Exploring the protective effect of Ascorbic acid on Amoxicillin and Clavulanic acid-induced lipid peroxidation in goat liver homogenate

 

Bibhas Pandit1*, Trilochan Satapathy1, Pooja Tiwari2

1Columbia Institute of Pharmacy, Near Vidhan Sabha, Tekari, Raipur, Chhattisgarh-493111, India.

2SLT Institute of Pharmaceutical Sciences, Guru Ghasidas Vishwavidyalaya, Koni, Bilaspur, Chhattisgarh, India-495009.

*Corresponding Author E-mail: bibhas.pandit@gmail.com

 

ABSTRACT:

Background: Therapeutic effect of ascorbic acid on amoxicillin and clavulanic acid-induced lipid peroxidation was carried out in got liver homogenates to investigate the relationship between drug-induced toxicity and drug-induced lipid peroxidation. Methods: The level of malondialdehyde, reduced glutathione and nitric oxide were estimated in control, drug-treated, drug-antioxidant treated and the only antioxidant-treated group at two hours and six hours of incubation time. Results: The level of malondialdehyde in the drug-treated group was found to be increased whereas the level of both reduced glutathione and nitric oxide decreased when compared to control. In the drug-antioxidant treated group, the level of malondialdehyde was found to be reduced whereas the extent of reduced glutathione, nitric oxide increased when compared to the drug-treated group. Conclusion: The present study established the potential of amoxicillin and clavulanic acid to induce lipid peroxidation. On the other hand, ascorbic acid repressed lipid peroxidation to a substantial level. Lipid peroxidation initiation capability of amoxicillin and clavulanic acid might be a contributing factor for their toxicity.

 

KEYWORDS: Malondialdehyde; nitric oxide; reduced glutathione; lipid peroxidation; amoxicillin; clavulanic acid; ascorbic acid; antibiotics.

 

 


 

 

1.       INTRODUCTION:

A substance capable of producing therapeutic effects can yield undesired or adverse effects also. “Side effects” and “toxic effects” are the two dimensions of adverse effects. Toxicity is always dose-related, whereas side effect might not be dose-related [1]. Many mechanisms developed to illustrate drug-induced lipid peroxidation. Generation of peroxide free radicals during the therapy is one of them. Peroxide free radicals are oxygenated species generated due to the oxidation of membrane phospholipids [2-4]. As drugs are the potential candidate to induce lipid peroxidation, they might contribute to drug-induced toxicity as an effect of lipid peroxidation [5-7].

 

 

Conversely, antioxidant plays a very vital role to suppress lipid peroxidation by scavenging peroxide free radicals [8-9]. The present work was intended to evaluate the antiperoxidative potentials of ascorbic acid [10-11] on amoxicillin and clavulanic acid-induced toxicity. The central themes of this hypothesis discussed below.

 

The development of a new drug molecule is always a massive investment of money and time. Whereas there are many molecules already in the market are limited to clinical uses, as they produce moderate to severe toxicities. We all know that an increase in Toxic Dose50 (TD50) value of a drug can raise the value of its therapeutic index (TI). It means the drug will produce toxic effects in a higher dose than previous dose. But how the TD50 value increased? As most of the drugs passed the omnipresent membrane, they are very liable to cause any perceptible changes in the membrane phospholipid, leads to lipid peroxidation. As lipid peroxidation is one of the causes of toxicity induced by the drugs, an antioxidant can be used to improve the TI of the drug. In this study, amoxicillin, and clavulanic acid used to cause lipid peroxidation in goat liver homogenates, and ascorbic acid used as an antioxidant. When clavulanic acid is an irreversible β-lactamase inhibitor, amoxicillin is susceptible to many β-lactamases. Use of amoxicillin in co-administration of clavulanic acid enhances the activity of amoxicillin to many folds [12] along with a severe side effect, hepatitis or liver injury [13-17]. As reactive oxygen and nitrogen species (ROS & RNS) damages the liver tissue [18-19], use of an antioxidant like ascorbic acid will be beneficial to reduce the chances of liver injury [20-21].

