according to the enzymatic Colorimetric test of Fossatti and prencipe (1980).

.Determination of creatinine:

Creatinine was determinaed according to kinetic method of Henry (1974).

.Determination of serum glutamic oxaloacetic transaminases (GOT) and glutamic pyruvic transaminase (GPT) alkaline phosphatase (ALP),total protein:

GOT and GPT activities were measured according to method   described by Thefeld(1974) while determination of total protin according to the colorimetric method of Henry(1964).

.Determination of sodium:

Sodium was measured according to the colorimetric method of Henry (1974).

.Determination of Potassium:

Potassium was measured according to the colorimetric method of Henry (1964).

.Determination of serum lipid:

1-Triglcerides

Enzeymatic colorimtric determination of triglycerides was carried out accolrding to Fossati and Prencipe (1980)                                                                                                                          

2- Total cholesterol:

Total cholesterol was determined by colorimetric method accolrding to NIHP(1984).

3-HDL- cholesterol:

Determination of HDL was carried out according to the method of Fnedewaid (1972) and Gordon and Amer(1977).                                                                                                                               

4-VLDL&LDL- cholesterol:

The determination of VLDL (very low density lipoproteins) and LDL (low density lipoproteins)were carried out according to the methold of  Lee and Nieman (1996) by calculatioln as  follows:              

*VLDL (mg/dl)= Triglycerides/5

*LDL(mg/dl)=Total cholesterol –HDL- VLDL

5-Determinatioln of atherogenic index (AI):

   Atherogenic index was calculated according to Nakabayashi et al .,(1995) as follows:                     

Atherogenic index (AI) =    

8-Statistical analysis

The obtained data were statistically analyzed using computerized SSPS (Statistic Program Sigmastat, Sataistical Soft- Ware, SAS Institute , Cry, NC ). Effects of different treatments were analyzed by one way ANOVA (Analysis of variance ) test using Duncan’s multiple range test  and (Snedecor and Cohran , 1967).

9- Histopathological examination :

At end of the examination which continued for 28 successive days, all rats were sacrificed, organs including liver , kidney and heart were taken for histopathological examination according  to Drury and Wallington (1980).

RESULTS AND DISCUSSION

A-Effect of almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on body weight gain(BWG), feed intake(FI) & feed efficiency ratio(FER) of reinopathic rats.

1. Body weight gain(BWG)g/28 days;

Table(1) show the mean value of body weight gain (BWG/28days) of reinopathic rats fed on various diets. It could be noticed that the mean value of BWGg/28 days of control (+) group was lower than control (-) group, being 6.20±0.68 & 10.78±0.27 respectively, showing significant difference with percent of increase +73.87% as compared to control (+).All reinopathic rats fed on various diets showed significant  increases in mean values  as compared  to control (+) group . The best BWG was recorded for group 3 (reinopathic rats fed on almond seeds) when compared to control (+) group.

1. Feed intake (F.I):

Table(2) results indicates the mean value of feed intake (g/day) of reinopathic rats fed on variable diets. Data revealed that the mean value of( FI) of control (+) group was higher  than control (-) group,being15.18±0.75 & 12.28±0.63g. respectively. All  reinopathic rats fed on various diets showed significant  decreases in mean values  as compared  to control (+) group. Rats fed on groups 3,4,5,6&7 showed nonsignificant differences  between them. Numerically the best FI was recorded for group7 (reinopathic rats fed on mixture of all  plants) when compared to control (-) group.

1. Feed efficiency ratio(FER).

Table(3) results illustrate the mean value of FER of reinopathic rats fed on different diets. Data show that the mean value of FER of control (+) group was lower  than control (-) group,being0.015±0.002 &0.031±0.003 respectively,indicating significant difference. All reinopathic rats fed on various diets showed significant  increases in mean values  as compared  to control (+) group . Rats fed on groups 3,4,5,6&7 showed nonsignificant differences  between them . Numerically the best FER was recorded for group3&4 (reinopathic rats fed on almond nut,sambucus herbs).

