Research Article Measurement of Capsaicinoids in Chiltepin Hot …downloads.hindawi.com/journals/jchem/2015/709150.pdf · AlbertoGonzález-Zamora, 1,2 ErickSierra-Campos, 3 RebecaPérez-Morales,

Research ArticleMeasurement of Capsaicinoids in Chiltepin Hot PepperA Comparison Study between Spectrophotometric Method andHigh Performance Liquid Chromatography Analysis

Alberto Gonzaacutelez-Zamora12 Erick Sierra-Campos3

Rebeca Peacuterez-Morales3 Cirilo Vaacutezquez-Vaacutezquez1 Miguel A Gallegos-Robles1

Joseacute D Loacutepez-Martiacutenez1 and Joseacute L Garciacutea-Hernaacutendez1

1Facultad de Agricultura y Zootecnia Universidad Juarez del Estado de Durango Km 35 Carretera Gomez Palacio-TlahualiloEjido Venecia 35111 Gomez Palacio DGO Mexico2Laboratorio de Biologıa Evolutiva Facultad de Ciencias Biologicas Universidad Juarez del Estado de DurangoAvenida Universidad SN Fraccionamiento Filadelfia 35010 Gomez Palacio DGO Mexico3Facultad de Ciencias Quımicas Universidad Juarez del Estado de Durango Avenida Artıculo 123 SN Fraccionamiento Filadelfia35010 Gomez Palacio DGO Mexico

Correspondence should be addressed to Erick Sierra-Campos ericksiergmailcom

Received 6 November 2014 Accepted 23 December 2014

Academic Editor Maria Roca

Copyright copy 2015 Alberto Gonzalez-Zamora et al This is an open access article distributed under the Creative CommonsAttribution License which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

Direct spectrophotometric determination of capsaicinoids content in Chiltepin pepper was investigated as a possible alternativeto HPLC analysis Capsaicinoids were extracted from Chiltepin in red ripe and green fruit with acetonitrile and evaluatedquantitatively using the HPLC method with capsaicin and dihydrocapsaicin standards Three samples of different treatment wereanalyzed for their capsaicinoids content successfully by these methods HPLC-DAD revealed that capsaicin dihydrocapsaicinand nordihydrocapsaicin comprised up to 98 of total capsaicinoids detected The absorbance of the diluted samples was readon a spectrophotometer at 215ndash300 nm and monitored at 280 nm We report herein the comparison between traditional UVassays and HPLC-DAD methods for the determination of the molar absorptivity coefficient of capsaicin (120576

280= 3 410 and

120576280= 3 720Mminus1 cmminus1) and dihydrocapsaicin (120576

280= 4 175 and 120576

280= 4 350Mminus1 cmminus1) respectively Statistical comparisons

were performed using the regression analyses (ordinary linear regression and Deming regression) and Bland-Altman analysisComparative data for pungency was determined spectrophotometrically and by HPLC on samples ranging from 2955 to 129mggwith a correlation of 091These results indicate that the twomethods significantly agreeThe described spectrophotometricmethodcan be routinely used for total capsaicinoids analysis and quality control in agricultural and pharmaceutical analysis

1 Introduction

Pepper (Capsicum annum L) is an important horticulturalcrop of major economic significance globally imparting fla-vor aroma and color to foods [1] The demand for new culti-vars with high levels of phytochemicals has received increas-ing attention for biochemists pharmaceutical companiesplant breeders and general public due to their health benefits

Chiltepines (Capsicum annuum var glabriusculum) are awild relative of domesticated peppers that grow throughoutMexico in undisturbed sites of tropical deciduous forestsand orchards in pasture grasslands and along roads [2 3]Chiltepin production in Mexico has been estimated at 50tonsyear having great importance for subsistence farmers ofthe central and northern regions of the country [3 4] TheareawhereChiltepin peppers are commercially harvested and

Hindawi Publishing CorporationJournal of ChemistryVolume 2015 Article ID 709150 10 pageshttpdxdoiorg1011552015709150

2 Journal of Chemistry

recognized for the quality flavor is in the mountains in thestate of Sonora Mexico which rarely exceed 1000m altitude[5]

Montoya-Ballesteros et al [6] observed that Chiltepinfruits in green and red stage ofmaturity showed differences incolor pungency and capsaicin and dihydrocapsaicin contentThese results suggest that during fruit ripening several bio-chemical physiological and structural modifications occurand these changes determine the attributes of fruit quality