 

Ascorbic acid is a monosaccharide found both in animals and plants. It works as a reducing agent [22]. Ascorbic acid was proved as an effective antioxidant to reduce the lipid peroxidation prompted by gentamycin, cisplatin, ceftazidime, flutamide, and tobramycin [23-28]. Estimation of MDA, GSH and NO was performed in goat liver homogenates. MDA is an oxidative by product of arachidonic acid. The level of MDA increases during lipid peroxidation [29-31]. GSH and NO give protection against lipid peroxidation. The decreased level of GSH and NO than usual indicated the commencement of lipid peroxidation [32-34].

 

2.    MATERIALS AND METHODS:

a.     Materials:

N-1-napthylethylenediamine dihydrochloride, thiobarbituric acid, 5,5’-Dithiobis-(2-nitrobenzoic acid); ascorbic acid, reduced glutathione; 1,1,3,3-tetraethoxypropane; trichloroacetic acid, sodium hydroxide, potassium dihydrogen phosphate and sodium nitrite were purchased from Loba Chemie Private Limited, India. Pure amoxicillin and clavulanic acid were purchased from Glenmark, India. Analytical grade reagents were used in this work.

 

b.    Methods:

 i.     Preparation of goat liver homogenates [26]:

Goat liver was obtained from Raipur Municipal Corporation (RMC) authorized slaughterhouse. It was selected due to its availability and similarities with the human liver [35]. The goat liver was harvested and cut into pieces with a sharp knife. Then the pieces were dipped into a germ-free conical flask previously filled with phosphate buffer (pH 7.4) solution. Before preparing the homogenates, the liver pieces were removed from the container and rinsed well with newly prepared phosphate buffer solution of pH 7.4. After that, the liver was ground into homogenates in the ratio of 1 g/mL of same phosphate buffer. The homogenate was then divided into four parts of equal volume and treated as mentioned below.

 

One of the portions was treated with 1 mL of distilled water and marked as a control group (C). The second portion of the homogenate was treated with the combination dose of amoxicillinand clavulanate potassium at 0.42 mg/g and 0.1042 mg/g of tissue homogenate respectively. This group was marked as drug-treated group (D). The third portion was treated with amoxicillin, clavulanate potassium, and ascorbic acid at a dose of 0.42 mg/g, 0.1042 mg/g and 0.166 mg/g of tissue homogenates respectively. It was kept as a drug- and antioxidant-treated group (DA). The fourth portion was incubated with ascorbic acid at a dose of 0.166 mg/g of homogenate and kept as an antioxidant-treated group (A). Then the samples were shaken for 6 h using an orbital shaker.

 

ii.       Estimation of MDA level from the tissue homogenate samples:

The extent of peroxidation of lipids in the samples was determined by measuring the level of MDA using thiobarbituric acid (TBA) method [36]. The extent of MDA in the samples was estimated at 3 h and 6 h of incubation. After each specified hour of incubation 2.50 mL of an incubated mixture was poured into the centrifuge tubes (5 centrifuge tubes for each set, i.e. for C, D, DA, and A) and 2.50 mL of 10 % w/v TCA solution was mixedwith each tube to precipitate protein. The samples were then centrifuged at 3000 r.p.m for 30 min, and the supernatant was separated through filtration. Then 2.50 mL of the filtrate was taken in a stoppered glass tube (5 stoppered glass tubes for each set, i.e. for C, D, DA, and A). 5.00mL of 0.002 M thiobarbituric acid (TBA) solution was added to each tubes. The volume was adjusted to 10.00 mL by the addition of distilled water. The tubes were then kept on a water bath and boiledtill a pink colour developed.The absorbance of solutions were measured and recorded at 530 nm against a solution prepared by 5.00 mL of TBA solution and 5.00 mL of distilled water. Shimadzu UV-1800 Double Beam Spectrophotometer was used to measure the absorbance. The concentration of MDA present in the test samples were calculated from the standard calibration curve.

