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Journal of Drug Delivery and Therapeutics

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Open Access Full Text Article                                                                        Research Article

Antimicrobial Activity of Curcuma longa Rhizome Ethanolic Extract Mediated Sonicated Biogenic Bismuth- Silver Oxide Nanocomposites: Characterization and Isolation of Turmeric Oil from Rhizomes.

Adithya Conjeevaram Gopal ¹, M.F. Valan *¹

Loyola Institute of Frontier Energy, Loyola College (Autonomous), Chennai, Tamil Nadu, India.

INTRODUCTION

Curcuma longa which is known as turmeric, is a medicinal spice used in cooking and treatment of ailments since the time of antiquity. This herb is consisted of phenolics and volatile compounds including oils ¹. Natural product mediated synthesis of nanomaterials have gained increasing attraction due to their environmental compatibility and cost- effectiveness. Darabi et al have demonstrated the use of platinum based bimetallic nanocomposites ². Mina Sarani ³ et al have demonstrated the role of Ag- doped- bismuth oxide and Cu- doped- bismuth oxide nanocomposites synthesized using the aqueous extract of B. multifoda and have demonstrated their antibacterial activity. 

The aim of the study was in the synthesis of C. longa ethanolic extract mediated bismuth- silver oxide nanocomposite and their application in the antimicrobial study; isolation, characterization and optimization of a protocol for isolation of turmeric oil from the rhizomes of C. longa was to be studied as well.

MATERIALS AND METHODOLOGY

Extraction of Turmeric Ethanolic Extract:  

Five grams of viralimanjal variety of Curcuma longa powdered rhizomes (freshly processed) were dissolved in 75 mL of ethanol. The contents were filtered to remove the residue and the filtrate was heated for a shorter duration at 60 °C to obtain a concentrated brown oily liquid.

Preparation of Ethanolic Extract Mediated Bismuth- Silver Oxide Nanocomposites: 

1 mL of the ethanolic extract was dissolved in 99 mL of double distilled water.  To this solution, 0.1 M of silver nitrate solution was added and stirred well. Subsequently 0.1 M of bismuth nitrate pentahydrate solution was added to this mixture and stirred well. The contents were sonicated in a sonication apparatus at a temperature reaching upto 50 °C, the contents were decanted in a way that a pink precipitate settling at the bottom of the beaker was scrapped and isolated. The precipitate consisted of bismuth and silver oxide nanocomposites. 

Antimicrobial Assay: 

Antimicrobial assay of the nanocomposites was carried as against 8 bacteria. The disk diffusion method was employed to study the antimicrobial activity at 1 mg/disc concentration in singlet as per the protocol of Duraipandiyan and Ignacimuthu ⁴.

Phytochemical Characterization of Ethanolic Extract of Turmeric Rhizomes: 

Phytochemical characterization of the ethanolic extract was carried out qualitatively for determination of the presence of terpenoids, steroids, oil and volatile organic compounds as follows:

1. Test for terpenoids: To the ethanolic extract of powdered turmeric rhizomes, one 1mL of chloroform and 1 mL of concentrated sulphuric acid were added. A red precipitate was obtained which determined the presence of terpenoids.

2. Test for sterols: To the ethanolic extract of powdered turmeric rhizomes one 1 mL of acetic anhydride and 1 mL of concentrated sulphuric acid was added and a red precipatate was obtained which determined the presence of sterols.

3. Test for oils and volatile compounds: 1 ml of ethanolic extract of powdered turmeric rhizomes was taken and applied on to a filter paper. A stain was observed which indicated the presence of oils and volatile compounds.  

Sequential Extraction of Turmeric Oil and Characterization: 

0.15 g of freshly processed turmeric rhizome powder was dissolved in 75 mL ethanol, heated at 70 °C and filtered. The solid yellow residue was discarded post filtration and the brown oily filtrate was extracted with hexane and petroleum ether in the ratio of 30:90. The organic phase was thus isolated from rhizomes and further subjected to characterization using UV- Vis, FT- IR using Bruker Alpha II FT-IR Instrument and GC- MS analyses at Gesra Labs, Porur, Chennai, Tamil Nadu, India.

Optimization of Extraction of Turmeric Oil: 

One gram of powdered turmeric rhizomes was dissolved in 25 mL of ethanol and filtered. 25 mL of hexane was added to the filtrate and further filtered. The oily fraction was obtained and extracted further with the help of hexane and petroleum ether in the ratio of 20: 10.   The oil rich fraction which was obtained post optimization was dried using anhydrous sodium sulphate to remove water molecules and was thus dissolved in petroleum ether. 

HPTLC Fingerprinting: 

Fifteen microlitre of the chloroform extract of the oily sample was subjected to HPTLC (High performance- thin layer chromatography) Fingerprinting. The process was carried out by applying the oil sample on to the TLC plate with the help of the mobile phase consisting of hexane, toluene and formic acid in the ratio of 5.5:0.5:0.1. The plates were sprayed with vanillin- sulphuric acid reagent and thereby heated at 105 °C. The HPTLC Chromatogram was visualized using CAMAG TLC Visualizer at 254 nm under UV light and 520 nm under white light with the help of a tungsten lamp. The process was evaluated by Dr. R. Shakila and co- workers at Siddha Central Research Institute, Arumbakkam, Chennai, Tamil Nadu, India. VisionCATS software was used to analyze the HPTLC Fingerprinting images.