In relation to the importance of phytochemicals for func-tional aspects of pepper the capsaicinoids are a specific classof compounds that causes the spicy sensation (pungency) ofchili pepper fruit The main capsaicinoid is capsaicin fol-lowed by dihydrocapsaicin nordihydrocapsaicin homodihy-drocapsaicin and homocapsaicin Of these compounds cap-saicin and dihydrocapsaicin account for approximately 90of capsaicinoids in chili pepper fruit These compounds arethe twomost potent capsaicinoids theirmolecules differ onlyin the saturation of an acyl group [7] and their amount rep-resents a quality parameter for different kind of products Inaddition capsaicin has been used traditionally for muscularpain and headache and to improve circulation and also com-monly added to herbal formulations because it acts as a cata-lyst for other herbs and aids in their absorption Moreovercapsaicin and other capsaicinoids have shown strong evi-dence that have promising potential in the fight against manytypes of cancer [8] High concentrations of capsaicin occur inChiltepin (C annuum L var glabriusculum (Dunal) Heiser ampPickersgill) [9] Bhut Jolokia (C chinense Jacq) [10] and theamounts vary with genotype fruit maturity and conditionsof cultivation [11 12]

The fertilization regime also affects the concentrationsof many secondary metabolites such as capsaicinoids andphenols [13 14] Golcz et al [15] stated that chili pepper hasthe greatest requirement for potassium (40) and nitrogen(31) in relation to the total amount of absorbed nutrientsLittle is currently known about the effect of soil nutrients onpepper fruit pungency Nitrogen availability may have amoredirect effect on capsaicin accumulation since the synthesis ofa single capsaicin molecule requires 3mol of nitrogen to beformed [16] Hence nitrogen availability may affect pepperpungency through its content in the fruit tissues On the otherhand potassium may also affect pepper pungency given itspositive effect on fruit development [17] Previously somestudies have documented N and K effect on hot pepper [1819] However the results obtained by researchers suggest thatK apparently does not notably interfere in capsaicin metabo-lization

In the past organoleptic tests such as the Scoville heattest were used to determine the pungency in peppers [20]These methods have been replaced by analytical methodsas colorimetry [21] gas chromatography [22] liquid chro-matography GCLC-MS [23] and NMR-flow probe analysis[24] Quantification of capsaicinoids often employs separa-tion methods combined with ultraviolet (UV) detection [25]Among these the most widely used is high performance liq-uid chromatography (HPLC) which has become one of themost important tools in the separation and identification ofcapsaicinoids from raw pepper fruit extracts In this method

a 46mm times 250mm C-18 column (10 120583m packing) is usedwith mobile phase consisting of a mixture of acetonitriledioxane water methanol and perchloric acid dependingon the sample concentration For samples with more than700 ppm absorption UV detection at 280 nm is used Forsamples with less than 700 ppm fluorescence detection isemployed with excitation at 288 nm and emission monitoredat 320 nm [26]

At present evaluation of capsaicinoids content in chilipeppers is of great interest to plant breeders food technol-ogists and nutritionists and also to assess breeding lines forgenetic selection purposes or monitor levels during storageor processing This interest has underscored the need forrapid and efficient quantification methods HPLC methodsalthoughmore precise are costly and require a trained opera-tor Simple spectrophotometric assays allowmore rapid anal-ysis [27 28] However ordinary UV-visible spectrophotome-try at a single wavelength generally requires that the analytebe separated from the other absorbing constituents in thecomplex sample matrix before the absorbance measurementsaremadeDavis et al [29] proposed a novel spectrophotomet-ric method for the determination of capsaicinoids in haba-nero pepper extracts that does not require prior analyte sep-aration We therefore took advantage of the enhanced sen-sitivity of currently available spectrophotometer (absorbancemeasured down to a sensitivity of 00001) to develop an assaythat uses capsaicinoids The theory of the calculation of themolar absorptivity is briefly illustrated

We address here the precision of spectrophotometer val-ues as a reflection of total amount of capsaicin and dihydroca-psaicin which represents the majority of capsaicinoids cont-ent in chili peppers and we compared values obtained withsimple spectrophotometer readings with the cumulative am-ounts of individual capsaicinoids identified through HPLCanalysis The presence of correlation between the two meth-ods is disclosed and discussed

2 Experimental

21 Reagents All the reagents standards (capsaicingt95 di-hydrocapsaicin about 90 from Capsicum sp) and solventsused were of high degree of purity or HPLC grade and werepurchased from Sigma-Aldrich (St Louis MO USA)