 

1,1,3,3-Tetrahydroxy propane (TEP) was used to construct the standard calibration curve. A series of standard solution were prepared by mixing 5.00 mL of freshly prepared TBA solution with several aliquots of standard TEP solution in graduated tubes. The volume of the tubes was then made up to 10.00 mL with distilled water. The tubes were then boiled in a steam bath form about 30 minutes, cooled, and their absorbances were recorded at 530 nm. The blank solution was prepared by mixing 5.00 mL of TBA solution and 5.00 mL of distilled water. Obsersed absorbances were plotted against concentrations to obtain the best-fit equation i.e. y=0.0069x, where y=absorbance, x=concentration. The value of R2 was found to be 0.9947 &standard error mean (sem)=0.015.

 

  iii.   Estimation of NO level from the tissue homogenate samples:

Estimation of NO was done as mentioned in Griess’s method [37-38]. The level of NO in the samples was determined at 3 h and 6 h of incubation. After each specified hour of incubation 2.50 mL of an incubated mixture was poured into the centrifuge tubes (5 centrifuge tubes for each set, i.e. for C, D, DA, and A) and 2.50 mL of 10 % w/v TCA solution was added to each tube for the precipitation of protein. The samples were then centrifuged at 3000 r.p.m for 30 min, and the supernatant was separated through filtration.5.00 mL of the filtrate was taken in stopper glass tube (5 stoppered glass tubes for each set, i.e. for C, D, DA, and A)and 0.50 mL of Griess reagent was added into it. Griess reagent was prepared by mixing 1:1 ratio of sulphanilamide (1 % w/v in 3N HCl) and 0.1 % w/v N-naphthyl ethylenediamine dihydrochloride. The absorbace of the solutions were measered at 540 nm, 10 minutes after the addtion of Griess reagent against a blank solution containing 5.00 mL of distilled water and 0.50 mL of Griess reagent. The concentrations of NO were calculated from the standard curve.

 

A standard curve was prepared by using Sodium nitrite solution.A series of standard solution were prepared by mixing 0.50 mL of freshly prepared Griess reagent with several aliquots of standard Sodium nitrate solution in graduated tubes. The volume of the tubes was then made up to 10.00 mL with phosphate buffer and the absorbance of the solutions was measured at 530 nm. The blank solution was prepared by mixing buffer and Griess reagent. Obsersed absorbances were plotted against concentrations to obtain the best-fit equation i.e. y=0.1096x, where y=absorbance, x=concentration. The value of R2=0.9977 & standard error mean (sem)=0.019.Shimadzu UV-1800 Double Beam Spectrophotometer was used to measure the absorbance.

 

  iv.   Estimation of GSH level from the tissue homogenate samples:

Estimation of GSH was done per Ellman’s methods [39]. The level of GSH in the samples was determined at 3 h and 6 h of incubation. After each specified hour of incubation 1.00 mL of an incubated mixture was transferred into the centrifuge tubes (5 centrifuge tubes for each set, i.e. for C, D, DA, and A) and one mL of 5 % w/v TCA solution in 1 mM EDTA was added to each tube. Then the mixture was centrifuged at 3000 r.p.m for 10 minutes and filtered to collect the supernatant. 1 mL of the filtrate was mixed with 5.00 mL of 0.10 M phosphate buffer (pH 8.0), and 0.40 mL of 5,5’-Dithiobis-(2-nitrobenzoic acid) (DTNB 0.01 % w/v in phosphate buffer) was added to it and the absorbance of the solutions was estimated at 412 nm.The blank solution was prepared from 6.00 mL of phosphate buffer and 0.40 mL of DTNB. The concentration of the test solutions was determined using standared curve, constructed as follows.

 

Different aliquots of the standard GSH solution were taken in 10.00 mL volumetric flasks. 0.40 mL of DTNB solution was added to each flask, and the volume was made upto the mark with freshly prepared phosphate buffer (pH 8.0) solution. The absorbance was recorded at 412 nm, against a blank. The blank solution was prepared by using 9.60 mL of phosphate buffer and 0.40 mL DTNB solution. Obsersed absorbances were plotted against concentrations to obtain the best-fit equation i.e. y=0.0005x, where y=absorbance, x=concentration. The value of R2=0.9959 & standard error mean (sem)=0.011.Shimadzu UV-1800 Double Beam Spectrophotometer was used to measure the absorbance.