The obtained results are in line with that of Sheir,Marwa(2009). Wien et al.,(2003);Kamil et al.,(2012)found that supplementation with Almond was associated with great reduction in weight. Kim et al.,(2008) suggested that Juniperus chinesis hot water extract reduse BWG in albino rats. Shen et al.,(2005)reported  that the scent of Grapefruit reduced appetite and body weight in rats.Fujioka et al.,(2006)suggested that  the fresh Grapefruit had effect on body weight in obese patient .Dow et al .,(2012)found that consuming Grapefruit may promote weight loss in overweight adult. Chrubasik et al.,(2008)reported that Sambucus nigra used for weight reduction.

Table (1): Effect of Almond  nut,Sambucus herbs,Juniperus seeds, Grapefruit albedo&the combination of all(as 5%) on body weight gain(BWG)par 28 days ,(FI)par day and(FER) of  reinopathic rats

Values denote arithmetic means ± standard deviation of the mean. (a, b ,c and d) in the same column differ significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are non significantly different.               

B-Effect of Almond nut,Sambucus herbs,Juniperus seeds,Grapefruit albedo&the combination of all on organs weight of reinopathic rats.

1-Liver weight (g):

Table(2) resalts show the mean value of liver weight (g) of reinopathic rats fed on various diets. It could be noticed that the mean value of liver( g.) of control (+) group was higher than control (-) group,being 5.6±0.19 & 3.98±0.19 respectively,showing significant difference.All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Numerically the best liver weight was recorded for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                   

2-Kidneys weight (g):

 Table(5) resalts indicate the mean value of kidneys weight (g) of reinopathic rats fed on different diets. It could be noticed that the mean value of kidneys( g.) of control (+) group was higher than control (-) group. All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Numerically the best kidneys weight was reveald for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                                                                                                                         

3-Spleen weight (g):

    Table(6) resalts show the mean value of spleen weight (g) of reinopathic rats fed on various diets. It could be observed that the mean value of spleen( g.) of control (+) group was higher than control (-) group. All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Rats fed on groups 7 showed nonsignificant differences  when compared to group 1 (healthy rats).Numerically the best kidneys weight was recorded for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                                                                                                      

4-Lungs weight (g):

Table(7) data show the mean value of lungs weight (g) of reinopathic rats fed on various diets. It could be revealed that the mean value of lungs (g.) of control (+) group was higher than control (-) group. All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Rats fed on groups 3,4,5,6&7 showed nonsignificant differences  between them. Numerically the better spleens weight was observed for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                                                               

5-Heart weight (g):

  Table(8) results show the mean value of heart weight (g) of reinopathic rats fed on different diets. It could be indicatled that the mean value of heart( g.) of control (+) group was higher than control (-) group,being 0.89±0.05 & 0.41±0.03 respectively. All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Rats fed on groups 4&5 showed nonsignificant differences  between them. It could be observed that rats fed on groups 3&7 showed nonsignificant difference as compared to control (-) group. Numerically the best heart weight was observed for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                                                                                                                                

Table(2):Effect of Almond  nut, Sambucus herbs, Juniperus seeds,    Grapefruit albedo & the combination of all (as5%) on organs weight (g)of reinopathic

Values denote arithmetic means ± standard deviation of the mean. (a, b ,c and d) in the same column differ significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are non significantly different.  

C-Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo&the combination of all on serum glucose(mg/dl) of reinopathic rats.

