Docking Studies of Piperine-Vitamin a Conjugate to Study the Increase in Bioavailability of Vitamin A

 

Animisha Mokkapati¹, Radha Krishna Nagumantri¹, Chinna Babu Pydi¹, Ramakrishna Chintala², Satyanarayana Rentala^1*

¹Department of Biotechnology, Institute of Technology, GITAM University, Gandhi Nagar, Visakhapatnam 530 045 India

²GITAM Institute of Science, GITAM University, Gandhi Nagar, Visakhapatnam 530 045 India

 

ABSTRACT:

Piperine, a major constituent of Piper nigrum (Black pepper), is one of the well known components in many Ayurvedic formulations. Piperine is most studied bioenhancer because it inhibits drug metabolizing enzymes in rodents and increases plasma concentrations of several drugs, including P-glycoprotein substrates. However, there are no evidences on piperine-vitamin A conjugate to inhibit human CYP450 3A4. In this paper the influence of piperine- vitamin A conjugate to study the metabolism of vitamin A with CYP450 3A4 was investigated. In silico results showed in the present study when piperine was conjugated with vitamin A, CYP450 3A4 was inhibited. This method improved the binding of piperine-vitamin A conjugate with CYP450 3A4 and increased bioavailability of Vitamin A.

 

 

 

 

INTRODUCTION:

The prevalence of underweight children and vitamin a deficiency xerophthalmia blindness in India is among the highest in the world, and is nearly double that of Sub-Saharan Africa with dire consequences for mobility, mortality, productivity and economic growth [1]. Vitamin A and its derivatives (retinoids) are essential components in vision; they contribute to pattern formation during development and exert multiple effects on cell differentiation with important clinical implications.

 

It has been known for 50 years that the key step in the formation of vitamin A is the oxidative cleavage of beta-carotene; however, this enzymatic step has resisted molecular analysis. Piperine (1-piperoyl piperidine) which is the active compound in both Piper longum and Piper nigrum is mainly responsible for the bioenhancing activity [2]. The mechanisms for the bio-enhancer activity of piperine have been proposed including DNA receptor binding, modulation of cell signal transduction and inhibition of drug metabolism. Piperine has been demonstrated to increase the serum levels and lengthen the serum half lives of some nutritional substances, such as coenzyme Q10 and beta-carotene [3]. Major categories of drugs that have shown increased bioenhancement include cardiovascular, respiratory, CNS, GIT antibiotics and anticancer. Some examples include tetracyclines, sulfadiazine, vasicine rifampicin, INH, pyrazinamide, ethambutol, phenytoin, phenobarbitone, carbamazepine nimesulide, indomethacin beta-carotene coenzyme Q10 (CoQ10), ciprofloxacin curcumin, dapsone, amino acids, glucose and several other classes of drugs [4-6]. Like Piperine, curcumin could modulate P-glycoprotein and CYP3A4 expression, and in turn modify the pharmacokinetic profiles of P-glycoprotein and CYP3A4 substrates in male Sprague-Dawley rats [7, 8]. However, piperine inhibits both the drug transporter P-glycoprotein and the major drug-metabolizing enzyme CYP3A4. Because both proteins are expressed in enterocytes and hepatocytes and contribute to a major extent to first-pass elimination of many drugs, which indicate that dietary piperine could affect plasma concentrations of P-glycoprotein and CYP3A4 substrates in humans, in particular if these drugs are administered orally [9-11]. Very recently, it is shown that a single administration of 1g of black pepper more than doubled area under the plasma concentration time curve and elimination half-life of phenytoin [12-15]. In this paper we discussed the in silico docking studies of piperine conjugated with vitamin A (Retinol) into Cytochrome P450 3A4 (CYP450 3A4). This implies the efficacy of conjugate on vitamin A metabolism using cytochrome P450 red-ox system.

 

MATERIAL AND METHODS:

Tools Employed:

Protein Data Bank server (PDB:www.rcsb.org/pdb) [16], What If server (http://swift.cmbi.ru.nl/servers/ html/index.html) [17], ACD Chem S ketch and Mole Gro Virtual Docker and Viewer were used during the research work.

 

Preparation of CYP450 3A4 and Piperine-Vitamin A conjugate:

Cytochrome P450 3A4 structure was downloaded from PDB. Server. The ID generated was 1W0E. The protein was optimized using Whatif server. The optimized protein was used for further analysis. Piperine-Vitamin Aconjugate structure was constructed using ACD Chem Sketch 12.01 software. The three dimensional structure of the Piperine-Vitamin Aconjugate was optimized using ACDChemSketch – Tools- - 3D structure optimization wizard.