 

3.    RESULTS:

Effects of ascorbic acid on amoxicillin and clavulanic acid-induced lipid peroxidation was reported as average percentage (%) change in MDA (Table I), NO (Table II) and GSH (Table III) concentration with respect to control group.  Interpretation of the results was supported by Student “t” test. Analysis of variance (ANOVA) was performed on the percentage changes data of samples D, DA and A with respect to control group at 2 h and 6 h of incubation period [40-41].


 

Table 1: Average % change in MDA concentration with respect to Control group.

After 2 h of incubation period

After 6 h of incubation period

Sets

D

DA

A

ANOVA

Sets

D

DA

A

ANOVA

1

33.83a

6.68a

-7.12a

F1=1.03

(df=6,12)

F2= 256.94

(df=2,12)

Critical difference:

(p = 0.05)#

Ranked mean**

(A)(B)(C)

1

28.37a

9.13a

-12.22a

F1=1.17

(df=6,12)

F2= 320.62

(df=2,12)

Critical difference:

(p = 0.05)#

Ranked mean**

(A)(B)(C)

2

21.85a

7.51a

-10.4a

2

34.25a

6.82a

-11.73a

3

24.09a

11.74a

-8.03a

3

38.85a

9.51a

-10.54a

4

27.31a

11.07a

-9.40a

4

33.20a

6.82a

-7.66a

5

27.45a

16.16a

-8.62a

5

34.36a

6.72a

-12.11a

6

24.09a

11.74a

-8.03a

6

32.61a

14.99a

-7.03a

7

27.31a

11.07a

-9.40a

7

28.37a

9.13a

-12.22a

Mean

±sem

26.57±0.84

10.86±0.64

-8.72±0.53

Mean±

sem

32.50±0.90

8.90±0.87

-10.00±0.91

*Percent changes of D, DA and A with respect to controls of corresponding hours are shown in the Table. sem= Standard Error of Means of four sets (n=7).  Significance of ‘t’ values of the changes of MDA content (df = 2) are shown as: a > 99%. #Critical values of F at p = 0.05 level, F1 = 2.99 [df = (6, 12)], F2 = 3.88 [df = (2, 12)] at p=0.05. F1 and F2 corresponding to variance ratio between groups and within groups, respectively. **Two means not included within the same parenthesis are statistically significantly different at p = 0.05 level.

 

Table 2: Average % change in NO concentration with respect to Control group.

After 2 h of incubation period

After 6 h of incubation period

Sets

D

DA

A

ANOVA

Sets

D

DA

A

ANOVA

1

-35.83a

-10.59a

32.26a

F1=1.20

(df=6,12)

F2= 677.7

(df=2,12)

Critical difference:

(p = 0.05)#

Ranked mean**

(A)(B)(C)

1

-32.28a

-9.54a

35.80a

F1=1.08

(df=6,12)

F2= 550.3

(df=2,12)

Critical difference

(p = 0.05)#

Ranked mean**

(A)(B)(C)

2

-33.92a

-11.76a

36.29a

2

-36.23a

-16.22a

34.56a

3

-28.48a

-14.08a

30.83a

3

-27.66a

-14.96a

25.24a

4

-33.51a

-9.14a

31.01a

4

-32.19a

-7.84a

33.54a

5

-32.91a

-14.00a

34.86a

5

-31.70a

-10.18a

35.02a

6

-37.24a

-8.90a

40.22a

6

-40.27a

-10.73a

31.25a

7

-36.67a

-15.11a

24.94a

7

-33.38a

-13.85a

26.64a

Mean±sem

-34.08±0.29

-11.94±0.72

32.92±1.18

Mean±sem

-33.39±0.43

-11.91±0.39

31.73±0.96

*Percent changes of D, DA and A with respect to controls of corresponding hours are shown in the Table. sem= Standard Error of Means of four sets (n=7).  Significance of ‘t’ values of the changes of NO content (df = 2) are shown as: a > 99%. #Critical values of F at p = 0.05 level, F1 = 2.99 [df = (6, 12)], F2 = 3.88 [df = (2, 12)] at p=0.05. F1 and F2 corresponding to variance ratio between groups and within groups, respectively. **Two means not included within the same parenthesis are statistically significantly different at p = 0.05 level.