22 PlantMaterial andGrowthConditions A sample of seedsof three different semidomesticated cultivars (named hereCh1 Ch2 and Ch3) was obtained from the semiarid regionsof the state of Sonora in Northwest Mexico These threecultivars are not significantly different morphologically andgrow in different regions of the Sonoran Desert where peopleharvest the fruits and aid in seed dispersalThe three cultivarswere cultivated in an experimental area within a rural land ofthe municipality of Francisco I Madero Coahuila Mexico(25∘ 461015840 3110158401015840 N 103∘ 161015840 2310158401015840 W) In this region the averagealtitude is 1105m the climate is predominantly semidry theaverage annual temperature is 22∘C with maximum temper-atures of 45∘C in summer and minimum of 4∘C in winterand the average annual rainfall is less than 400mm [30]

Journal of Chemistry 3

According to technical recommendations by Molina et al[31] seeds were treated before germination as follows theywere soaked in water without chlorine for about 72 h andafterward they were germinated in greenhouse conditionsat 25∘C in Jan 2013 Seeds were placed within polystyrenetrays of 200 cavities filled with a mixture of peat moss andvermiculite as substrate one seed per cavity was placed ata depth of 5mm from the base and sprayed with water Thetrays were placed under shade at 22∘C and covered with blackplastic Irrigation was performed once a week until the dayof transplant which was performed 82 days after the date ofgerminationThe transplant was directly from the trays to thefield when seedlings had an approximateminimumheight of20 cm in Apr 2013 Distance between plants and rows was1m Vermicompost was used as fertilizer in an amount of45 t haminus1 The irrigation drip was provided every 20 days atfield capacityThe fruitwas harvestedGM in July 2013 77 daysafter to transplant and then RR 87 days after transplant

23 Collection of Fruits from Plant Materials Samples werecollected at two stages of maturity green mature (fresh) andred ripening and then dried using a cabinet-type convectivedryer at 65∘C by 24 h and triturated

24 Capsaicinoids Extraction The capsaicinoids were extra-cted from the dried fruits following the methods proposedby Al Othman et al [32] and Parrish [33] slightly modifiedas follows briefly the dried fruits were powdered and 1 g wastreated with 10mL of acetonitrile at 65∘C along 20min undersonication with a working frequency of 35 kHz The extractswere evaporated to dryness at 60∘C resuspended in 05mLof acetonitrile and filtrated through 045 120583mcellulose acetatemembrane filter (GVS Filter Technology Indianapolis INUSA) Samples were stored atminus20∘Cuntil theywere analyzed

25 Capsaicinoids Analysis byHPLC-DAD Chromatographyconditions were based on the validated method reportedpreviously [9] The analyses of capsaicinoids were performedby HPLC-DAD (Agilent 1200 Agilent Technologies PaloAlto CAUSA) employing a reversed phase columnKromasilEternity-5-C18 (46 times 150mm) with Precolumn (SUPELCOAnalytical Sigma-Aldrich) at 25∘C Elution was performedwith an isocratic mixture of water acetonitrile 50 50 Detec-tion was set at 202 222 and 280 nm Injection volumewas 20120583L All peaks related to capsaicinoids were eluted inabout 15min Quantitative analysis was performed followingthe external standard method Calibration curves were builtby injecting by triplicate of ten increasing concentrationsof standard Nordihydrocapsaicin was quantified using thecalibration curve of dihydrocapsaicin since it was not com-mercially available

26 Spectrophotometric Method The absorbance of the unk-nown samples solutions was read using one cm quartz cell ina UV-Vis spectrophotometer (HACH DR-5000)

Preparation of Standard Stock Solution Pure capsaicin ordihydrocapsaicin was weighed and dissolved in acetonitrile

up to 20mL to get the stock solution of 2mgmL Thesolution was then filtered with 045120583m syringe filter Thestock solution of capsaicin or dihydrocapsaicin was diluted asrequired Calibrate daily with at least six working standardsover the range of 10 to 200mgL for each standard Thesedilutions were scanned in the UV-Vis spectrometer and 120582maxof capsaicin was selected as wavelength of detection Theinstrument was set at 280 nm The capsaicinoids content inthe unknown extract solutions was calculated based on theabsorbance values of known standard solutions

27 Pungency Capsaicinoid content was converted to Scov-ille heat unit (SHU) by multiplying the concentration of cap-saicinoid in dry weight of pepper in parts per million (ppm)by the coefficient of the heat value for each compound 93 fornordihydrocapsaicin and 161 for both capsaicin and dihydro-capsaicin for HPLC [22]

Total SHU

= [C (ppm) + DHC (ppm)] times 161

+ [n-DHC (ppm) times 93]