Table(3) data reveal the mean value of serum glucose (mgldl) of reinopathic rats fed on different diets. It could be noticed that the mean value of glucose of control (+) group was higher than control (-) group, being  155.00 ± 5.00 & 100.00 ± 4.34  respectively, showing significant difference . All reinopathic rats fed on different diets indicated significant decreases in mean values  as compared  to control (+) group . Numerically the better serum glucose was observed for group 7 (reinopathic rats fed on mixture of all plants) when compared to control (-) group.                                                                                                                                           

The obtained  results are in line with that of Ali,Amal (2011) found that nephritis inflicted rats by gentamicin revealed  increase of glucose in serum. Lovejoy  et al.,(2002)reported that Almond- enriched diets do not alter insulin sensitivity in health adults or glycemia in patients with diabetes. Jenkins et al.,(2006),(2008) however suggested that Almond may decrease the risk of oxidative damage to protein by decreasing the glycemic excrution. Wien et al.,(2010)found that the Almond- enriched intervention group exhibited greater reduction in insulin. Li et al.,(2011)reported that the Almond consumption is associated with amelioration in hyperglycemia. Kamil and chen(2012)concluded that Almond was usfuel for diabetes. Gray et al,.(2000)demonstrated that Sambucus nigra was a antidiabetic plant. Ciocoiu et al.,(2009)reported that S. nigra represents a possible dietary adjunct for the treatment of diabetes and a potential source for the discovery of new orally active agents for future diabetes therapy. Orhan et al.,(2011),(2012)reported that the treatment of diabetic rats with Juniperus oxycedrus decreased the blood glcose levels in liver and kidney tissues .Ju et al.,(2008) suggested that  the ethanolic extract of Chinese juniperus berries reduced the blood glucose levels of alloxan-induced diabetic  rats. Fujioka et al.,(2006)suggested that insulin resistance was improved with fresh Grapefruit of obese patient.                                      

Table(3):Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo &the combination of all(as5%) on serum glucose (mg/dl) of reinopathic rats

Values denote arithmetic means ± standard deviation of the mean. Means with different letters(a, b and c) in the same

column different significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are non significantly different

D-Effect of Almond  nut,Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on total cholesterole (T.C.), triglycerides (T.G), high density lipoprotein cholesterol (H.D.L.c) , Low density lipoprotein cholesterol of (L.D.Lc), very low density lipoproteins cholesterol (V.L.D.Lc) & atherogenic index(A.I.) of reinopathic rats .                     

1-Total cholesterole(T.C.)mg/dl :               

Data of table(4) illustrate the mean value of serum(T.C.) (mgldl) of reinopathic rats fed on different diets. It could be observed that the mean value of(T.C.) of control (+) group was higher than control (-) group,being  141.00 ±3.46 & 118.00 ±2.65  respectively,showing significant difference. All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group . Numerically the better serum (T.C) was showed for group 5 (reinopathic rats fed on Juniperus seeds) when compared to control (-) group.                                                                                                                                             

2-Triglycerides(T.G.)mgldl :

  Table(4) results show the mean value of serum(T.G.) (mgldl) of reinopathic rats fed on different diets. It could be noticed that the mean value of(T.G.) of control (+) group was higher than control (-) group, being  152.00 ±7.21 & 130.00 ±4.55  respectively, indicating significant difference . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group . Feeding on Juniperus group revealed nonsignificant different of(T.G.) as compared to control (-) group. Numerically the best serum (T.G.) was recorded for group 7 (reinopathic rats fed on mixture of plants) when compared to control (-) group.                                                                                                                             

3-High density lipoprotein cholesterol(H.D.Lc)mgldl:

Table(4) data indicate the mean value of serum (HDLc) (mgldl) of reinopathic rats fed on different diets. It could be observed that the mean value of (HDLc) of control (+) group was lower than control (-) group, being  74.30 ±5.14 & 88.30 ±2.86  respectively, showing significant difference . All reinopathic rats fed on different diets revealed significant increases in mean values  as compared  to control (+) group . Numerically the best serum (HDLc) was obseved for group 6& 7 (reinopathic rats fed on Grapefruit albedo &  mixture of plants) when compared to control (-) group.                                                                                                       

4-Low density lipoprotein cholesterol(L.D.Lc)mgldl:

Data in table(4) illustrat the mean value of serum(LDLc) (mgldl) of reinopathic rats fed on different diets. It could be observed that the mean value of(LDLc) of control (+) group was higher than control (-) group,being  36.30 ±4.07 &3.70 ±0.08 respectively,showing significant . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group . Rats fed on group 3( Almond nut) revealed nonosignificant differences as compared to control (-) group, and recorded the best serum (LDLc).                    