 

Table 3: Average % change in GSH concentration with respect to Control group.

After 2 h of incubation period

After 6 h of incubation period

Sets

D

DA

A

ANOVA

Sets

D

DA

A

ANOVA

1

-23.06a

-8.41a

18.98a

F1=1.03

(df=6,12)

F2= 256.94

(df=2,12)

Critical difference:

(p = 0.05)#

Ranked mean**

(A)(B)(C)

1

-24.84a

-8.35a

13.37a

F1=1.17

(df=6,12)

F2= 320.6

(df=2,12)

Critical difference:

(p = 0.05)#

Ranked mean**

(A)(B)(C)

2

-24.23 a

-8.28a

19.88a

2

-25.16a

-7.77a

15.96a

3

-22.68 a

-7.98a

14.51a

3

-23.59a

-9.27a

11.80a

4

-19.34 a

-12.00a

22.53a

4

-24.43a

-6.32a

14.80a

5

-24.54 a

-10.43a

13.92a

5

-23.62a

-10.00a

11.07a

6

-21.13 a

-7.74a

19.46a

6

-23.00a

-12.53a

15.61a

7

-19.84 a

-11.75a

21.67a

7

-22.12a

-5.18a

20.73a

Mean±sem

-22.12±0.38

9.52±0.54

-18.72±0.49

Mean±sem

-23.83±0.32

-8.49±0.32

-14.77±0.33

* Percent changes of D, DA and A with respect to controls of corresponding hours are shown in the Table. sem= Standard Error of Means of four sets (n=7).  Significance of ‘t’ values of the changes of GSH content (df = 2) are shown as: a > 99%. #Critical values of F at p = 0.05 level, F1 = 2.99 [df = (6, 12)], F2 = 3.88 [df = (2, 12)] at p=0.05. F1 and F2 corresponding to variance ratio between groups and within groups, respectively. **Two means not included within the same parenthesis are statistically significantly different at p= 0.05 level.

 


 

Fig. 1: Average % change in MDA content after 2 h of incubation period

 

Fig. 2: Average % change in MDA content after 6 h of incubation period

 

 

Fig. 3: Average % change in NO content after 2 h of incubation period

 

Fig. 4: Average % change in NO content after 6 h of incubation period

 

Fig. 5: Average % change in GSH content after 2 h of incubation period

 

 

Fig. 6: Average % change in GSH content after 6 h of incubation period


 