(1)

Then the conversion to SHU from total capsaicinoids contentobtained by spectrometric method in dry weight of pepperwas done by multiplying the coefficient corresponding to theheat value for capsaicin which is 16 times 107 [34]

28 Statistical Analysis Experimental data are shown as themean plusmn standard error of assay run in triplicate for capsaicin-oids contentThe statistical test was anANOVA for which weused SPSS version 150 forWindowsThemeans of treatmentswere compared with Tukeyrsquos multiple range test (119875 le005) Linear correlation analysis for total capsaicinoids usingHPLC and spectrophotometric method was conducted usingDeming regression analysis with Bland-Altman plots Allstatistical calculations of correlation were performed usingMedcalc version 123 (Mariakerke Belgium)

3 Results and Discussion

31 Calculation of the Molar Absorptivity Coefficient by HPLCand Spectrometric Method Few attempts have been made tocalculate absorption coefficient for capsaicinoidsThe major-ity of capsaicinoids exhibit absorption in the UV region ofthe spectrum between 200 and 350 nm Because these obeythe Beer-Lambert law (absorbance is linearly proportional tothe concentration) absorbance measurement can be used toquantify the concentration of a pure (standard) capsaicin-oid or to estimate the total capsaicinoid concentration in amixture or extract of capsaicinoids in a sample Standardsof capsaicinoids (capsaicin and dihydrocapsaicin) commer-cially purchased were dissolved in acetonitrile and the spec-trum was scanned in order to evaluate their fine structure(Figures 1(a) and 1(b)) Using the absorbance measured at itsmaximum wavelength (280 nm) and taking into considera-tion the dilution factor purity degree and molecular mass(capsaicin 30541 gmol dihydrocapsaicin 30743 gmol) the


200 250 300 3500

05

1

15

2

25

Abso

rban

ce

120582 (nm)

000020040060080100

0 20 40 60 80 100 120Ab

sorb

ance

Concentration (mgL)

y = 0009x + 00384

R2 = 09942

100mgL80mgL60mgL

40mgL20mgL10mgL

(a)

200 250 300 350120582 (nm)

100mgL80mgL60mgL

40mgL20mgL10mgL

0

05

1

15

2

25

Abso

rban

ce 000020040060080100120

0 20 40 60 80 100 120

Abso

rban

ce

Concentration (mgL)

y = 00106x + 00331

R2 = 0999

(b)

Figure 1 UV-Vis spectra of different concentrations of capsaicin (a) and dihydrocapsaicin (b) Insert calibration curve of each capsaicinoidThe regression line equations for capsaicin and dihydrocapsaicin were (119910 = 00097119909 + 003) and (119910 = 00106119909 + 0031) respectively

molar absorption of each standard capsaicinoid was calcu-lated using the following well-known equation

119860 = 120576 sdot 119887 sdot 119888 (2)

where 119860 is absorbance (dimensionless) 119887 is the path lengththrough the sample in cm 120576 is the molar absorbance coeffi-cient in liter molminus1 and 119888 is the molar concentration in molliterminus1

Regarding linearity spectrophotometric test exhibited alinear response between 10 120583gmL and 100 120583gmL and therepresentative equation of analysis was 119910 = 0009119909 + 0038a collection of 3 calibration curves presented a correlationcoefficient 0994 for capsaicin and 119910 = 001119909 + 0033 witha correlation coefficient of 099 for dihydrocapsaicin (insetFigures 1(a) and 1(b)) The molar absorptivity coefficient ofcapsaicin is (120576

280= 3410Mminus1 cmminus1) and of dihydrocapsaicin

is (120576280

= 4175Mminus1 cmminus1) LOD and LOQ were found to be0070 and 0213 120583gsdotmL for capsaicin while LOD and LOQfor dihydrocapsaicin were 0161 and 049120583gsdotmL respectivelyThe results indicated excellent recoveries ranging from 985to 993 and 99 to 1016 for capsaicin and dihydrocapsaicinrespectively and reliability of the method

The same experimental conditions were applied to allcapsaicinoids obtained by theHPLC-DADmethod Lambert-Beerrsquos law is valid for a DAD detector which is a spectropho-tometer submitted to a dynamic and continuous HPLC flowoperating on chromatographically separated molecules Asproposed by Locatelli et al [35] (3) can be used when 120576 is cal-culated as the slope of a straight-line regression built usingthe injected mass and the corresponding areas recorded bythe DAD detector as follows

Area = 120576 sdot006 sdot 119887 sdot 119898

0

119865 sdotMW (3)

