STUDIES ON IN-VITRO ANTICANCER AND ANTIOXIDANT PROPERTIES FROM MARINE BACTERIAL PIGMENT ISOLATED FROM THE COASTAL AREA OF MARAKANAM (TN)HTML Full Text
STUDIES ON IN-VITRO ANTICANCER AND ANTIOXIDANT PROPERTIES FROM MARINE BACTERIAL PIGMENT ISOLATED FROM THE COASTAL AREA OF MARAKANAM (TN)
Department of Microbiology, School of Life Sciences, Pondicherry University, Puducherry- 605014, India.
ABSTRACT: Marine bacteria have the potentiality to produce diverse bioactive molecules such as pigment. Therefore, it needs to exploit and identifying a novel type of pigment from marine bacteria for Industrial applications. This study aimed to investigate the marine bacterial pigment against antioxidant and anti-cancer properties; the marine bacteria producing pigment were isolated from water samples collected at the coastal of Marakkanam (TN), India. The isolates were screened out based on the growth characteristics and performance of different media and the strain designated as MB4, which was taken as further studies. The strain MB4 characterized by SEM analysis showed that coccoid cell morphology, nonsporulating, Gram-positive with yellow pigmentation and positive for MR-VP, catalase, lipase, acetoin production, and hemolysis. The cells were able to tolerate 10 percent NaCl concentration and ability to grown pH 9. The MB4 strain was shown a higher wave-number (1395.77) cm-1 against Raman Intensity to identify pigment production. The methanolic extracted pigment was produced at a maximum peak at 260 nm. The yellow-pigmented crude extract checked for anti-cancer properties using the colon cancer cell line (HCT15), the cell viability has been reduced after treatment of the extract (25-500 µg ml-1) and also exhibits IC50 value of 255.58 ± 43.51 mg ml-1 antioxidant DPPH radical scavenging activity. Due to their yellow pigment productions which have antioxidant activity and anti-cancer properties, this could be a novel pigment-producing strain for biomedical and industrial applications.
MB4 strain, Yellow pigment, DPPH activity, Anti-cancer properties, Raman spectroscopy, Marine water
INTRODUCTION: Synthetic pigments made up of heavy metals and petroleum compounds are reported to be carcinogenic, allergic, induce hyperactivity, toxic, and organ damage, which are unsafe for both environment and human health 1. Due to their low cost, increased stability, and wide range of spectra, the synthetic pigments are widely used 2.
In recent years, natural pigments are highly demanding for the use of colouring agents in foods, fabrics, feed, printing ink, and cosmetics, which is nonpolluted, eco-friendly, and less cost.
Naturally occurring colourants are safe to use produced by microflora and fauna, which are nontoxic, noncarcinogenic and easily degradable 3. Generally, microbial pigments are the micro-organisms that produce colour, which is more attractive nowadays due to easy methods available for cultivation; pigments are highly stable and year-round availability 4. An alternative source for natural pigment produces from bacteria, fungi, and microalgae 5-9.
Bacteria have great potential to produce various bioproducts among the microorganisms, and one of the physiological characters produced by the bacteria is pigment production 10. Biomedical and pharmaceutical industries utilize marine natural products, and the bioactive compounds are produced by marine bacteria 11. Microorganisms are a promising source of natural pigments like carotenoids, flavins, chlorophyll, quinines, and prodigiosin are the pigments produced by coloured microorganisms found in the different environmental niche. In survey 12, yellow (31.3 %), orange (15.2%), brown (9.9%), and red or pink (5.4%) were found to be microorganisms recovered from marine sources. Various concentrations of minerals, a wide range of temperature, and it should be the ability to tolerate different pH is the good qualities of pigment producers 5. Marine organisms produce a variety of metabolites that are treated against antitumor, antioxidant, and antimicrobial activities.
In recent years, Raman spectroscopy is a popular analytical tool applied for the study of microorganisms, especially pigments and biomolecules, through qualitative and quantitative analysis. Strong Raman signals were exhibits when applied to the microbial pigments for the understanding of different pigments, even pure cultures as well as extracted pigments 13, 14. It has been used to monitor different types of pigments present in the microorganisms directly through pure cultures and environmental samples 15. Antioxidants may reduce the risk of diseases, particularly heart and cancer, in the health sector. It can absorb free radicals that may oxidize lipid or DNA, proteins, nucleic acids and cause degenerative disease 16. Many other findings reported that the β-carotene and other carotenoid extracted pigments have antioxidant activities 17. Antioxidant able to donate a hydrogen atom reduces DPPH as a result of colour loss which determines the scavenging capacity of a molecule 18. The carotenoid pigment producer Planococcus sp. TRC1 was showed appreciable antioxidant activity leading to pharmaceutical and food applications 19.