5- Very low density lipoprotein cholesterol(V.L.D.Lc)mgldl:

Data of table(4) indicate the mean value of serum (VLDLc) (mgldl) of reinopathic rats fed on different diets. It could be observed that the mean value of (VLDLc) of control (+) group was higher than control (-) group, being  30.40 ±3.98 &26.00 ±2.57 respectively, showing significant difference . All reinopathic rats fed on different diets showed significant decreases in mean values  as compared  to control (+) group . Rats fed on group . The best treatment  was recorded for group7 Atherogenic Index(A.I.)ratio.                                                                       

Data of table(4) revealed that the mean value of serum(A.I.) (mgldl) of reinopathic rats fed on different diets. It could be noticed that the mean value of(A.I.) of control (+) group was higher than control (-) group,being  0.892 ±0.09 & 0.34 ±0.01 respectively, showing significant difference . All reinopathic rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . Rats fed on group Numerically the best treatment  was recorded for group7 considering serum (A.I.).                                                                        

 The obtained  results are in line with that of Ali, Amal (2011)who found that nephritis inflicted rats by gentamicin caused increase of T.L.  , T.G.  , LDLc  , VLDLc & decrease of HDLc in serum. Wien et al., (2003) reported that supplementation with Almond was associated with great reduction in T.C., T. G., LDLc , VLDLc &increase in HDLc with type 2 diabetics. Li et al.,(2011) found that Almond consumption is associated with hypolipidemia. Rajaram et al ., (2010)showed that consumption of Almond nut lower blood lipids and risk of CHD. Spiller et al.,(1992),(1998)found that there was a rapid reduction in low density LDLc, TC without change in HDL  in humans when consumption  Almond nut. Jenkins et al.,(2002) found that the Almond decreased the level of blood lipids(LDLc) and (LDLc: HLDc) reduce coronary heart disease risk factor for the hiperlipidemic subjects. Lovejoy et al .,(2008) found that Almond-enrich diet had beneficial effects on serum lipids in healthy adults. Jaceldo et al .,(2011)  repoted that Almond had cholesterol lowering effect. Ciocoiu et al.,(2009)found that the polyphnols of Sambucus nigra reduce the lipid peroxides, neutralize  the lipidi peroxil radicals and inhibit the LDL oxidation. Murkovic et al., (2004) reported that Sambucus spray –dried extract at a low dose exerts a minor effect on serum lipids and antioxidative capacity higher ,but nutritionally relevant doses might significantly reduce  post prandial serum lipids. Ju et al.,(2008)suggested that the aqueous extract of Chinese juniperus had a potential hypolipidemic effect. Kim et al.,(2008) reported that Juniperus chinesis hot water extract reduce blood lipids levels of alino rats.Orhan et al.,(2011)found that the treatment of diabetic rats with Juniperus oxycedrus decrease both the levels of  lipid peroxidation in liver &kidney tissues. Rincon et al .,(2005) suggested that Grapfruits peels had the highest ascorbic  acid and carotenoid content ,good source of dietary fiber and phenolic compound whose use could be useful in the formulation of fuctional foods and suitable to reduce risk of some disease such as cardiovascular and some associated to lipid oxidation. Vinson et al.,(2002)reported that the juice of grapefruit was able to significantly inhibit atherosclerosis and lowerd cholesterol & triglycerids.                                                         

Table(4): Effect of Almond  nut, Sambucus herbs, Juniperusseeds, Grapefruit albedo & the combination of all on total cholesterole (T.C.), triglycerides(T.G), high density lipoprotein cholesterol (H.D.L.c) , Low density lipoprotein cholesterol of (L.D.Lc), very low density lipoproteins cholesterol (V.L.D.Lc) & atherogenic index(A.I.) of reinopathic rats

Values denote arithmetic means ± standard deviation of the mean. Means with different letters (a,b and c) in the same column different  significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are nonsignificantdifferent.

