solvent extraction and measurement of antioxidant activity
TRANSCRIPT
Chiang Mai J. Sci. 2019; 46(4) : 661-671http://epg.science.cmu.ac.th/ejournal/Contributed Paper
Solvent Extraction and Measurement of Antioxidant Activity and Total Phenolic Content from Capsicum chinense Jacq. Cv Habanero at Different Maturity Stages Víctor Manuel Moo-Huchin* [a], María de Lourdes Vargas y Vargas [a], Jorge Abraham Tamayo-Cortez [a], Daniel Arcángel López-Sauri [a], Enrique Sauri-Duch [a], Alejandro Ortiz-Fernández [b], César Abraham Can-Cauich [a] and David Betancur-Ancona [c][a] Tecnológico Nacional de México. Instituto Tecnológico de Mérida, km 5 Mérida-Progreso, C.P. 97118
Mérida, Yucatán, México.[b] Tecnológico Nacional de México. Instituto Tecnológico Superior de Calkiní, Av. Ah-Canul, C.P. 24900
Calkiní, Campeche, México.[c] Facultad de Ingeniería Química, Universidad Autónoma de Yucatán, Periférico Norte km 33.5, Tablaje
Catastral 13615, Colonia Chuburná de Hidalgo Inn, C.P. 97203, Mérida, Yucatán, México.*Author for correspondence; e-mail: [email protected]
Received: 20 July 2018Revised: 8 February 2019
Accepted: 18 February 2019
ABSTRACT Theaimof thepresentworkwastoinvestigatetheinfluenceof maturity(MS1:green,
MS2:greenchangingtoorangeandMS3:orange),theextractionprocedure(EP1,withpH=7.0±0.1andEP2,withpH=2.0±0.1)andtheliquidtosolidratio(from10to60mL/g)usedfortheextraction,onthecontentof totalphenoliccompounds(TPC)andontheantioxidantactivitymeasuredwith theassaysDPPH,ABTSandFRAP.TheaqueousextractsobtainedwithEP2presentedahighercontentof TPCandhigherantioxidantactivity,measuredwiththeABTS.Theliquidtosolidratioof 60mL/gandMS3permittedtheHabanerochilipeppertoobtainthehighestquantityof TPCandantioxidantactivity.TheTPCcontentwashighlycorrelated (P≤ 0.05)with the antioxidant activity,measuredwith the assaysDPPH (R=0.88),ABTS(R=0.81)andFRAP(R=0.91).TheresultsobtainedallowustosuggestthatthedifferenceinpHof theextractionsolventcouldexertthemostsignificantimpactonthereactionmechanismof theTPCwithDPPH,ABTSandFRAP.Moreover,theconsumptionof orangecoloredHabanerochilipepperisproposed,givenitshighercontentof TPCanditsantioxidantactivity.
Keywords:antioxidantactivity,Capsicum chinense, fruits,maturity
1. INTRODUCTIONFreeradicalsareproducedinnormaland
pathological cell metabolism, consideringthat oxidation is essential to most living
organisms for the production of energy tofuel biological processes. Oxygen-centeredfreeradicalsandotherreactiveoxygenspecies
Chiang Mai J. Sci. 2019; 46(4)662
(ROS)havebeenassociatedwiththeonsetof manydiseasesanddegenerativeprocessesinageing. The ROS readily attack and induceoxidative damage to various biomolecules,including proteins, lipids, lipoproteins andDNA[1].Thisoxidativedamageisacrucialetiological factor implicated in several chronic human diseases such as diabetes mellitus, cancer, atherosclerosis, arthritis,neurodegenerative diseases and also in the ageingprocess[2].Almostallorganismsarewellprotectedagainstfreeradicaldamagebyenzymes such as superoxide dismutase andcatalase,orcompoundssuchasascorbicacid,tocopherol and polyphenolic compoundsobtained from the consumption of fruit and vegetables[3].