The activation, viability, and proliferation of the cells were measured quantitatively in calorimetric for MTT Assay 20. The dehydrogenase enzymes associated with the endoplasmatic reticulum and mitochondria in the living cell convert MTT into a purple-blue formation which is water-insoluble. The viable cell number is directly proportionate to product formation and inversely proportional to cytotoxicity 21. In this study, we are investigating the pigmented bacteria isolated from marine water samples to evaluate the antioxidant and anti-cancer properties of the extracted pigment.
MATERIALS AND METHODS:
Collection of MARINE WATER SAMPLE: Samples collected from the marine water surface at different sites along with the coastal areas of Marakanam (TN), India. The marine water samples from the sea surface were collected by using Teflon plates dipped into water, lifted horizontally, and scrapped off the adhering surface film till the procedure repeated the total volume of 30 to 50 ml water sample was collected. The samples were stored at 4°C until the isolation was carried out within 24 h 22. The salinity of the collected marine water samples was determined by using the protocol reported by 23.
Isolation of Yellow-Pigmented Bacterial Isolates from Marine Water: Isolation of pigment-producing colonies from collected seawater samples using selective Zobell marine agar medium 24. The plates were incubated at 37°C for 48 h, the colonies showing yellow, red, orange, and brown pigmentation subcultured for purification.
Screening, Morphological and Biochemical Characterisation of the Isolates: The bacterial isolate was plated on Zobell marine agar, Luria Bertani agar and Tryptic soy agar incubated at 37°C for three days. By using standard Microbiological techniques, the different cultural characteristics, cell morphological parameters, and Gram's reaction were studied to the bacterial cells.
Scanning Electron Microscopic (SEM) Analysis for MB4 Isolate: The yellow-pigmented bacterial culture isolate was grown on Luria Bertani broth were centrifuged, phosphate buffer saline (pH 7.0) used to wash for thrice to remove salts, glutaraldehyde (2%) solution was fixed with sample and allowed for alcoholic dehydration at 6-12 h. The dehydrated sample was prepared and analysed on SEM 25.
Biochemical Characterisation of the Pigmented Isolate: The biochemical characteristics of the yellow-pigmented bacterial MB4 isolate.
Kovac's Oxidase Test: By using a sterile toothpick, a well-isolated colony of MB4 was picked and thoroughly rubs into an area of the moist test disc impregnated with oxidase reagent. After 30 sec, the inoculated area was observed for colour change. A bluish-purple colour indicated a positive reaction 26.
Catalase Test: Cultures grown on MB4 slants for 24 to 48 h flooded with 0.5 ml of 3 per cent hydrogen peroxide. Rapid effervescence shows a positive result for catalase activity 27.
Methyl Red-Voges Proskauer Test (MR-VP test): Methyl Red-Voges Proskauer tests were used to differentiate acid producers from those producing a neutral product, acetoin. The isolate was inoculated in 5 ml MRVP tubes and incubated at 35 °C for 48 h. Methyl red positive tubes were observed by the change of the colour of the media from yellow to red. Voges-Proskauer test was recorded positive by the development of red colour due to the addition of Barritt's reagent-I and Barritt's reagent-II 28.
Lipase Activity: The isolate MB4 was inoculated on egg-yolk agar and incubated at 37°C for 48 h. The positive reaction of the lipase activity indicated that the development of opalescent precipitates 29.
Urease Test: It was performed on 5 ml urea broth in test tubes containing phenol red (pH 6.8) as the pH indicator. The urea broth tubes inoculated with isolates were incubated for 24 h. The positive tubes were developed on red colour 30.
Assay of Phosphatase Activity: The loopful of bacterial growth of strain was collected and deposited on the surface of the tangible medium containing the phosphatase substrate (para-nitrophenyl phosphate) to a final concentration of 1 mg ml-1. The plates were then incubated at 37°C as described by 31.
Coagulase and DNAse Production: It was detected by the method using ethylenediamine-tetraacetic acid (EDTA) treated coagulase plasma by the formation of a clot after 1, 2, 4 or 24 h recorded as positive. During the investigation, 0-1 % DNA (BDH) was added to this medium to enhance the detection of DNase 32.
Physiological Characterisation of the MB4 Isolate: The physiological characteristics such as pH and NaCl tolerance were determined. The effect of pH was determined by the preparation of nutrient broth with incremental pH values ranging from 4 to 14. Strain MB4 was inoculated and incubated for 48 h at 37°C, the growth in the culture broth read at 620 nm. The pigments were extracted and observed for maximum pigment production 33. The inoculation of MB4 also examined the effect of NaCl tolerance in nutrient broth in different concentrations ranging from 6 to 12% of NaCl. The flasks were incubated at 37°C for 48 h, and the results were observed for growth and pigment production.