4.       DISCUSSION:

Lipid peroxidation is a common phenomenon occurs invariably in almost all the cells of living organisms. It serves as an instance of cells involving in free radical reaction by generating reactive oxygen species (ROS). Drug-induced toxicity was linked to peroxidation effects. In this work change of MDA, NO, and GSH content in goat liver samples was measured to quantify the lipid peroxidation induction potency of chemotherapeutic agents. The change in MDA, NO, and GSH content were observed after the specified time intervals and the results were verified by statistical analysis methods. The percentage of change in MDA, NO, and GSH level of different samples were calculated with respect to the corresponding control. The level of MDA increased when the goat liver samples were treated with a drug shown in table-1. It suggested the drug has induced lipid peroxidation. But when the goat liver sample was treated with drug and antioxidant both, the MDA content decreased in comparison with the drug-treated group. It indicates that ascorbic acid can suppress Amoxicillin and Clavulanic induced lipid peroxidation. Being a good free radical scavenger, ascorbic acid may inhibit lipid peroxidation. When the goat liver sample was treated with only ascorbic acid then there was some increase in MDA content. Maybe it is due to the fact that ascorbic acid can reduce ferric to ferrous which promote the generation of hydroxyl radicals and other reactive oxygen species through Fenton’s reaction. NO is an important bio-regulatory molecule interferes in with many physiological and pathological processes within the mammalian body. NO is sometime show beneficial and sometimes shows detrimental effects. Sufficient production of NO from vascular endothelial cells is required to protect organ such as the liver from ischemic damage. Sustained levels of NO production result in tissue toxicity which leads to vascular collapse [42]. In table-2, the goat liver sample treated with the drug produced a decreased level of NO. But when treated with drug and antioxidant both the NO level increased. Again when the goat liver sample was treated with ascorbic acid alone the NO contents were little decreased in comparison to the control samples. This increase in NO content in both drug and antioxidant-treated group suggests the free radical scavenging property of ascorbic acid. The average present changes in reduced glutathione (GSH) level of seven samples were shown in table-3. It was observed that ascorbic acid increased the GSH level when compared to drug treated groups.  It has been observed that when the goat liver sample was treated with Amoxicillin and Clavulanic acid-treated treated group. Again when the goat liver sample was treated with Morin alone the GSH contents were little decreased in comparison to drug and drug + antioxidant treated group. Histograms of average percentage change in MDA, NO and GSH along with standard error mean were shown in Fig. 1, 2,3,4,5 and 6 respectively.

 

Multiple comparison analysis along with an analysis of variance was performed on the % changes data with respect to the control group of corresponding hours to compare the means of the samples. There were significant differences among various groups (F1) of D, DA and A, statistically different from each other whereas there were no significant differences among various groups (F2). Drug-induced lipid peroxidation may be considered as a possible mechanism of drug-induced toxicity. The potential of antioxidants in the reduction of drug-induced lipid peroxidation may be subjected to reduce drug induce toxicity. Antioxidants have been proven to be general cytoprotective agents of therapeutic benefit by many researchers. The possible role of antioxidants in reducing drug-induced lipid peroxidation provides a scope for consideration of antioxidant as a protective candidate in therapy for reducing drug-induced toxicity and consequence increase therapeutic index.

 

 

5.       CONCLUSIONS:

The lipid peroxidation induction capacity of amoxicillin and clavulanic acid might be a contributing factor for their liver toxicity. Ascorbic acid was found to be a good supressor of lipid peroxidation induced by amoxicillin administered with clavulanic acid. Co-administration of antioxidants along with antibiotics or chemotherapeutic agents might be implemented clinically with an aim of reducing drug-induced toxicity. Though no ultimate conclusion could be drawn from these investigations it was understood that the using of antioxidant would be beneficial on combination therapy of amoxicillin and clavulanic acid.

 

6.       CONFLICTS OF INTEREST:

The authors declare that there is no conflict of interest.

 

7.       REFERENCES:

1.        Edwards IR, Aronson JK. Adverse drug reactions: definitions, diagnosis, and management. London: Lancet. 2000. pp. 1255-1259.

2.        Cheeseman KH. Mechanisms and effects of lipid peroxidation. Molecular Aspects of Medicine. 1993; 14(3): 191-197.

3.        Girotti AW. Mechanisms of lipid peroxidation. Journal of Free Radicals Biology Med. 1985; 1(2): 87-95.

4.        Gurudharshini N, Madhumitha M, Muthusaravanan S, Perianayaki P, Poornimmashree A, Kumaravel K. A big picture on antimicrobial strategies then and now. Research Journal of Engineering and Technology. 2017; 8(4): 361-364.

5.        Kasapovic J, Pejic S, Stojiljkovic V, Todorovic A, Radosević-Jelic L, Saicic ZS. Antioxidant status and lipid peroxidation in the blood of breast cancer patients of different ages after chemotherapy with 5-fluorouracil, doxorubicin and cyclophosphamide. Clinical Biochemistry. 2010; 43(16-17): 1287-1293.