Chili peppers have been extensivelyusedover theyearsasapreservativeandasa spice to add flavor to food preparations.ThegenusCapsicum,whichoriginatesfromthe tropical and humid zones of Central and Southern America, belongs to theSolanaceae family and includes peppers of importanteconomicvalue[4].Thesoutheastof Mexico,specificallytheStateof Yucatan,is the leading producer of Habanero chili(Capsicum chinense); which is a traditional cropof theMayanpeoplewithasignificanteconomic relevance for the areas in which it is cultivated. The C. chinensecv.Habanero isveryaromaticandisalsooneof thehottestintheworld[5].Thedemandforthefruitof theHabanerochiliproducedinYucatanhasincreased in the international market with its exportationtoCanada,Europe,JapanandtheUSA,whereitisusedasaspiceinfoodstuffsand also to obtain derivatives which are used inthetreatmentof neuropathies[6].
TheMexicanOfficialStandard (NOM-189-SCFI-2012) indicates that the nationaland international markets distinguish the Habanero chili produced in the YucatanPeninsula from those originating from other
productionareasbasedonitscharacteristics,including taste, aroma, pungency, color andshelf-life, all of which are a result of thespecial conditions of the region, such asclimate,soilandlocation[6].
Anumberof reportsareavailablewhichdescribe the influenceof thematurationof Capsicum annuum and Capsicum baccatum on the content of bioactive compounds and their antioxidantactivityusingdifferentextractionsolvents[7,8].However,itiswellknownthatthe content of bioactive compounds and the antioxidantactivityof plantmaterialsuchaschili are influencedbyadiversityof factorswhichincludeclimaticconditions,genotype,maturity, cultivation techniques, type andvolumeof solventtoextracttheantioxidantcompounds and the number of extractionstages[4].
In the literature, different solventcombinations have been used to extractantioxidants from plant materials, such asfruits, vegetables and other foodstuffs. Themost widely used solvents for extractingphenolic compounds are water, ethanol,methanol,acetone,andtheirwatermixtures,with or without acid [9]. The solubility of phenolicsisgovernedbythechemicalnatureof the plant sample, aswell as the polarityof thesolventsused.Thereforethetypeof extraction solvent, as well as the isolationprocedures, may have a significant impactontheextractionyieldof polyphenolsfromplantmaterial[9].
Given the importance of Capsicum chinense as a typical ingredient in the gastronomyof the Mexican Southeast, and taking intoconsideration its high commercial value,there ismuch interest inexploringsomeof the factors which could have an influenceon the extraction of antioxidants from theedible fraction of this fruit; moreover, noinformation is currently available regardingthe effect of three degrees of maturity
Chiang Mai J. Sci. 2019; 46(4) 663
(MS1,MS2andMS3)of theHabanerochilipepper on the content of TPC and on the antioxidant activity. The aim of this studywas to evaluate the effectof the extractionsystem composition, liquid–powder ratioand the degree of maturity on the contentof TPCandantioxidantactivityof Capsicum chinenseJacq.CvHabanero.
2. MATERIALS AND METHODS 2.1 Procurement of Habanero Chili Pepper
The fruits of C. chinense cv. HabanerousedinthisstudywereacquiredinSeptember
2017 in a localmarket of Merida,Yucatan,Mexico. The fruits were selected accordingto size and integrity, eliminating thosepresenting lesions or signs of insect attack. Taking into consideration the color, theywere then divided according to their degree of maturity (5 kg of fruit per degree of maturitywereused):green(MS1) (4.8°Brixand 0.034 g of citric acid /100 g of fresh weight), green changing to orange (MS2)(5.5°Brixand0.029gof citricacid/100gof freshweight),andorange(MS3)(9.5°Brixand 0.024 g of citric acid/100 g of fresh weight)[10](Figure1).
Figure 1.Degree of maturity of Capsicum chinense used in this work. Green (MS1); green changing to orange (MS2) and orange (MS3).
After washing the fruits with distilledwater (3 times), the edible part of thefruit (which includes the pericarp and the placenta) was separated, frozen at -80°Cand lyophilized. The lyophilized fruitswereground in a blender (Osterizer®) and the resulting powder was stored at -20°C inhermeticallysealedbagsuntilrequired.