Extraction of pigments from bacterial isolates: The yellow-pigmented MB4 isolate was grown in LB broth for seven days kept in a rotary shaker at 160 rpm at 37°C. The broth was centrifuged at 8,000 rpm for 15 min, and the cells were harvested and poured off the supernatant. The cell pellet was rewashed with sterile distilled water centrifuged. The pellets were resuspended with 5 ml methanol and sonicate the mixture until all visible pigments were extracted. The solvent mixtures were centrifuged at 4,000 rpm for 15 min; the pigment supernatant was separated and filtered through a Whatman no. 1 filter paper and analyzed by scanning the absorbance in the wavelength region from 300-700 nm using a UV-VIS spectrophotometer 34.
Raman Spectroscopy Analysis: The MB4 strain was grown on LB agar medium after 48 h of incubation and the pigmented pure culture to determine the strong Raman signalling for Raman spectroscopy 15.
Determination of DPPH Scavenging Assay: 35 was reported the method of determination of DPPH radical scavenging activity. An aliquot of 0.5 ml of the extracted pigment sample solution in methanol was mixed with 2.5 ml of 0.5 mM methanolic solution of DPPH. The sample mixture was shaken vigorously and incubated for 30 min in the dark at room temperature. The absorbance was measured at 517 nm using a UV spectrophotometer. Ascorbic acid was used as a positive control. DPPH free radical scavenging ability (%) was calculated by using the formula.
% of inhibition = absorbance of control – absorbance of sample/absorbance of control ×100
Cell Culture and MTT Assay: The MB4 pigmented extract was carried out with MTT assay. The HCT15 colon cancer cell line was plated separately using 96 well plates with the concentration of 1×104cells/well in DMEM media with 10% fetal bovine serum and 1X Antibiotic Antimycotic Solution in a CO2 incubator at 37°C with 5% CO2. The 200 μl of 1X PBS cells were washed, and then the cells were treated with various concentrations of crude extract of the pigmented compound in serum-free media and incubated for 24 h. At the end of the treatment period, the medium was aspirated from cells. Prepare 0.5 mg ml-1 MTT mixed in 1X PBS and incubated at 37°C for 4 h using CO2 incubator. After the incubation period, the medium containing MTT was discarded from the cells and washed using 200 μl of PBS. The formed crystals were dissolved with 100 μl of DMSO and thoroughly mixed. The formazan dye turns to purple-blue colour. The colour intensity was measured at 570 nm using a microplate reader 36.
Statistical Analysis: Statistical analysis was performed with Mean and Standard Deviation (SD) in excel, and all analyses were carried out in triplicates.
RESULTS AND DISCUSSION:
Isolation of Pigmented Bacteria from Marine Water: The bacteria producing pigment were isolated from the marine water sample by using Zobell Marine Agar media.
TABLE 1: SCREENING OF PIGMENT-PRODUCING BACTERIAL ISOLATES
|Location||Total no. of isolates||Pigment producing strains|
|Marakanam coastal region (12.1899° N, 79.9249° E)||MB1||MB2, MB4, MMB8
Ten isolated strains were purified and named viz., MB1, MB2, MB3, MB4, MMB5, MMB8, MBS7, MBS8, SB2 and SB10; the results were shown in Table 1. Based on the growth performance and pigment production, the strain MB4 was screened for further studies.
Morphological Characters of Bacterial Isolates: Pigment-producing bacteria are isolated from marine samples, which are ubiquitous 37. A result of Fig. 1, the isolated colony was round, smooth with yellow-pigmented, non-motile, non-sporulating, and gram-positive bacteria. 38 were reported that the carotenoid-producing microbes were isolated from the extreme environmental niche. The sixty marine species of yellow (19), orange (5), pink or salmon colour (5), brown (5) and red (1) were described by 39. The marine organism isolated in the present study was yellow-colored. All bacterial pigmented isolates need not be carotenogenic 40. Some pigmented bacteria from marine are Alteromonas (yellow, violet) 41-43, Flavobacterium (yellow) 44, Deleya, Marinomonas, Pseudomonas and Shewanella (yellow) 45, Erythrobacter (yellow) 46, Pseudoal-teromonas (purple, red, or yellow pigments), Xanthomonas (yellow) 47, Exiguobacterium (yellow, orange) 48, 34, and Bacteroidetes (yellow, orange, pink or red) 49 were reported to possess antagonistic activities. The SEM image of Fig. 2 was clearly shown that the isolate belongs to Cocci with irregularly arranged cells.
FIG. 1: GROWTH OF MB4 STRAIN ON LB AGAR MEDIUM
Biochemical Reaction of the Strain: The yellow-pigmented bacterium from deep-sea sediment was oxidase, DNase, methyl red, urease negative, and catalase, Voges–Proskauer positive exhibited by Croceicoccus marinus gen. nov., sp. nov., 50. The present investigation of the MB4 strain was shown in Table 2, which is positive for MR-VP, catalase, lipase, acetoin production, and hemolysis and negative for oxidase, coagulase, urease, and phosphatase.