Area is expressed in (120583AUs) MW (gmolminus1) is the molecularweight and 119898

0(ng) is the injected mass The application of

(3) at an HPLC flow of 10mLminminus1 yielded for capsaicin (C)120576280= 3 720 plusmn 36Mminus1 cmminus1 and for dihydrocapsaicin (DHC)

120576280= 4 350 plusmn 30Mminus1 cmminus1 The ratio between C and DHC 120576

values at 280 nm was 120576C 280120576DHC 280 = 0855 Similarlythe lower limit of quantitation (LOQ) was determined byinjecting a standard at a concentration that resulted in an SNratio of 10The determination of the limit of detection (LOD)was analysed on the basis of SN ratio of 3 The LOD andLOQ for this method were 00038 120583gsdotmL and 00116120583gsdotmLfor capsaicin respectively Relative standard deviation (RSD)of the within-day and day to day was 002ndash005 and 008ndash01 for capsaicin and dihydrocapsaicin respectively

32 Assessment of the Accumulation of Capsaicinoids at TwoRipening Stages in Chiltepin Fruits The pungent metabolitesin the fruits of Capsicum species are called capsaicinoidswhich are a group of 12 or more alkaloids with a structureof vanillylamide of branched fatty acids with 9ndash11 carbons[36] but capsaicin and dihydrocapsaicin are responsible formore than 90 of the pungency [37] The capsaicin contentin chili peppers is variable and ranges from 01 to 1 of thefruit weight approximately but the amount varies dependingon the temperature at which the plant is grown the age ofthe fruit the light and soil composition [32] The addition ofmineral supplements to the pepper crops causes an increasein the capsaicinoid content of the fruit [38] and this nitrogensupply is essential for the synthesis of such compound[39] However due to adverse effects of chemical fertilizersinterest has been stimulated for the use of organic manuresVermicompost is produced by biodegradation of organicmaterial through interactions between earthworms andmicr-oorganisms [40]


Chiltepin has high phenotypic plasticity shown by thevariation of traits such as leaf morphology fruit shape seedgermination pattern or resistance to pathogens [2] In Mex-ico it can be found from the Yucatan peninsula and the Gulfof Mexico where it grows in deep soils with dense evergreenvegetation to xeric regions in the Sonoran desert or thecentral plateau where it is commonly associated with nursetrees [41] Harvesting the fruits of Chiltepin is still a commonpractice in central and northernMexico and the total harvesthas been estimated to be 50metric tons per year [3] Howeverenvironmental degradation grazing and harvesting haveseverely impacted many native populations of this species[41]

In the present work we have determined the contentof capsaicin dihydrocapsaicin and nordihydrocapsaicin byHPLC-DAD to compare the total capsaicinoids level obtainedby spectrometric method to study the influence of maturitystage and vermicompost on quality in capsaicinoids compo-sition in Chiltepin fruits

Chiltepin chili samples showed a high concentration ofcapsaicinoids in the two stages of maturity GM and RR(Table 1) Nordihydrocapsaicin capsaicin and dihydrocap-saicin were observed in the fruit of all samples However theamount of nordihydrocapsaicinwas lower (5 to 9) than cap-saicin (30 to 59) and dihydrocapsaicin (32 to 63) relativeto the total of capsaicinoids Homocapsaicin and homodi-hydrocapsaicin were only present in some samples and insmall percentages (from less than 1 to 4) compared tototal capsaicinoids contentHPLC revealed that capsaicin anddihydrocapsaicin comprised up to 93 of total capsaicinoidscontent detected in the studied samples Capsaicin contentof the control samples without vermicompost ranged from163plusmn004 to 541plusmn029mgg dryweight in red ripe fruit Lev-els of capsaicin assessed of sampleswith vermicompost variedfrom 323plusmn018 to 643plusmn022mgg dryweight in red ripe fruitCapsaicin content was higher in green mature fruit than redripe fruit (see Table 1) The content ranged from 2378 plusmn 087to 4014 plusmn 025mgg dry weight in green mature sampleswithout vermicompost (control) and 4202 plusmn 193 to 5668 plusmn149mgg dry weight in green mature samples treated withvermicompost In most cases capsaicin levels were greaterthan dihydrocapsaicin except for Ch1 RR-Cr Ch1 Gm-Crand Ch2 Gm-Cr Ch2 GM-Vc was the cultivar showing thehighest values of total capsaicinoids (10701 plusmn 155mg gminus1)and therefore the highest value of pungency Lowest values ofcapsaicinoids were observed in Ch1 to GM (7112 plusmn 251mggminus1) and Ch3 to RR (3070 plusmn 107mg gminus1) except for thecase of dihydrocapsaicin where Ch1 to GM (2949 plusmn 132mggminus1) and Ch3 to RR (1196 plusmn 026mg gminus1) showed the lowestvaluesThe difference between the lowest and highest value is35-fold