2.2 Extract PreparationForthepurposeof evaluatingtheeffect
of thecompositionof theextractionsystemandof theratioof extractant(mL)topowder(g)of Habanerochilipepper,twoprocedureswereused:EP1(withpH=7.0±0.1);amixtureof methanol/water(50:50,v/v)followedby
an extraction with a mixture of acetone/water (70:30, v/v) and EP2; a mixture of methanol/water(50:50,v/v)acidifiedatapH=2followedbyanextractionwithamixtureof acetone/water(70:30,v/v).
For the EP1 extraction, 1.0 g of dry powder was mixed with a volume of methanol/water solvent (10, 20, 40 and60mL) for 1 h at 73°C (This temperaturewas selected in aprevious assay, resultsnotshown). After agitation, the mixture wascentrifuged for 10 min at 1500 rpm andthe supernatant was recovered. The residue wassubjected toasecondextractionwithavolumeof acetone/watersolvent(10,20,40and60mL),andfurtheragitationfor1hat
Chiang Mai J. Sci. 2019; 46(4)664
73°C, the supernatants of both extractionswere mixed together. Finally, the extractswerestoredat─20°Cuntiltheiranalysis.
The other procedure for antioxidantextraction,EP2,wascarriedoutinasimilarmannerexceptfortheadjustmentof pH=2withhydrochloricacid0.1Nforthedifferentvolumesof themethanol/watermixture.
2.3 TPC TPCof theextracts(previouslydiluted
in an appropriate solvent) was analyzedin accordance with the Folin-Ciocalteaucolorimetric method described by Moo-Huchin et al. [11]. Absorbancewas read at765 nmwith a UV–Vis spectrophotometer(PerkinElmerLambda11).Theconcentrationof total soluble phenolic compounds was calculated using a standard curve of aqueoussolutionsof gallicacid(1–10ppm)and expressed asmg gallic acid equivalents(GAE)/g dry weight (DW) of Habanerochili.
2.4 Antioxidant ActivityIn order to determine the antioxidant
activityof theextracts(previouslydilutedinanappropriatesolvent)bymeansof theassaysDPPH,ABTSandFRAP, themethodologypreviously published byMoo-Huchin et al.[11],Can-Cauich et al. [12] andBenzie and
Strain [13]was employed. For each assay, acalibration curve was prepared using ascorbic acidasastandardandresultsareexpressedasascorbicacidequivalentsing/100gof DW.
2.5 Statistical Analysis The data were analyzed by analysis of
variance (ANOVA) and the comparisonof means was performed by Tukey testusing Statgraphics Plus software, version2.1 (Manugistic, Inc., Rockville,MD,USA).Statistical differences were considered to be significant(P≤0.05).
3. RESULTS AND DISCUSSION3.1 ANOVA Results
According to the results, the threeindependentfactors(method,liquidtosolidratio and maturity) and their interactions significantly affected (P ≤ 0.05; Table 1)thecontentof TPCandantioxidantactivity(DPPH, ABTS and FRAP) in Capsicum chinenseJacq.Cv.Habanero.
3.2 Content of Total Phenolic Compounds (TPC)
Thisstudyreports the influenceof theextraction process, the liquid/powder ratiointheextractionandthedegreeof maturityof Capsicum chinense on the TPC content (Figure2).
Table 1.Analysisof variancefortheTPCcontentandantioxidantactivity(DPPH,ABTSandFRAP)of extractsof Capsicum chinense Jacq. CvHabanero.
FactorsTPC DPPH ABTS FRAP
F-Ratio P-value F-Ratio P-value F-Ratio P-value F-Ratio P-value
A:Method 211.42 0.0000* 65.30 0.0000* 54.10 0.0000* 67.33 0.0000*
B:Liquidtosolidratio 2011.36 0.0000* 486.26 0.0000* 1855.12 0.0000* 654.00 0.0000*
C:Maturity 2125.47 0.0000* 204.95 0.0000* 131.99 0.0000* 477.79 0.0000*
AB 1.50 0.2355 5.79 0.0030* 10.64 0.0001* 8.50 0.0003*
AC 13.90 0.0001* 2.02 0.1501 0.61 0.5507 4.19 0.0248*
BC 98.99 0.0000* 10.30 0.0000* 29.79 0.0000* 49.64 0.0000*
*Significantatthe0.05probabilitylevel.