TABLE 2: BIOCHEMICAL CHARACTERS OF THE PIGMENTED STRAIN
|S. no.||Biochemical reaction||MB4 strain|
Positive (+), Negative (-)
FIG. 2: SEM IMAGES OF PIGMENTED MARINE BACTERIAL MB4 STRAIN
Physiological Characterisation of the Isolates: Studies were reported the yellow-pigmented Polaribacter butkevichii sp. nov. were grown in pH between 7.6 and 8.2 isolated from marine water samples 51. The MB4 strain exhibits maximum growth at pH nine as compared to pH from 4 to 14 Fig. 3. The extreme halophiles were unable to grow in the presence of NaCl less than 12%, and it can also be able to grow in saturated NaCl, including halobacteria and Halocooci. The isolate MB4 was the ability to tolerate salt concentration of 10% NaCl which was shown in Fig. 4. It has been reported that the growth was exhibited up to 8.5% NaCl concentration by P. balearica strain 52. The KMM 1447T yellow-orange pigmented strain isolated from marine ascidian was the ability to grow in 8% NaCl 53.
FIG. 3: PIGMENTED MB4 STRAIN WAS GROWN AT DIFFERENT pH
FIG. 4: THE BACTERIAL PIGMENTED MB4 STRAIN WAS GROWN AT DIFFERENT NACL CONCENTRATION
Extraction of Yellow Pigment from the Strain MB4: The isolated autotrophic cell cultures were extracted yellow coloured pigments using methanol solvent results much efficient in the quantification was better than acetone 54. The yellow pigments were separated from cell pellets by methanol extraction from the bacterial broth Fig. 5. Based on the absorption spectrum, the characterisation of crude extracted yellow pigment exhibited a maximum at 260 nm Fig. 6. As compared to water, PBS or acetone extraction, pigments recovered even better by using methanol extraction 34, 55, 56. The lemon yellow colour pigment is extracted by methanol from Halomonas aquamarina MB598, also reported by 2.
FIG. 5: PELLETS ARE SEPARATED FROM THE PIGMENTED MB4 STRAIN
FIG. 6: METHANOLIC EXTRACTION OF YELLOW-PIGMENTED BACTERIAL ISOLATES
Raman Spectroscopy Analysis for Pigmented Strain: Raman spectroscopy, one of the highly sophisticated instruments applied in Microbiology research in recent years for the characterisation of microbial pigments. The pure cultured MB4 strain was analyzed by using Raman spectroscopy for pigment production, and the strain was showed that maximum wavenumber (1395.77) cm-1 against Raman intensity Fig. 7. The excitation of electronic absorption of the spectrum which produces a specific enhancement of certain Raman intensity (bands) correspondingly the portion of the molecule electronic transition occurs indicates moving of atoms in the chromophore when vibration takes place 57, 58.
FIG. 7: RAMAN SPECTROSCOPY ANALYSIS OF PIGMENTED MB4 BACTERIAL ISOLATE
Antioxidant Activity of Pigment Extract: Natural products like pigments, frequently used to evaluate antioxidant activity, the ascorbic acid, glutathione, cysteine, and tocopherols, were used as reducing agents which are decolorizing involving in DPPH radicals 59.
The reducing activity of free radical owing to antioxidant depending on the reduction of one electron which exhibits scavenging of DPPH; as a result development of antioxidant properties due to reducing power 60. The anticarcinogenic and antioxidants properties are present in carotenoid pigments 61.
FIG. 8A: GROWTH PERFORMANCE OF A PIGMENTED EXTRACT OF MB4 AND ASCORBIC ACID
FIG. 8B: SCAVENGING ACTIVITY OF MB4 PIGMENTED EXTRACT AND ASCORBIC ACID
In-vitro, the potential of antioxidant activity of yellow-pigmented crude extract (YPCE) was scavenged due to free radicals in 500 µg ml-1 concentrations, and the percentage of inhibition for DPPH showed 68.30 percent Fig. 8A with IC50 255.58 ± 43.51 among different concentrations from 25 to 500 µg ml-1 Fig. 8B.
The carotenoid pigment is the biological compound available for human beings that have properties of ulcer prevention 62. The DPPH free radical scavenging activity was shown in different concentrations by using the methanol extract 59. The pigment production by halophilic bacteria and their relation to radical scavenging property was reported by 63. The yellow pigment was extracted from Kocuria flava SIF3 discovered to have DPPH radical scavenging activity with an IC50 value of 1.25 mg ml-1 in-vitro antioxidant assay 64.
Pigment Extract Evaluated for Anti-cancer Activity: Although advanced techniques developed in the treatment, prevention, and diagnosis of the disease. Still, one of the most serious human health problems in the world is cancer despite their understanding of its biology, which causes severe to mankind 65. Microbial pigments possess anti-cancer activity, and prodigiosin pigment produces cytotoxicity on U937 leukemia cells extracted from Pseudoalteromonas sp. 1020R 66. The present investigation was performed on a potential crude extract of yellow pigment against colon cancer cell line (HCT15), which showed that the different dose concentrations of the methanolic fraction of MB4 isolated from seawater could largely inhibit cell proliferation at a concentration of 500 μg ml-1 Fig. 9A and Fig. 9B and no cytotoxicity was detected against the standard untreated cell. It was found to be reported similar studies by 67.