E-Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on kidney function of reinopathic rats.

1-Serum creatinine (mg/dl):

Data of table(5) indicated the mean value of serum creatinine (mg/dl) of reinopathic rats fed on various diets. It could be observed that the mean value of creatinine of control (+) group was higher than control (-) group, being  1.20 ±0.03 & 0.50 ±0.03respectively,showing significant difference . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group . Groups 3&7 indicated nonosignificant differences when compared to control (-) group and revealed the best treatment.

2-Serum urea (mg/dl):         

Data of table(5) illusterate the mean value of serum urea (mg/dl) of reinopathic rats fed on various diets. It could be noticed that the mean value of urea of control (+) group was higher than control (-) group,being  110.00 ±4.58 & 20.00 ±1.76 mg/dl  respectively,indicating significant difference . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group . The best treatment was recorded for group 7(mixture of all plants) when compared to control (+) group.

3-Serum uric acid(U. A.) (mg/dl):

Table(5) results show the mean value of serum(U. A) (mg/dl) of reinopathic rats fed on various diets. It could be observed that the mean value of uric acid of control (+) group was higher than control (-) group,being  7.70 ±0.03 & 1.70 ±0.03 mg/dl  respectively,indicating significant difference . All reinopathic rats fed on various diets revealed significant decreases in mean values  as compared  to control (+) group . The best treatment was recorded for group 7(mixture of all plants) when compared to control (+) group.                                                      

It seems possible that the used plants could correct the changes in kidneys function due to injection of rats with gentamicin .

Table(5): Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on kidney function of reinopathic rats

Values denote arithmetic means ± standard deviation of the mean. Means with different letters (a,b and c) in the same column different  significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are   nonsignificantdifferent.

4-Electrolytes (m.eq/l):

A-Serum sodium(Na) &potassium(k)(m.eq/l)

Table (6) results show the mean value of serum (Na)& (k)(m.eq/l) of reinopathic rats fed on various diets. It could be observed that the mean values of Na & k of control (+) group were lower than control (-) group,being(  36.20±2.45.03 & 199.00 ±7.94)&(2.80±0.26&12.40±0.35) m.eq/l  respectively,indicating significant differences . Rats fed on various diets revealed significant increases in mean values  as compared  to control (+) group . The best treatment was recorded for group 7(mixture of all plants) when compared to control (-) group.                                                      

It seems possible that the used plants could correct the changes in serum electrolytes Na & K due to injection of rats with gentamicin .                                                                                                           

B-Urine sodium(Na) &potassium(k)(m.eq/l)

Table (6) results indicate the mean value of urin (Na)& (k)(m.eq/l) of reinopathic rats fed on various diets. It could be noticed that the mean values of Na & k of control (+) group were higher than control (-) group, being(  245.00±15.09& 95.00 ±3.46)&(96.30±5.48 &44.10±2.7) m.eq/l  respectively, indicating significant differences . Rats fed on various diets showed significant decreases in mean values  as compared  to control (+) group . The best treatment was recorded for group 7(mixture of all plants)for both Na & k when compared to control (-) group.                              