Chiang Mai J. Sci. 2019; 46(4) 665
Based on the statistical analysis, theextraction procedure, EP2, significantlyincreased the TPC content of Capsicum chinense,incomparisonwiththeTPCcontentreportedforEP1(Figure2).Inthisrespect,it has been reported that the extraction of TPC improves when an organic solvent is used (ethanol,methanol, acetone)mixed inwater at different proportions. Moreover,the yield of TPC extraction has beenreported to increase with the addition of an acid to an aqueous solvent, so that theacid hydrolysis can produce the liberationof the largest amount of hydrolysables,whicharelinkedtotheplantfiber[14].Theaddition of acids also causes hydrolysis of phenolic compounds that can transform the glycosylated form of the phenoliccompoundsintoaglyconformswhichcanbedetected in the spectrophotometric analysisof phenoliccompounds [15].ThisprovidesapartialexplanationastowhytheprocedureEP2 (usinghydrochloric acid0.1N inorderfor the methanol-water mixture to reach apH=2) obtained the highest TPC content.In addition, it is important to take intoconsiderationthattheextractionof theTPCalso depends on their chemical structure,where each radical group of the TPC reacts differentlytothepHof theextractionsolvent.
The stability and reactivity of theTPC canalso be affected by their nature (electron-donating or electron-withdrawing) and the size(stericallyhindering)of thesubstituteinthearomaticring[16].
In another result, itwas found that, astheliquid/powderratioincreases(from10to60mL/g)there isalsoasignificant increaseintheextractionof phenoliccompoundsforallthedegreesof maturityof theHabanerochili (Figure 2). The highest TPC contentwas obtained with the liquid/solid ratio of 60mL/gandisattributedtothefactthatagreatervolumeof solventcanenterthecell,whereas more phenolic compounds could be retained in the solvent under the highest conditionsof liquid/powderratio[17].Thisresult concurswith those reportedbySoodandGupta[18]whofoundthatagreaterratioof liquidtosolidallowsagreaterquantityof solventtoprovokemoredissolutionof TPC,in pomegranate peel.
Regardingthematurityof theHabanerochili, the highest TPC content was foundforMS3, followedbyMS2andMS1;whichwould support the use of Habanero chilipeppers when they are completely orange(MS3)asaningredientinthegastronomyof theMexicansoutheast.However,Kappeletal.[8]andConfortietal.[7]foundquitethe
1
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Figure 2. TPC content in Habanero chili pepper with three degrees of maturity (MS1:green, MS2: 22
green changing to orange, MS3: orange), using the extraction procedure EP1 (A) and EP2 (B). 23
24
1
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Figure 2. TPC content in Habanero chili pepper with three degrees of maturity (MS1:green, MS2: 22
green changing to orange, MS3: orange), using the extraction procedure EP1 (A) and EP2 (B). 23
24
Figure 2. TPCcontentinHabanerochilipepperwiththreedegreesof maturity(MS1:green,MS2:greenchangingtoorange,MS3:orange),usingtheextractionprocedureEP1(A)andEP2(B).
Chiang Mai J. Sci. 2019; 46(4)666
opposite, reporting that green chili peppers(not mature) of other cultivars (Capsicum annum and Capsicum baccatum) showed a higher quantityof TPCincomparisonwithredchilipeppers (mature). The enzymes, such aspectinmethylesteraseandpolygalacturonase,are associated with the alteration of fruit cell walls during maturation and their action reduces the complexation of the phenoliccompoundswiththepolysaccharides,whichmay explain why the mature chili MS3obtained a higher TPC content when it was exposed to the solvent, in comparisonwithMS2andMS1[19].Furtherstudiesarerequired inorder to characterize theprofileof phenolic compounds and obtain a better understanding of the metabolic changes of these secondary metabolites during thematuration of Capsicum chinense, given thatHowardetal.[20]alsofoundthatthecontentof total phenolic compounds of various species of Capsicum was higher with the use of the assayFolin-Ciocalteu, in comparisonwiththequantificationbyHPLC; it is likelythat the additional detection of capsaicinoids wasreflected.