FIG. 10: EFFECT OF MB4 STRAIN CRUDE EXTRACT ON COLON CANCER CELL LINE
Microbial pigments possess antimicrobial, anti-inflammatory, anti-cancer, and antioxidant activities, which also act as colouring for various industries like food processing and cosmetics 4. The extracted crude pigment MB4 was checked against the colon cancer cell line (HCT15) for cytotoxic activity Fig. 10.
Many studies had reported that the inhibition of cell cycle and apoptosis induced by microbial pigments 68, 69. It inferred that the crude pigment extract of marine MB4 shown significant antioxidant and potential anti-cancer activity against (HCT15) cell lines of colon cancer. The cytotoxic activity against cervical cancer cells (HeLa) and HepG2 were demonstrated in yellow pigment from Streptomyces griseoaurantiacus 70. The potential breast cancer cell lines and lung cancer cells have experimented with pigment carotenoid extracted from Kocuria sp. QWT-12 71. The pigment was found to exhibited inhibitory action against the growth of human cancer cell lines, which develops anti-cancer drugs from Salinicoccus sp. 72. The novel compound from yellowish pigment produced by Rhodococcus maris reduced the risk of breast cancer 73. The Natrialba sp. M6 produce carotenoid pigments that were effective against anti-cancer and antiviral activities 74.
CONCLUSION: Bacterial pigments for different therapeutics are an area of recent research interest. The present study attempts to validate the anti-cancer potential of bacterial pigment using colon cancer cell line (HCT15) cells and antioxidant properties. We used in-vitro cultured cells (HCT15) to determine the anti-cancer activity of extracted pigments from MB4 strain by using MTT assay for measuring the cell viability, and our results depict a dose-dependent by increasing concentration of the extract which decreases in cell viability. The DPPH radical scavenging activity revealed an increased concentration of the extract with an IC50 value of 255.58 ± 43.51 mg ml-1. The anti-cancer and antioxidant activity were induced by the extracted marine bacterial pigments. The presence study envisages the antioxidant and anti-cancer potential of bacterial pigments isolated from marine water, which can find applications in therapeutic treatments. The organisms were isolated from marine water, and yellow color pigment-producing bacteria were used for pigment extraction. Biochemical identification studies have confirmed the MB4 organism to be gram-positive, Cocci with a distinct yellow color, positive for MR-VP, catalase, lipase, acetoin production, and hemolysis which can tolerate 10 percent NaCl and it can also grow pH at 9.
ACKNOWLEDGEMENT: We are thankful to the Department of Microbiology, Pondicherry University, Puducherry, for providing the facilities to carry out this research work successfully.
DECLARATION OF INTEREST: The authors declare no conflict of interest exists.
- Usman H, Farouq A, Baki A, Abdulkadir N and Mustapha G: Production and characterization of orange pigment produced by Halophilic bacterium Salinococcus roseus isolated from Abattoir soil. Journal of Microbiological Experiment 2018; 6(6): 238-43.
- Fariq A, Yasmin A and Jamil M: Production, characterization and antimicrobial activities of biopigments by Aquisalibacillus elongatus MB592, Salinicoccus sesuvii MB597, and Halomonas aquamarina MB598 isolated from Khewra Salt Range, Pakistan. Extremophiles 2019; 23(4): 435-49.
- Cristea D and Vilarem G: Improving light fastness of natural dyes on cotton yarn. Dyes and pigments 2006; 70(3): 238-45.
- Venil CK and Lakshmanaperumalsamy P: An insightful overview on microbial pigment, prodigiosin. Electronic Journal of Biology 2009; 5(3): 49-61.
- Joshi VK, Attri D, Bala A and Bhushan S: Microbial pigments. Indian Journal of Biotechnology 2003; 2: 362-69.
- Choi SY, Yoon KH, Lee JI and Mitchell RJ: Violacein: properties and production of a versatile bacterial pigment. Biomedical Research International 2015.
- Rao N, Prabhu M, Xiao M and Li WJ: Fungal and bacterial pigments: secondary metabolites with wide applications. Frontiers in Microbiology 2017; 8: 1113.
- Pandey N, Rahul J, Anita P and Sushma T: Optimisation and characterisation of the orange pigment produced by a cold adapted strain of Penicillium (GBPI_P155) isolated from mountain ecosystem. Mycology 2018; 9(2): 81-92.
- Ramesh C, Vinithkumar NV and Kirubagaran R: Multifaceted applications of microbial pigments: current knowledge, challenges and future directions for public health implications. Microorganisms 2019; 7(7): 186.
- Margalith PZ: The carotenoid pigments. In Pigment Microbiology. Chapman and Hall London 1992; 32-76.