It seems possible that the used plants could correct the changes in urine electrolytes  Na & K  due to injection of rats with gentamicin .                                                                                                          

Such results are in agreement with that recorded by EL – Moselhy , Shimaa (2007), Sheir , Marwa (2009)&Ali , Amal (2011),who reported that creatinine, urea& uric acid level was decreased by treatment with herbal combination of some plants.                                                                               

Anon (2012a),found that  Almond oil had diuretic effect in human body.                                            

Anon (2012b)  ,reported that the decoction of the inner bark  of Sambucus is used freely as a diuretic in the treatment of acute nephritis. Anon (2012c),found that Sambucus used for nephritis –edema with 12 -25g drug material from entire plant in decoction. Dzharullaeva et al .,(2009) found that the main biologically active substances are flavonoids (1.61%) in Juniperus oxycedrus and possess the diuretic effect. Tumen et al.,(2012) reported that Juniperus sp. is mainely used as diuretic  in Turkish folk.                                                                                                                                                         

Goldfarb et al., (2001) suggested that grapefruit juice ingestion was associated with an increase in mean oxalate excretion and citrate excretion. Trinchieri et al.,(2002) showed that grapefruit juice increased urinary excretion of citrate and calcium, magnesium in healthy subjects. Honow et al .,(2003)reported that both grapefruit and apple juices reduced the risk of calcium oxalate stone formation.

Table(6): Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all(as5%) on serum and urine sodium Na and k(m.eq/l) of reinopathic rats

Values denote arithmetic means ± standard deviation of the mean.  Means with different letters (a, b, c and d) in the same column different significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are nonsignificantdifferent.

F-Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on liver function of reinopathic rats.

1-Serum glutamic oxaloacetate transaminase (GOT) or (AST) enzyme (U/L):

Data of table(7) illustrate the mean value of serum (GOT) (U/L) of reinopathic rats fed on various diets. It could be noticed that the mean value of (GOT)of control (+) group was higher than control (-) group,being  60.00 ±5.00 & 16.00 ±2.51respectively,showing significant difference . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group. Mixture of all plants(group7) revealed the best treatment when compared to control (+) group considering (GOT) activity.

2-Serum glutamic pyruvate transaminase(GPT) or(ALT)enzyme (U/L):

Data of table(7) show the mean value of serum (GPT) (U/L) of reinopathic rats fed on different diets. It could be observed that the mean value of (GPT)of control (+) group was higher than control (-) group,being  11.20 ±1.05 & 4.00 ±0.61 respectively,showing significant difference . All reinopathic rats fed on various diets revealed significant decreases in mean values  as compared  to contro l (+) group. Group( 7) which was the best treatment  considering the GPT  activity showed nonosignificant difference,in comparison with group (1) of healthy rats.

3-Serum alkaline phosphatase(ALP) enzyme (U/L):

Data of table(7) illustrate the mean value of serum (ALP) (U/L) of reinopathic rats fed on various diets. It could be noticed that the mean value of (ALP)of control (+) group was higher than control (-) group,being  249.20 ±10.15 & 87.00 ±2.13 respectively,indicated significant difference . All reinopathic rats fed on various diets revealed significant decreases in mean values  as compared  to control (+) group.Numerically the mixture of all pants diet recorded the better treatment of serum ALP.                                                                                                                                                       

4-Serum (GOT)/(GPT)or (AST)/(ALT) ratio (U/L):

Data of table(7) indicate the mean value of serum(GOT)/ (GPT)ratio (U/L) of reinopathic rats fed on various diets. It could be noticed that the mean value of (GOT)/(GPT)ratio of control (+) group was higher than control (-) group, being  5.40 ±0.25 & 4.00 ±0.34 respectively, showing significant . All reinopathic rats fed on various diets revealed significant decreases in mean values  as compared  to control (+) group. The best treatment  considering the GOT/ GPT ratio was recorded for group “7” in comparison with group (2).                                                                                           

Table(7): Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo & the combination of all on liver function of reinopathic rats.

(U/L)* means unit per liter Values denote arithmetic means ± standard deviation of the mean. Means with different letters (a and b) in the same column differ significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are nonsignificantdifferent.