3.3 Antioxidant ActivityThe antioxidant activityof a foodstuff
istheexpressionof thedifferentantioxidantcomponents which use different action mechanismstodelayorinhibittheoxidativedeterioration caused by free radicals.Measurement of the antioxidant activity of vegetables isof great relevance,withwhichit is possible to determine the resistance of a product to oxidation and the antioxidantactivityproducedbythefruitintheorganismat the moment of consumption. The chemical diversity of the antioxidants of aplantmakesitextremelydifficulttocarryoutanindividualquantificationof theiractivityin vitro;therefore,theuseof differentmethodsarerequiredinordertomeasurethelevelof antioxidantactivity,directlyinplantextracts.Inthepresentstudy,theantioxidantactivityof aqueous extracts from Habanero chilipeppers (obtained through two extractionprocedures with different liquid to powderratiosandwiththreedegreesof fruitmaturity)wasevaluatedwiththeassaysDPPH,ABTSandFRAP(Figures3,4and5).
2
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
Figure 3. Antioxidant activity (measured with the assay DPPH) of Habanero chili pepper with 48
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 49
procedure EP1 (A) and EP2 (B). 50
51
2
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
Figure 3. Antioxidant activity (measured with the assay DPPH) of Habanero chili pepper with 48
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 49
procedure EP1 (A) and EP2 (B). 50
51
Figure 3. Antioxidantactivity(measuredwiththeassayDPPH)of Habanerochilipepperwiththreedegreesof maturity(MS1:green,MS2:greenchangingtoorange,MS3:orange),usingtheextractionprocedureEP1(A)andEP2(B).
Chiang Mai J. Sci. 2019; 46(4) 667
Theantioxidantactivity(measuredwiththeassaysDPPHandFRAP)of theaqueousextracts obtained with the EP1 procedure(pH= 7.0±0.1) was significantly higher (p≤ 0.05) in comparisonwith the antioxidantactivityreportedfortheprocedureEP2(pH=2).Phenoliccompoundsarerecognizedasacid groups and tend to be deprotonated in moderated basicmedia, forming the anion,nucleophilic phenoxide, which becomeshighly soluble inwater [21], this provides apartialexplanationforthehigherantioxidant
activitymeasuredwiththeassayDPPH,usingthe procedureEP1whichwas not acidified(Figure 3). Dawidowicz and Olszowy [22]postulated that a change in the concentration of hydrogen ion can give rise to changein the mechanism of the DPPH radical-scavenging process due to the phenolic compounds(HAT,hydrogenatomtransfer).The reduction in pH leads to the presenceof the transfer mechanism of electrons coupledtoprotons(PC-ET)[23].Thus, theproperties of the medium have the most
3
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
Figure 4. Antioxidant activity (measured with the assay ABTS) of Habanero chili pepper with 75
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 76
procedure EP1 (A) and EP2 (B). 77
78
79
3
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
Figure 4. Antioxidant activity (measured with the assay ABTS) of Habanero chili pepper with 75
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 76
procedure EP1 (A) and EP2 (B). 77
78
79
Figure 4. Antioxidantactivity(measuredwiththeassayABTS)of Habanerochilipepperwiththreedegreesof maturity(MS1:green,MS2:greenchangingtoorange,MS3:orange),usingtheextractionprocedureEP1(A)andEP2(B).