- Karbalaei-Heidari HR, Partovifar M and Memarpoor-Yazdi MJAJOMB: Evaluation of the Bioactive Potential of Secondary Metabolites Produced by a New Marine Micrococcus Species Isolated from the Persian Gulf 2020; 12: 61-65.
- Venil CK, Zakaria ZA and Ahmad WA: Bacterial pigments and their applications. Process Biochemistry 2013; 48(7): 1065-79.
- Huang WE, Li M, Jarvis RM, Goodacre R and Banwart SA: Shining light on the microbial world: the application of Raman microspectroscopy. In Advances in Applied Microbiology 2010; 70: 153-86.
- Lu X, Al-Qadiri HM, Lin M and Rasco BA: Application of mid-infrared and Raman spectroscopy to the study of bacteria. Food and Bioprocess Technology 2011; 4(6): 919-35.
- Jehlička J, Edwards HG and Oren A: Raman spectroscopy of microbial pigments. Applied Environmental Microbiology 2014; 80(11): 3286-95.
- Kaur P, Heggland I, Aschner M and Syversen T: Docosahexaenoic acid may act as a neuroprotector for methylmercury-induced neurotoxicity in primary neural cell cultures. Neurotoxicology 2008; 29(6): 978-87.
- Britton G: Functions of intact carotenoids. In carotenoids 2008; 189-212.
- Kurechi T, Kikugawa K and Kato T: Studies on the antioxidants. XIII. Hydrogen donating capability of antioxidants to 2, 2-diphenyl-1-picrylhydrazyl. Chemical and Pharmaceutical Bulletin 1980; 28(7): 2089-93.
- Majumdar S, Priyadarshinee R, Kumar A, Mandal T and Mandal DD: Exploring Planococcus TRC1, a bacterial isolate, for carotenoid pigment production and detoxification of paper mill effluent in immobilized fluidized bed reactor. Journal of Cleaner Production 2019; 211: 1389-1402.
- Mosmann T: Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. Journal of Immunological Methods 1983; 65(1-2): 55-63.
- Fotakis G and Timbrell JA: In-vitro cytotoxicity assays: comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride. Toxicology Letters 2006; 160(2): 171-77.
- Kjelleberg S, Stenström TA and Odham G: Comparative study of different hydrophobic devices for sampling lipid surface films and adherent microorganisms. Marine Biology 1979; 53(1): 21-25.
- Müller TJ: Determination of salinity. In K.K.K. Grassh off and M. Erhardt (eds.), Methods of seawater analysis; Wiley-VCH, Weinheim, Germany 1999.
- Button DK, Schut F, Quang P, Martin R and Robertson BR: Viability and isolation of marine bacteria by dilution culture: theory, procedures, and initial results. Applied Environmental Microbiology 1993; 59(3): 881-91.
- McDougall LA, Holzapfel WH, Schillinger U, Feely DE and Rupnow JH: Scanning electron microscopy of target cells and molecular weight determination of a bacteriocin produced by Lactococcus lactis International Journal of Food Microbiology 1994; 24(1-2): 295-308.
- Seeley W and Van demark J: Microbes in action: A laboratory manual of Microbiology. The W.H. Freeman and Company Inc, San Francisco 1981.
- Smibert RM and Krieg NR: General characterization. In: Methodology for General Bacteriology (ed.) P. Gerhardt, Academic Publisher New York 1981; 400-50.
- O'meara RA: A simple delicate and rapid method of detecting the formation of acetylmethylcarbinol by bacteria fermenting carbohydrate. The Journal of Pathology and Bacteriology 1931; 34(4): 401-06.
- Aneja KR: Biochemical activities of Microorganisms. Experiments in Microbiology, Plant Pathology, Tissue culture and Mushroom cultivation. New Age International (P) Ltd (2nd). New Delhi 1996; 190-217.
- Christensen WB: Urea decomposition as a means of differentiating Proteus and paracolon cultures from each other and from Salmonella and Shigella types. Journal of Bacteriology 1946; 52(4): 461.
- Baird-Parker AC: A classification of Micrococci and Staphylococci based on physiological and biochemical tests. Journal of General Microbiology 1963; 30: 409-27.
- Menzies RE: Comparison of coagulase, deoxyribonuclease (DNase), and heat-stable nuclease tests for identification of Staphylococcus aureus. Journal of Clinical Pathology 1977; 30(7): 606-08.
- Benjamin S, Pawar Y, Surekha, Dhanya P, Josh MKS and Pradeep S: Micrococcus luteus Strain BAA2. A Novel Isolate Produces Carotenoid Pigment. Electonic Journal of Biology 2016; 12(1): 83-89.
- Sasidharan P, Raja R, Karthik C, Ranandkumar S and Indra AP: Isolation and characterization of yellow pigment producing Exiguobacterium Journal of Biomolecular Technology 2013; 4(4): 632-35.
- Blois MS: Antioxidant determinations by the use of a stable free radical. Nature 1958; 181(4617): 1199-1200.