 

Docking studies of Piperine-Vitamin Aconjugate with 1W0E:

The protein was imported into MoleGro Virtual Docker version 4.0.2.0 and surface was created. Cavities were detected in the protein surface. Five cavities were found and they were represented in green color. Piperine-Vitamin Aconjugate was imported into MoleGro Virtual Docker software in .mol format. This ligand was docked into cavities and it produced five docking sites with different amino acid sequences. The MolDock score and RMSD values were calculated.

DISCUSSION:

Optimization of CYP450 3A4 and Piperine-Vitamin A conjugate

The optimized structure of CYP450 3A4 (PDB ID: 1W0E) was shown in Figure 1. The protein was subjected to detect cavities using MoleGro Virtual Docker version 4.0.2.0. The cavities were shown in green in (Figure 1). Five cavities were detected as shown in Black color in Figure 1.

 

Figure 1: 1W0E protein with five cavities

 

Piperine-Vitamin A conjugate was optimized using ACDChemSketch 12.01. The cavities and a pose of piperine-Vitamin A conjugate was shown in Figure 2.

 

 

Figure 2: Piperine – Vitamin A conjugate within five cavities

 

Docking in each cavity generated five poses (Pose 1 through 5). The five poses generated are shown in Figure 3. It should be noted that each pose has its own spatial arrangement.

 

Figure 3: Five poses were generated for piperine-vitamin A conjugate. The 3D optimized structures (using ACDChemsketch software) of all the five poses are shown

 

Figure 4: A). Pose 1 docked into the Cavity – residues and their positions are shown; B). Pose 2 docked into the Cavity – residues and their positions are shown; C). Pose 3 docked into the Cavity – residues and their positions are shown; D). Pose 4 docked into the Cavity – residues and their positions are shown.

 

Docking studies of Piperine- Vitamin A conjugate with CYP450 3A4

Docking of Piperine- Vitamin A conjugate with cavities of CYP450 3A4 generated five poses with unique chemical arrangement. The cavity structures were shown in Figure 4A, 4B, 4C and 4D.

 

The Mol Dock scores were given in Figure 5A. A Graph and RMSD values were generated during docking studies as shown in Figure 5B and 5C.

 

Figure 5: A). After docking the Ligand with 1W0E, lowest energy – that is lowest Mol Dock score poses were generated. B). The graph showing the process of docking C). RMSD matrix was generated during docking process

 

Docking results were shown in Table 1. The Mol Dock Score and cavity volume of Pose 1 was high; the structure in the pose 1 would be superior to other poses. This implies that the structure in pose 1, was firmly bound with CYP450 3A4, making the protein more inactive. This leads to the reduction or inhibition of metabolism of Piperine- Vitamin A conjugate. A similar report was also shown with crude piperine extract. To increase bioavailability of vitamin A, we prepared Piperine- Vitamin A conjugate and this ligand was docked with CYP450 3A4.

 

Table 1: Poses and residues around the surface cavity of 1W0E protein

Pose	Name	Residues at Proximity  - 1.50
1	Ligand 1	Arg 105; Met 452; Ile 118
2	Ligand 2	Arg 105, 440, 130;  Gly 444, 306; Ile118
3	Ligand 3	Ala 448; Arg 105;  Cys 442; Ile 369, 443, 118;  Ser119
4	Ligand 4	Ala 448; Arg 105, 372;  Gly 444; Phe302
5	Ligand 5	Arg 105, 375;  Cys 442;Ile 443

 

This report suggests that conjugating piperine with vitamin A may slow down the metabolism of vitamin A; thereby piperine probably enhances the bioavailability of vitamin A.

 

CONCLUSION:

Many drug-drug interactions can be /explained by inhibition of P-glycoprotein and/or CYP3A4. A broad variety of drugs are substrates for both P-glycoprotein and CYP3A4 and because many compounds are inhibitors of both proteins, elevated plasma concentrations of a drug by a concomitantly administered substance can be due to a dual effect on drug transport and metabolism. This was evidenced by our previous data on docking of piperine with CYP3A4, Ferritin and P- glycoprotein [12]. The reports presented in this paper clearly suggest that piperine probably influence the metabolism of vitamin A, a substrate for CYP3A4.

 

ACKNOWLEDGEMENT:

We thank University Grants Commission, Govt. of India, for sponsoring the communicated research work (F. No 42-221/2013 (SR)) and GITAM University for providing infrastructure to conduct the project.

 

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

 

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15.     PDB server: www.rcsb.org/pdb

16.     What if server: swift.cmbi.ru.nl/

 

 

 

 

 

Accepted on 05.06.2017          © RJPT All right reserved