4
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
Figure 5. Antioxidant activity (measured with the assay FRAP) of Habanero chili pepper with 102
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 103
procedure EP1 (A) and EP2 (B). 104
105
106
107
4
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
Figure 5. Antioxidant activity (measured with the assay FRAP) of Habanero chili pepper with 102
three degrees of maturity (MS1:green, MS2: green changing to orange, MS3: orange), using the extraction 103
procedure EP1 (A) and EP2 (B). 104
105
106
107
Figure 5.Antioxidantactivity(measuredwiththeassayFRAP)of Habanerochilipepperwiththreedegreesof maturity(MS1:green,MS2:greenchangingtoorange,MS3:orange),usingtheextractionprocedureEP1(A)andEP2(B).
Chiang Mai J. Sci. 2019; 46(4)668
significantimpactonthereactionmechanismof phenolic compounds with the DPPHradicaland,inconsequence,ontheiractivity.
The values expressed using theFRAP assay represent the correspondingconcentration of electron donating antioxidantswiththereduction in theferriciron (Fe3+) to ferrous ion (Fe2+). However,the use of low pH can significantly inhibittransferof theelectronfromtheantioxidanttotheferricion[24].Thisfactexplainswhythe extraction procedure EP1 (with pH= 7.0±0.1) favored the higher antioxidantactivity (FRAP)of the fruit, in comparisonwith the procedure of extraction EP2(pH=2.0±0.1)(Figure5).
Moreover, when the ABTS assay wasused,theantioxidantactivityvalueof aqueousextracts obtained with the procedure EP2(withadjustmentof pH=2)wassignificantlyhigherthanthatof theextractionprocedureEP1 (Figure 4). The assay of antioxidantactivity with ABTS can be performed in awiderangeof pHvalues[25].However,thereaction mechanism of the radical ABTSwiththeantioxidantcanchangewiththepHof themedium;forexample,Lemańskaetal.[26] reported that the transfer of electronswasfacilitatedinconditionsof acidpHwiththe use of the assay ABTS, which couldexplain why a higher value of antioxidantactivitywasobtainedwithEP2.
Theantioxidant activity,measuredwiththe three assays,was found to be higher inorange Habanero chilis (MS3) and whenthe liquid to solid ratio of 60 mL/g wasused.If theliquidtosolidratioisincreased(> 60 mL/g) (under the conditionsstudied), probably no more solublephenolic compounds are extracted (sincethe maximum extraction is reached) andtherefore,thecontentof TPCanditsactivityantioxidant would be unchanged.However,futurestudiesarerequiredtoinvestigatetheoptimalconditionsforextractionof phenoliccompounds from Capsicum chinenseJacq.usingthe information reported in this research as a starting point.
The values of the antioxidant activityreported in this work are not comparable to that reported in the literature. This fact is explainedbecausedifferentauthorsapplydifferentextractionconditionsanddifferentsolvents for the extraction of phenoliccompounds of Capsicum chinense.Inaddition,the expression of the units reported forantioxidantactivityaredifferent.
3.4 CorrelationThe correlation coefficient between
the antioxidant activity (ABTS, DPPH andFRAP)andthecontentof TPCareshowninTable2.Antioxidantactivity,measuredwiththeassaysABTS,DPPHandFRAP,presented
Table 2.Pearson’sCorrelationbetweendifferentparametersof antioxidantactivityandtotalphenolic compounds.
Parameters TPC ABTS DPPH FRAP
TPC 1
ABTS 0.81* 1
DPPH 0.88 0.89 1
FRAP 0.91 0.85 0.97 1
*Allthevaluesaresignificant(P≤0.05).
Chiang Mai J. Sci. 2019; 46(4) 669
apositivecorrelation(R=0.81;R=0.88andR = 0.91, P ≤ 0.05, respectively, with thecontent of total phenolic compounds. These results suggest that the total phenolic compounds may be the most importantmetabolites contributing in the antioxidantactivity of Habanero chili. However,according to the results, the value of thecorrelation coefficient between the contentof totalphenolic compounds and the assayFRAP is higher than the correlation valueobtainedwiththeothertwoassays;thiscanbeexplainedbythefactthattheFRAPassaydoesnotreactwiththethiolcompounds(i.e.,whichcontain–SH)norwiththecarotenoids[27, 28], in contrast with the assays ABTSandDPPH, which can react with the thiolcompounds and carotenoids [29, 30]. It isalso important to take into consideration that,inthiswork,solventsmixedwithwaterwere used, including acetone, which is asolvent traditionally used for the extractionof carotenoids[30].