- Meerloo JV, Gertjan JL, Kaspers and Cloos J: Cell Sensitivity Assays: The MTT Assay. Cancer Cell Culture 2011; 731: 237-45.
- Franks A, Haywood P, Holmström C, Egan S, Kjelleberg S and Kumar N: Isolation and structure elucidation of a novel yellow pigment from the marine bacterium Pseudoalteromonas tunicata. Molecules 2005; 10(10): 1286-91.
- Arunkumar K, Ramasamy and Udayasuriyan V: Isolation and characterization of a yellow pigmented colony forming bacterium for carotenogenesis. Biotechnology 2006; 5(1): 79-82.
- ZoBell CE and Upham HC: A list of marine bacteria including descriptions of sixty new species. Bulletin Scripps Institute of Oceanography, University of California 1944; 5: 239-92.
- Moss M: Bacterial pigments. Microbiologist 2002; 3(4): 10-12.
- Gauthier MJ and Flatau GN: Antibacterial activity of marine violet-pigmented Alteromonas with special reference to the production of brominated compounds. Canadian Journal of Microbiology 1976; 22(11): 1612-19.
- Gauthier MJ: Alteromonas citrea, a new Gram-negative, yellow-pigmented species from seawater. International Journal of Systematic and Evolutionary Microbiology 1977; 27(4): 349-54.
- Mccarthy SA and Johnson RM. Effects of various agents on the pigment (violacein) and antibiotic production of Alteromonas luteoviolacea. Bullet. Japan Society of Science Fishery 1985; 51(7): 1115-21.
- Reichenbach H, Kohl W, Böttger-Vetter A and Achenbach H: Flexirubin-type pigments in Flavobacterium. Archives of Microbiology 1980; 126(3): 291-93.
- Akagawa-Matsushita M, Itoh T, Katayama Y, Kuraishi H and Yamasato K: Isoprenoid quinone composition of some marine Alteromonas, Marinomonas, Deleya, Pseudomonas and Shewanella Microbiology 1992: 138(11): 2275-81.
- Denner EB, Vybiral D, Koblízek M, Kämpfer P, Busse HJ and Velimirov B: Erythrobacter citreus nov., a yellow-pigmented bacterium that lacks bacteriochlorophyll a, isolated from the western Mediterranean Sea. International Journal of Systematic and Evolutionary Microbiology 2002; 52(5): 1655-61.
- Kirti K, Amita S, Priti S, Mukesh Kumar A and Jyoti S: Colorful world of microbes: carotenoids and their applications. Advances in Biology 2014; 837-91.
- Balraj J, Pannerselvam K and Jayaraman A: Isolation of pigmented marine bacteria Exiguobacterium from peninsular region of India and a study on biological activity of purified pigment. International Journal of Science Technology and Research 2014; 3: 375-84.
- Kirchman DL: The ecology of Cytophaga–Flavobacteria in aquatic environments. FEMS Microbiology Ecology 2002; 39(2): 91-100.
- Xu XW, Wu YH, Wang CS, Wang XG, Oren A and Wu M: Croceicoccus marinus nov., sp. nov., a yellow-pigmented bacterium from deep-sea sediment, and emended description of the family Erythrobacteraceae. International Journal of Systematic and Evolutionary Microbiology 2009; 59(9): 2247-53.
- Nedashkovskaya OI, Kim SB, Lysenko AM, Kalinovskaya NI, Mikhailov VV, Kim IS and Bae KS: Polaribacter butkevichii nov., a novel marine mesophilic bacterium of the family Flavobacteriaceae. Current Microbiology 2005; 51(6): 408-12.
- Bennasar A, Rossello-Mora R, Lalucat J and Moore ER: 16S rRNA Gene Sequence Analysis Relative to Genomovars of Pseudomonas stutzeri and Proposal of Pseudomonas balearica nov. International Journal of Systematic and Evolutionary Microbiology 1996; 46(1): 200-05.
- Lyudmila AR, Uchino M, Falsen E, Anatoly ML, Natalia VZ and Valery VM: Pseudomonas xanthomarina nov., a novel bacterium isolated from marine ascidian. Journal of General and Applied Microbiology 2005; 51: 65-71.
- Henriques M, Silva A and Rocha J: Extraction and quantification of pigments from a marine microalga: a simple and reproducible method. Communicating Current Research and Educational Topics and Trends in Applied Microbiology Formatex 2007; 2: 586-93.
- Sørensen L, Hantke A and Eriksen NT: Purification of the photosynthetic pigment C‐phycocyanin from heterotrophic Galdieria sulphuraria. Journal of the Science of Food and Agriculture 2013; 93(12): 2933-38.
- Godinho A and Bhosle S: Carotenes produced by alkaliphilic orange-pigmented strain of Microbacterium arborescens-AGSB isolated from coastal sand dunes. Indian Journal of Marine Science 2008; 37(3): 307-12.