6.        Selvakumar K, Madhan R, Srinivasan G, Baskar V. Antioxidant assays in pharmacological research. Asian Journal of  Pharmacy and Technology. 2011; 1(4): 99-103.

7.        Karunakar Hegde, Cijo Issac, Arun B. Joshi. Inhibitory Response of Carissa carandas root extract on lipid peroxidation. Research Journal of Pharmacy and Technology.  2010; 3 (4): 1072-1076.

8.        Niki E, Yoshida Y, Saito Y, Noguchi N. Lipid peroxidation: Mechanisms, inhibition, and biological effects. Biochemical and Biophysicxal Research Communication. 2005; 338(1): 668-676.

9.        Jiby E, Rajesh MG, Anish NP, Deepa P, Jayan N. In vitro antioxidant activity of the methanolic extract of Simaruba glauca DC. Asian Journal of Research in Chemistry. 2010; 3(2): 312-315.

10.      Dima Al Diab, Nour Al Asaad. Comparative analysis of ascorbic acid content and antioxidant activity of some fruit juices in Syria. Research Journal of Pharmacy and Technology.  2018; 11(2): 515-520.

11.      Shrada BK, Dhanraj M. Role of Vitamin C in body health. Research Journal of Pharmacy and Technology. 2018; 11(4): 1378-1380.

12.      Todd PA, Benfield P. Amoxicillin/clavulanic acid: an update of its antibacterial activity, pharmacokinetic properties and therapeutic use. Drugs. 1990; 39(2): 264-307.

13.      Gresser U. Amoxicillin-clavulanic acid therapy may be associated with severe side effects-review of the literature, European Journal of Medical Research. 2001; 6(4): 139-149.

14.      Fontana RJ, Shakil AO, Greenson JK, Boyd I, Lee WM. Acute liver failure due to amoxicillin and amoxicillin/clavulanate. Digestive Diseases and Sciences. 2005; 50(10): 1785-1790.

15.      Salvo F, Polimeni G, Moretti, Conforti A, Leone R, Leoni O. Adverse drug reactions related to amoxicillin alone and in association with clavulanic acid: Data from spontaneous reporting in Italy. Journal of Antimicrobial Chemotherapy. 2007; 60(1): 121-126.

16.      Cundiff J, Joe S. Amoxicillin-clavulanic acid-induced hepatitis. American Journal of Otolaryngol-Head and Neck Medicine and Surgery. 2007; 28(1): 8-30.

17.      Garcia Rodriguez LA, Stricker BH, Zimmerman HJ. Risk of acute liver injury associated with the combination of amoxicillin and clavulanic acid. Archives of Internal Medicine. 1996; 156(12): 1327-1332.

18.      Muriel P. Role of free radicals in liver diseases. Hepatology International.  2009; 3(4): 526-536.

19.      Zhu R, Wang Y, Zhang L, Guo Q. Oxidative stress and liver disease. Hepatology Research. 2012; 42(8): 741-749.

20.      Beattiea D, Sherlock S. Ascorbic acid deficiency in liver disease. Gut.1976; 17(8): 571-575.

21.      Karunakar H, Cijo I, Arun B. Joshi. Inhibitory response of Carissa carandas root extract on lipid peroxidation. Research Journal of Pharmacy and Technology. 2010; 3 (4): 1072-1076.

22.      Hamid AA, Aiyelaagbe OO, Usman LA, Ameen OM, Lawal A. Antioxidants: its medicinal and pharmacological applications. African Journal of Pure and Applied Chemistry. 2010; 4(8): 142-151.

23.      Devbhuti P, Saha A, Sengupta C. Gentamicin induced lipid peroxidation and its control with ascorbic acid, Acta Polania Pharmaceutical Drug Research. 2009; 66(4): 363-369.

24.      Ray S, Roy K, Sengupta C. Cisplatin-induced lipid peroxidation and its inhibition with ascorbic acid. Indian Journal of Pharmaceutical Sciences. 2006; 68(2): 199-204.

25.      Devbhuti P, Devbhuti, Saha A, Sengupta C. Effect of ascorbic acid on lipid peroxidation induced by ceftazidime. Journal of Pharmaceutical Sciences and Technology. 2011; 1(1): 51-53.