The direct-absorbance measurement of total capsaici-noids levels could have interferences from other compoundspresent in the pepper fruit extractsTherefore after the valid-ation byHPLC we evaluated the spectrophotometricmethodapplied for total capsaicinoids in Chiltepin fruit samplesFigure 2 shows the typical spectrum of Ch1 RR-Cr and Ch1RR-Cr-C+DHC complex overlain spectra and the results

0

02

04

06

08

1

12

14

16

200 250 300 350

Abso

rban

ce

120582 (nm)

Ch1 RR-Cr 008mL (dilutionC + DHCCh1 RR-Cr dilution

10mgL10mgL1 1000) +

1 1000

Figure 2 UV absorption spectrum of dry Chiltepin fruit extractwithout standards (white circle) and with standards (capsaicin ldquoCrdquoand dihydrocapsaicin ldquoDHCrdquo black circle)

obtained are shown in Table 1 in accordance with the HPLC-DAD analysis Recoveries obtained for the standards didnot differ significantly from 100 showing that there wasno interference from other common compounds present inextract and thus indicating accuracy and reliability of themethod

The capsaicinoids content as determined by the spec-trophotometric method was generally higher in GM ripefruit than red fruit (see Table 1) The content varied from2018ndash9773mgg for RR samples to 5947ndash11893mgg for GMstage Samples showed a considerable percentage of decreasein capsaicinoids levels depending on the stage of maturationfromGM toRR Ch3was the cultivar that showed the greatestvariation in the capsaicinoids content The values of thesemetabolites in RR stage decreased by 628 512 and 456for nordihydrocapsaicin capsaicin and dihydrocapsaicinrespectively This produces fruits with an average 50 lesspungency in the RR stage regarding GM For Ch1 and Ch2pungency levels are 20 lower in RR than in GM

33 Effects of Vermicompost Fertilization on CapsaicinoidsContent in Chiltepin Pepper Fruit Vermicompost and com-post can meet the nutrient demand of crop and significantlyreduce the excessive use of chemical fertilizers in agriculture[42] and for vermicompost in particular increase soil fer-tility without polluting the soil as well as the quantity andquality of harvested product [43] According to Ramesh et al[44] organic production is an alternative for consumers whoprefer food free of pesticides and synthetic risk-free fertiliz-ers with high nutritional value and growingmedia promotesthe development and productivity of pepper [45 46]

Under the studied conditions the fruit in the vermicomp-ost treatment experienced very high capsaicinoids accumula-tion in different samples The results presented in Table 1clearly demonstrated that vermicompost had a significanteffect on the capsaicin content of Chiltepin peppers this


Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(

RR)greenmature(GM)control(Cr

)andverm

icom

postsupp

lement(Vc

)no

rdihydrocapsaicin(n-D

HC)

capsaicin

(C)anddihydrocapsaicin

(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20

Mean of HPLC and SPT

HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140

Mean of SPT and SPT1

Mea

n of

HPL

C an

d H

PLC1

(b)

Figure 3 Bland-Altman plot for total capsaicinoids in Chiltepin fruit extracts (a) The solid line represents the mean difference in measuredcapsaicinoids concentrations betweenmethods and the dashed lines are plusmn2 SDMean difference minus37 standard deviation 196 and 95 limitof agreement minus218 to 143 (b) By Deming regression of the scatter plot the slope is 0716 the intercept 1759 and the correlation coefficient(119903) = 088 These results indicate a fair degree of correlation between HPLC and spectrometric (SPT) values

treatment showed about two times more capsaicin thancontrol treatment Also dihydrocapsaicin concentration wasaffected significantly by the vermicompost treatment butconsiderable variability was observed for this capsaicinoidin the different samples The accumulation of capsaicinoidsdepends on synthesis and degradation Capsaicinoids synthe-sis generally beginswith oneweek after fruit set and continuesas placental tissues developThe compounds reachmaximumlevels approximately 40ndash50 days after fruit set [47 48] Uponmaturation capsaicinoids levels in most cultivars show anoticeable variable rate of decline associated with an increasein the activity of peroxidase enzymes in ripe fruit [49 50]Our results have also indicated that capsaicin levels in GMstage were higher than in RR stage and vermicompostfertilization does not alter this behavior This is apparentlythe first report in Chiltepin pepper of fertilization treatmentaffecting the metabolism of capsaicinoids responsible forpungency We can deduce that in RR stage the peroxidasesmay be involved in the degradation of capsaicinoids