RESULTS AND DISCUSSION

Qualitative Phytochemical Test Analyses: The qualitative phytochemical test analyses revealed the presence of terpenoids, steroids, oils and volatile organic compounds.

Antimicrobial activity of the bimetallic nanocomposites: The antimicrobial activity was observed and the nanocomposites inhibited the growth of seven out of eight tested bacteria. The antimicrobial activity was not observed against the gram-negative bacterium Proteus sp. This is the first time we have used sonicated bismuth- silver oxide nanocomposites for the determination of antimicrobial activity. In this study, this activity was carried out in singlet and the antibacterial activity was considered to be moderate. The bacterial strains used were multi drug resistant (MDR) by nature. On comparison with the control, only Klebsiella pneumoniae exhibited comparatively higher susceptibility to the nanocomposites when compared to ciprofloxacin in terms of the zone of inhibition.

UV-Vis, FT-IR and GC-MS based characterization of turmeric oil: The UV-Vis, FT-IR and GC-MS based characterizations of the turmeric oil revealed the presence of aromatic compounds, fatty acids and volatile organic hydrocarbons. The ethanolic extract of C. longa has served as a reducing agent for bismuth- silver oxide nanocomposites. From the GC- MS Table, Dibutyl Phthalate showed the highest peak area. 1- octadecene also showed higher peak areas when compared to other components present in the turmeric oil. This indicated the presence of fatty acid esters, volatile organic compounds and terpenoids. Ar- turmerone which is a well- known terpenoid present in turmeric oil was however found to be present in smaller quantity, as shown in the peak area section of the GC- MS table presented above.

HPTLC Fingerprinting: From the HPTLC Fingerprinting Tables, it can be observed that after derivatization using vanilln- sulphuric acid reagent 8 peaks were obtained and the image of the chromatogram was recorded at 520 nm. The nature of the compounds were volatile in nature. The presence of volatile organic compounds and terpenoids was confirmed in the HPTLC Fingerprinting technique.

Figure 1: UV- Vis Spectroscopy of turmeric oil

Figure 2: FT-IR Spectroscopy of turmeric oil.

Figure 3: GC- MS chromatogram of turmeric oil.

Figure 4: HPTLC Fingerprint of TLC Plate under 254 nm.

Figure 5: HPTLC Fingerprint of TLC Plate under visible white light at 520 nm. This was derivatized using vanillin and sulphuric acid reagent.

Figure. 6: HPTLC fingerprinting of turmeric oil at 254 nm post derivatization.

Figure 7: HPTLC fingerprinting at 520 nm.

Table 1: Antimicrobial Activity of Bismuth- Silver Oxide Nanocomposites.

Table 2: GC- MS Profiling of Turmeric Oil

Table 3: FT-IR Table

CONCLUSION

Thus it could be inferred that the nanoparticles inhibited the growth of the microorganisms tested. Characterization of the turmeric ethanolic extract revealed the presence of terpenoids, sterols, oils, fatty acids and volatile organic compounds. This is the first time we have carried such a study and the turmeric oil is assumed to be the reducing agent for the formation of nanocomposites. The turmeric oil was isolated from the viralimanjal variety of Curcuma longa and the constituents were analyzed and compared with the standard NIST libraries.

Future Scope

The turmeric oil may be used in formulation of turmeric oil based antimicrobial agents for treatment of infectious diseases. The biogenic silver oxide and bismuth nanocomposites may be used in formulation of antimicrobial agents in combating multi drug resistant pathogens. Further studies in pre- clinical models shall be undertaken for determination of toxicity and safety potential. Cost shall be optimized from pilot scale to industrial scale production of Bismuth- silver oxide nanocomposite preparation. 

Future Direction: The characterization of the sonicated biogenic nanocomposites using UV- Vis, FT- IR, XRD, SEM- EDAX will be carried out in the near future. Also, the toxicity profiling of the biogenic sonicated nanocomposites will be carried out in pre-clinical models. The densitometric analysis using HPTLC will be carried out for estimation of terpenoids.

Acknowledgements: The authors would like to place in record of sincere gratitude to Dr. V. Duraipandiyan, Senior Scientist, Entomology Research Institute, Loyola College (Autonomous), Chennai, Tamil Nadu, India for evaluation of the antimicrobial assay. The authors would further thank the Jesuit Management, Entomology Research Institute and Loyola Institute of Frontier Energy, Loyola College (Autonomous) for provision of facilities. The Staff at Gesra Labs, Porur and SCRI, Arumbakkam, Chennai are sincerely acknowledged for their affirm in carrying out the characterization and HPTLC Studies respectively.

Conflict of Interest: The authors do not have any conflict of interests.

Author’s Contributions: Adithya C.G. prepared the bimetallic nanoparticles. Dr. M.F. Valan supervised the overall study.

REFERENCES

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