In another result, a highly positivecorrelationwasfoundbetweenthethreeassaymethods used to measure the antioxidantactivityof Habanerochili(DPPHvs.ABTS,R = 0.89; FRAP vs. ABTS, R = 0.85 andFRAP vs. DPPH, R = 0.97; P ≤ 0.05),whichindicatesthattheaqueousextractsof Habanerochilishowsimilaractivitiesamongthe assays. These results have also beenreportedbyCan-Cauichetal.[12]inastudyconducted on the peel of tropical fruits.
4. CONCLUSIONSFrom the two extraction procedures
evaluated,theresultsshowthattheextractionwhich includes aqueous methanol acidifiedto pH= 2, followed by an extractionwithaqueousacetone,significantlyincreasedboththeTPCcontentandtheantioxidantactivity,measuredwith theassayABTS, in fruitsof Capsicum chinense.At a higher liquid to solid
ratio(60mL/g)andwithacompletelyorangecolored Habanero chili (MS3), the highestquantity of TPC and antioxidant activitywas obtained. Based on the correlationcoefficients, it is possible to conclude thatthe TPC may be the compounds whichcontribute to the antioxidant activity of the Habanero chili and that their aqueousextracts show similar activities between thethreeassays.Theconsumptionof Habanerochili, as an ingredient in the gastronomy, isrecommendedwhen the fruit is completelyorange,givenitshighercontentof TPCandantioxidantactivity.Moreover,itissuggestedthat thepHof the extractionmediummayhave the most significant impact on thereactionmechanismof theTPCwithDPPH,ABTSandFRAPand that this aspectmustbeexploredindepthinfuturestudiesontheHabanerochilipepper.
ACKNOWLEDGMENTSSpecial thanks to Mrs. Sandra Rae
Anderson for the English-native correctionof the manuscript.
DISCLOSURE STATEMENTNo potential conflict of interest was
reportedbytheauthors.
REFERENCES[1] FarberJ.L.,Environ. Health Perspect.,1994;
102: 17-24.
[2] RaoA.L.,BharaniM.andPallaviV.,Adv. Pharmacol. Toxicol.,2006;1: 29-38.
[3] WahleK.W.,BrownI.,RotondoD.andHeysS.,Adv. Exp. Med. Biol.,2010;698: 36-51.
[4] Menichini F., Tundis R., BonesiM., Loizzo M.R., Conforti F., StattiG. and Menichini F., Food Chem.,2009; 114: 553-560. DOI 10.1016/j.foodchem.2008.09.086.
Chiang Mai J. Sci. 2019; 46(4)670
[5] Pino J., Sauri-Duch E. andMarbot R.,Food Chem., 2006; 94: 394-398. DOI10.1016/j.foodchem.2004.11.040.
[6] Islas-FloresI.,González-EstradaT.andVillanueva M. A., The Capsaicin: Just as Hot as Hell?; InPandalai S.G., ed.,Recent Research Developments in Biochemistry,Kerela,India,2005:121-132.
[7] ConfortiF.,StattiG.A.andMenichiniF.,Food Chem.,2007;102:1096-1104.DOI10.1016/j.foodchem.2006.06.047.
[8] Kappel V.D., Costa G.M., Scola G.,SilvaF.A.,LandellM.F.,ValenteP. andMoreiraJ.C.,J. Med. Food,2008;11: 267-274.DOI10.1089/jmf.2007.626.
[9] Azmir J.,Zaidul I.S.M.,RahmanM.M.,Sharif K.M., Mohamed A., Sahena F.,JahurulM.H.A.,GhafoorK.,NorulainiN.A.N. and Omar A.K.M., J. Food Eng.,2013;117:426-436.DOI10.1016/j.jfoodeng.2013.01.014.