34 Spectrophotometer Compared toHPLC It is assumed thatspectrophotometric estimation of the capsaicinoids is inac-curate because of interference of other compounds presentin the extract However in a previous study high levels ofagreement were found between colorimetric and chromato-graphic techniques [21] Although HPLC measurements arein general more precise and reproducible than spectropho-tometric methods these techniques can be an alternative todetermine the total capsaicinoids in spite of the fact that theirefficacy is limited perhaps partly because of carotenoids andchlorophylls Thus such testing requires standardized andreproducible techniques

The agreement between quantification by HPLC-DADand spectrophotometric method was evaluated by the graph-ical investigation of difference-plots [51] and by Deming

regression analysis [41] Difference-plots are useful tools todisplay differences betweenmethods but they do not providestatistical tools for inference of the differences The Demingregression analysis is suitable if measurement errors are pro-portional and the procedure accounts for errors in bothmethods [3] In this case samples were not analyzed in repli-cates and equal variance between the two methods was assu-med Medcalc was used for Deming regression analysis anddifference-plots

In difference-plot (Figure 3(a)) and Deming regressions(Figure 3(b)) for capsaicinoids in Chiltepin fruit extracts thedifferences between the two methods (HPLC values subtrac-ted from spectrophotometer values) were plotted against themean concentrations shown by the two methods (Figure 3)The mean differences are shown (solid lines) as are thelimits of agreement (broken lines) corresponding to meanplusmn2 SD Concentrations quantified by HPLC-DAD and spec-trophotometer are plotted against each other (dots) fortotal capsaicinoids (Figure 3(a)) and the Deming regression(capsaicinoids 119910 = 1759 + 0716119909) is indicated (unbrokenlines) as the identity lines (broken lines)

When comparing simple spectrophotometric and HPLCquantificationmethods identical trends among varieties afterthe performance of means separation were revelated betweentotal capsaicinoids concentrations (spectrophotometer) andtotal capsaicinoids concentration (HPLC) (Figure 4) A highlinear relationship (119903 = 0913 119875 lt 0001) was observed aftercorrelation analysesDifferences between spectrometricmea-surements and HPLC analyses in total capsaicinoids contentamong different samples have shown slightly higher 2 to 26difference ranging 2ndash24mgkgminus1 dry wt (Table 1)

Total capsaicinoids quantified byHPLC correlated closelywith spectrophotometer values The correlation coefficientwas 091 (119875 lt 0001) for capsaicinoids in Chiltepin pepperfruit indicating a strong positive association between these


0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120

HPLC-DAD total capsaicinoids (mgg dwt)

tota

l cap

saic

inoi

ds (m

gg d

wt)

Figure 4 Correlation between cumulative capsaicinoids of Chilte-pin acetonitrile extract separated by HPLC and total capsaicinoidsby spectrophotometer (119903 = 091) 119899 = 11

values The simple spectrophotometric procedure proved tobe a valid efficient and cost-effective method for the quan-tification of total capsaicinoids in these conditions which infact adequately represent total capsaicinoids content in hotchili pepper extracts

4 Conclusion

This study describes a spectrophotometric method for theprecise determination of total capsaicinoids in pepper fruitsamples and yields a close estimation but it is not as accurateas HPLC-DAD Further investigation is needed to under-stand the origin of these analytical errors The method is aneffective mean for quality control and is a viable alternative tothe existing analytical methods for routine analyses allowinga rapid and accessible quantitation of capsaicinoids in fruitssamples without any time-consuming sample separation Inaddition our results indicate that in contrast to other studieswhere capsaicin content was generally higher in ripe fruitthan green fruit Chiltepin red ripening fruits showed lowerlevels of capsaicin and dihydrocapsaicin than green maturefruits in all samples Vermicompost treatment affected cap-saicin and dihydrocapsaicin levels and favors the accumula-tion of capsaicin

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This studywas possible with the support of researchGrant no2009-134382 from Consejo Nacional de Ciencia y Tecnologıain Mexico (CB-SEP-CONACYT) The first author was sup-ported by a Doctoral Research Fellowship no 327844 CVU

no 164727 (CONACYT) Erick Sierra-Campos acknowledg-ed SNI (CONACYT) for the stipend receivedThe authors arevery thankful to Monica A Valdez-Solana Jaime A de Lira-Sanchez Jorge A Sosa-Gutierrez and Eliab M Gonzalez-Olvera for helpful discussions and critical reading of thispaper and to RaymundoMontelongo Miguel de la Cruz andJ Guadalupe Luna for agronomic support and work in theexperimental field