5-Serum and urine total protein(mg/dl):

a-Serum total protein(T.P.):

Data of table(8) show the mean value of serum(T.P.) (mg/dl) of reinopathic rats fed on various diets. It could be observed that the mean value of (T.P.) of control (+) group was lower than control (-) group, being  5.60 ±0.36 & 11.30 ±0.26 respectively, indicating significant . All reinopathic rats fed on different diets showed significant increases in mean values  as compared  to control (+) group. The best treatment  considering the serum T.P. in sehwan was recorded for group (7) in comparison with group (2) .                                                                                                           

b-Urine total protein(T.P.):

Data of table(8)  indicate the mean value of urine(T.P.) (mg/dl) of reinopathic rats fed on various diets. It could be noticed that the mean value of (T.P.) of control (+) group was higher than control (-) group,being  4.90 ±0.52 & 3.20 ±0.13 respectively, indicating significant difference . All reinopathic rats fed on different diets revealed significant decreases in mean values  as compared  to control (+) group. Numerically the best treatment  considering the urine T.P. was recorded for group (5).                                                                                                                                                           

These results seemed to agree with that found by  El- Moselhy , Shimaa (2007)and Ali, Amal (2011) rising plants and herbes. Jenkins et al .,(2006) suggested that Almond may decrease the risk of oxidative damage to protein by decreasing glycemic excursion and decreased risk CHD . Zhu et al.,(2008)reported that the extraction of S.chinensis by 75% alcohol showed very significant protective effects on mice acute hepatic injury induced by CCl4. Orhan et al.,(2012)reported that Juniperus berries decreased liver enzymes ,ALP of diabetic rats. Dakovic et al.,(1999) found that the liver protein enzyme content is significantly decreased after the multiple oral Grapefruit juice ingestion of male mice .                                                                                                                           

Table(8): Effect of Almond  nut, Sambucus herbs, Juniperus seeds, Grapefruit albedo &the combination of all (as5%) on serum protein(mg/dl) of reinopathic rats

Values denote arithmetic means ± standard deviation of the mean.  Means with different letters (a, b, c and d) in the same column different significantly at p ≤ 0.05 using one way ANOVA test, while those with similar letters are nonsignificantdifferent.

HiSTOPATHOLOGICAL RESULTS.

Kidneys

Microscopically, kidneys of rat from (control – group) revealed the noral histological structure of renal parenchyma (Photo.1). While mean, kidneys of rat from (control + group)showed vacuolations of epithelial lining renal tubules, presence of protein cast in the lumen of renal tubules (Photo..2), Atrophy of some glamerulars tufts (Photo..3) and peritubular inflammatory cells infiltration (Photo.4) .However, kidneys of rat from Almond  nut 5%showed no changes except slight congestion of capillary tufts (Photo..5). Moreover, kidneys of rat from Sambucus herbs 5% revealed vacuolations of epithelial lining some renal tubules (Photo.6). Most examined sections from Juniperus seeds 5%showed no histopathological changes (Photo..7,8), whereas, some sections from this group showed distension of bowmen's space (Photo..9) Some examined sections from Grapefruit albedo 5%and mixture of all plants5% revealed no histopatholigcal changes (Photo.. 11, 12).             

Liver

Microscopically, liver of rat from (control -group) revealed the normal histological structure of hepatic lobule (Photo.1). While mean, liver of rat from (control + group) showed      cytoplasmic vacuolization of hepatocytes (Photo. 2, 3). However, liver of rat from Almond  nut 5%    

revealed local hepatic necrosis associated with inflammatory cells infiltration (Photo. 4) and kupffer cells activation (Photo.5). No histopatholigcal changes were noticed in liver of rat from Sambucus herbs 5%  (Photo.6). Examined sections from Juniperus seeds 5% revealed kupffer cells activation and dilatation of hepatic sinusoids (Photo. 7, 8). However, liver of rat Grapefruit albedo 5% showed

dilatation and cougestion of hepatic sinusoids (Photo. 9,10) and slight vacuolations of hepatocytes (Fig.12). Moreover, liver of rat from mixture of all Plants5% revealed no changes except hydropic degeneration of some hepatocytes (Photo.11).