- Vítek P, Jehlička J, Edwards HG and Osterrothová K: Identification of β-carotene in an evaporitic matrix-evaluation of Raman spectroscopic analysis for astrobiological research on Mars. Analytical and Bioanalytical Chemistry 2009; 93(8): 1967-75.
- Vítek P, Osterrothová K and Jehlička J: Beta-carotene—a possible biomarker in the Martian evaporitic environment: Raman micro-spectroscopic study. Planetary and Space Science 2009a; 57(4): 454-59.
- Nishino T, Shibahara-Sone H, Kikuchi-Hayakawa H and Ishikawa F: Transit of radical scavenging activity of milk products prepared by Maillard reaction and Lactobacillus casei strain Shirota fermentation through the hamster intestine. Journal of Dairy Science 2000; 83(5): 915-22.
- Tanaka M, Kuei CW, Nagashima Y and Taguchi T: Application of antioxidative maillrad reaction products from histidine and glucose to sardine products. Nippon Suisan Gakkaishi 1998; 54(1): 1409-14.
- Bendich: What have we learned about the Biological actions of Beta-Carotene? American Journal of Nutrition Science 2004; 32-2: 225-30.
- Bowen J, Soutar C, Serwata RD, Lagocki S, White DA, Davies SJ and Young AJ: Utilization of (3S, 3′ S)‐astaxanthin acyl esters in pigmentation of rainbow trout (Oncorhynchus mykiss) Aquaculture Nutrition 2002; 8(1): 59-68.
- Subramanian P and Gurunathan J: Differential production of pigments by halophilic bacteria under the effect of salt and evaluation of their antioxidant activity. Applied Biochemistry and Biotechnology 2020; 190: 391-409.
- Sahar AM, Ghada SI, Maha IA, Ahmed MA, Bataweel NM and Abu-Zaid M: Production and partial characterization of yellow pigment produced by Kocuria flava isolate and testing its antioxidant and antimicrobial activity. International Journal of Life Science and Pharmaceutical Research 2020; 10(2): 58-66.
- Hanahan D and Weinberg RA: The hallmarks of cancer. cell 2000; 100(1): 57-70.
- Wang Y, Nakajima A, Hosokawa K, Soliev AB, Osaka I, Arakawa R and Enomoto K: Cytotoxic prodigiosin family pigments from Pseudoalteromonas 1020R isolated from the Pacific coast of Japan. Bioscience, Biotechnology and Biochemistry 2012; 76(6): 1229-32.
- Yuvaraj N, Kanmani P, Satishkumar R, Paari KA, Pattukumar V and Arul V: Extraction, purification and partial characterization of Cladophora glomerata against multidrug resistant human pathogen Acinetobacter baumannii and fish pathogens. World Journal of Fish Marine Science 2011; 3(1): 51-57.
- Montaner B, Navarro S, Piqué M, Vilaseca M, Martinell M, Giralt E, Gil J and Pérez‐Tomás R: Prodigiosin from the supernatant of Serratia marcescens induces apoptosis in haematopoietic cancer cell lines. British Journal of Pharmacology 2000; 131(3): 585-93.
- Pandey R, Chander R and Sainis KB: Prodigiosins: A novel family of immunosuppressants with anti cancer activity. Indian Journal of Biochemistry and Biophysics 2007; 44: 295-302.
- Prashanthi K, Suryan S and Varalakshmi KN: In-vitro anticancer property of yellow pigment from Streptomyces griseoaurantiacus JUACT 01. Brazilian Archives of Biological Technology 2015; 58: 869-76.
- Rezaeeyan Z, Safarpour A and Amoozegar MA: High carotenoids production by a halotolerant bacterium, Kocuria strain QWT-12 and anticancer activity of its carotenoids. EXCLI Journal 2017; 16: 840-51.
- Srilekha V, Krishna G, Mahender P and Charya MAS: Investigation of in-vitro cytotoxic activity of pigment extracted from Salinococcus isolated from Nellore sea coast. Journal of Marine Medical Society 2018; 20: 31-33.
- Elsayed Y, Refaat J, Abdelmohsen UR and Fouad MA: The genus Rhodococcus as a source of novel bioactive substances: a review. Journal of Pharmacognosy and Phytochemistry 2017; 6: 83-92.
- Ghada EH, Marwa MA, Gehan MA, Hanan Ghozlan, Soraya AS, Nadia AS and Yasser RA: In-vitro dual (anticancer and antiviral) activity of the carotenoids produced by haloalkaliphilic archaeon Natrialba M6. Scientific Reports 2020; 10: 5986.
How to cite this article:
Aroumougame S: Studies on in-vitro anticancer and antioxidant properties from marine bacterial pigment isolated from the coastal area of Marakanam (TN). Int J Pharm Sci & Res 2021; 12(4): 2370-79. doi: 10.13040/IJPSR.0975-8232.12(4).2370-79.
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