26.      Ray S. Evaluation of protective role of ascorbic acid on Flutamide-induced lipid peroxidation. International Journal of Pharm Tech Research. 2012; 4(1): 135-140.

27.      Aruna P, Shruti B, Archana A, Sameer H, Villasrao J. Tobramycin-induced lipid peroxidation and its control with ascorbic acid. Indo American Journal of Pharmaceutical Research. 2013;3(8): 6076-6082.

28.      Roy K., De AU, Sengupta C. Evaluation of glutathione and ascorbic acid as suppressors of drug-induced lipid peroxidation, Indian Journal of Experimental Biology. 2000; 38(6): 580-586.

29.      Gaweł S, Wardas M, Niedworok E, Wardas P. Malondialdehyde (MDA) as a lipid peroxidation marker. Wiadomości Lekarskie. 2004; 57(9-10): 453-455.

30.      Ayala A, Muoz MF, Argelles S. Lipid peroxidation: Production, metabolism, and signaling mechanisms of malondialdehyde and 4-hydroxy-2-nonenal. Oxidative Medicine and Cell Longevity. 2014; 2014: 1-31.

31.      Huda MK, Baydaa ST, Sura AS. Effect of the (ɣ-ray) and laser tadiation on the important antioxidant enzyme glutathione (GSH) level in serum. Research Journal of Pharmacy and Technology. 2017; 10(10): 3386-3390.

32.      Niedernhofer LJ, Daniels JS, Rouzer CA, Greene RE, Marnett LJ. Malondialdehyde, a product of lipid peroxidation, is mutagenic in human cells. The Journal of Biological Chemsitry. 2003; 278(33): 31426-31433.

33.      Ferreira ALA, Machado PEA, Matsubara LS. Lipid peroxidation, antioxidant enzymes and glutathione levels in human erythrocytes exposed to colloidal iron hydroxide in vitro. Brazilian Journal of Medicine and Biological Reearch. 1999; 32(6): 689-694.

34.      Hogg N, Kalyanaraman B. Nitric oxide and lipid peroxidation. Biochimica Et Biophysica Acta. 1999; 1411(2-3): 378-384.

35.      Hilditch TP. The chemical constituents of natural fats. Champel and Hall Ltd. London. 1956, 3rd ed. pp. 664.

36.      Ohkawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Analytical Biochemistry. 1979; 95(2): 351-358.

37.      Sun J, Zhang X, Broderick M, Fein H. Measurement of nitric oxide production in biological systems by using griess reaction assay. Sensors. 2003; 3(8): 276-284.

38.      Supratim Ray. Exploring the protective role of water extract of Spirulina platensis on flutamide-induced lipid peroxidation using 4-Hydroxy nonenal and nitric oxide as model markers. Research Journal of Pharmacy and Technology.  2011; 4(12): 1857-1860.

39.      Ellman GL. Tissue sulfhydryl groups. Archives of Biochemistry and Biophysics. 1959; 82(1): 70-77.

40.      Ray S. Exploring the protective role of ascorbic acid on Busulfan-induced lipid peroxidation. Research Journal of Pharmaceutical, Biological and Chemical Sciences. 2011; 2(4): 702-708.

41.      Joseph PR, Paul B. Remington: The science and practice of pharmacy. In: Statistics. Lippincott Williams & Wilkins, Philadelphia 2006. 21st ed.  pp. 127-150.

42.      Balakrishnan N, Panda A B, Raj N R, Shrivastava A, Prathani R. The Evaluation of Nitric Oxide Scavenging Activity of Acalypha Indica Linn Root. Asian Journal of Research in Chemistry. 2009; 2(2): 148-150.

 

 

 

 

 

Received on 29.12.2018           Modified on 17.01.2019

Accepted on 20.02.2019         © RJPT All right reserved

Research J. Pharm. and Tech. 2019, 12(7):3301-3306.

DOI: 10.5958/0974-360X.2019.00556.0