[10]González M., Centurión A., Sauri E.and Latournerie L.,Acta Hortic., 2005;682: 1297-1302. DOI 10.17660/ActaHortic.2005.682.174.
[11]Moo-Huchin V.M., Estrada-Mota I.,Estrada-León R., Cuevas-Glory L.,Ortiz-Vázquez E., Vargas M.D.L.V.and Sauri-Duch E., Food Chem., 2014; 152: 508-515. DOI 10.1016/j.foodchem.2013.12.013.
[12]Can-Cauich C.A., Sauri-Duch E.,Betancur-AnconaD.,Chel-GuerreroL.,González-Aguilar G. A., Cuevas-GloryL. F. andMoo-Huchin V.M., J. Funct. Foods,2017;37:501-506.DOI10.1016/j.jff.2017.08.028.
[13]BenzieI.F.F.andStrainJ.J.,Anal. Biochem.,1996; 239: 70-76.
[14]Ruenroengklin N., Zhong J., Duan X.,Yang B., Li J. and Jiang Y., Int. J. Mol. Sci.,2008;9:1333-1341.DOI10.3390/ijms9071333.
[15]MelloB.C.andHubingerM.D.,Int. J. Food Sci. Technol., 2012; 47: 2510-2518.DOI10.1111/j.1365-2621.2012.03129.x.
[16]GanapathiM.R.,NaumovS.,HermannR.andBredeO.,Chem. Phys. Lett.,2001;337:335-340.
[17]PrasadN.K.,ChunY.,EnY.,ZhaoM.andJiangY., Innov. Food Sci. Emerg. Technol.,2009; 10: 155-159. DOI10.1016/j.ifset.2008.11.007.
[18]SoodA.andGuptaM.,Food Biosci.,2015;12: 100-106. DOI 10.1016/j.fbio.2015.09.004.
[19]BrummellD.A.,DalCinV.,CrisostoC.H.andLabavitchJ.M.,J. Exp. Bot.,2004;55: 2029-2039.
[20]HowardL.R.,TalcottS.T.,BrenesC.H.andVillalonB.,J. Agric. Food Chem.,2000;48: 1713-1720.
[21]Smith M.B. and March J., March´s Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 6th Edn., JohnWileyandSons,NewJersey,2013.
[22]Dawidowicz A.L. and Olszowy M.,Talanta, 2012; 97: 312-317. DOI10.1016/j.talanta.2012.04.036.
[23]Pękal A. and Pyrzynska K., Int. J. Food Sci. Nutr., 2015; 66: 58-62. DOI10.3109/09637486.2014.959899.
[24]Ou B., Huang D., Hampsch-WoodillM.,FlanaganJ.A.andDeemerE.K.,J. Agric. Food Chem., 2002;50: 3122-3128. DOI10.1021/jf0116606.
[25]ShalabyE.A.andShanabS.M.,Indian J. Mar. Sci.,2013;42:556-564.
Chiang Mai J. Sci. 2019; 46(4) 671
[26]LemańskaK.,SzymusiakH.,TyrakowskaB.,ZielińskiR.,SoffersA.E.andRietjensI.M.,Free Radic. Biol. Med.,2001;31: 869-881.
[27]PulidoR.,BravoL.andSaura-CalixtoF.,J. Agric. Food Chem.,2000;48: 3396-3402. DOI10.1021/jf9913458.
[28]Payne A.C., Mazzer A., Clarkson G.J.and TaylorG.,Food Sci. Nutr., 2013; 1: 439-444.DOI10.1002/fsn3.71.
[29]Walker R.B. and Everette J.D., J. Agric. Food Chem., 2009; 57: 1156-1161. DOI10.1021/jf8026765.
[30]Moo-Huchin V.M., Gonzalez-AguilarG.A., Moo-Huchin M., Ortiz-VazquezE., Cuevas-Glory L., Sauri-Duch E.andBetancur-AnconaD.,Chiang Mai J. Sci.,2017:44:605-616.