References

[1] N Caporaso A Paduano G Nicoletti and R Sacchi ldquoCapsai-cinoids antioxidant activity and volatile compounds in olive oilflavoredwith dried chili pepper (Capsicum annuum)rdquoEuropeanJournal of Lipid Science and Technology vol 115 no 12 pp 1434ndash1442 2013

[2] S Hernandez-Verdugo R Luna-Reyes andKOyama ldquoGeneticstructure and differentiation of wild and domesticated popu-lations of Capsicum annuum (Solanaceae) from Mexicordquo PlantSystematics and Evolution vol 226 no 3-4 pp 129ndash142 2001

[3] E J Votava G P Nabhan and PW Bosland ldquoGenetic diversityand similarity revealed via molecular analysis among andwithin an in situ population and ex situ accessions of chiltepın(Capsicum annuum var glabriusculum)rdquoConservationGeneticsvol 3 no 2 pp 123ndash129 2002

[4] J G Almaza R K Maiti P R Foroughbakhch et al ldquoBro-matologıa del chile piquın (Capsicum annuum L var aviculare(Dierb) D amp E)rdquo in Resumenes 15th Congreso Mexicano deBotanica Queretaro Mexico 2001

[5] W G DrsquoArcy and W H Eshbaugh ldquoNew world peppers(Capsicum-Solanaceae) north of Colombia a resumerdquo Baileyavol 19 pp 93ndash105 1974

[6] L C Montoya-Ballesteros A Gardea-Bejar G M Ayala-Chavez Y Y Martınez-Nunez and L E Robles-Ozuna ldquoCap-saicinoides y color en chiltepın (Capsicum annuum var avicu-lare) processing effect on sauces and picklesrdquo Revista Mexicanade Ingenieria Quımica vol 9 no 2 pp 197ndash207 2010

[7] M A Bernal A A Calderon M A Pedreno R Munoz A RBarcelo and FMerinoDeCaceres ldquoCapsaicin oxidation by per-oxidase from Capsicum annuum (var annuum) fruitsrdquo Journalof Agricultural and Food Chemistry vol 41 no 7 pp 1041ndash10441993

[8] X-F Huang J-Y Xue A-Q Jiang and H-L Zhu ldquoCapsaicinand its analogues structure-activity relationship studyrdquoCurrentMedicinal Chemistry vol 20 no 21 pp 2661ndash2672 2013

[9] A Gonzalez-Zamora E Sierra-Campos J G Luna-OrtegaR Perez-Morales J C R Ortiz and J L Garcıa-HernandezldquoCharacterization of different Capsicum varieties by evaluationof their capsaicinoids content by high performance liquidchromatography determination of pungency and effect of hightemperaturerdquoMolecules vol 18 no 11 pp 13471ndash13486 2013

[10] Y Liu andMG Nair ldquoCapsaicinoids in the hottest pepper BhutJolokia and its antioxidant and antiinflammatory activitiesrdquoNatural Product Communications vol 5 no 1 pp 91ndash94 2010

[11] Y Zewdie and P W Bosland ldquoEvaluation of genotype environ-ment and genotype-by-environment interaction for capsaici-noids inCapsicum annuum Lrdquo Euphytica vol 111 no 3 pp 185ndash190 2000

[12] G F Barbero A G Ruiz A Liazid M Palma J C Vera and CG Barroso ldquoEvolution of total and individual capsaicinoids inpeppers during ripening of the Cayenne pepper plant (Cap-sicum annuum L)rdquo Food Chemistry vol 153 pp 200ndash206 2014


[13] J P Bryant T P Clausen P B Reichardt M C McCarthyand R A Werner ldquoEffect of nitrogen fertilization upon the sec-ondary chemistry and nutritional value of quaking aspen(Populus tremuloides Michx) leaves for the large aspen tortrix(Choristoneura conflictana (Walker))rdquo Oecologia vol 73 no 4pp 513ndash517 1987

[14] P Gremigni J Hamblin D Harris and W A Cowling ldquoTheinteraction of phosphorus and potassium with seed alkaloidconcentrations yield and mineral content in narrow-leafedlupin (Lupinus angustifolius L)rdquo Plant and Soil vol 253 no 2pp 413ndash427 2003

[15] A Golcz P Kujawski and B Markiewicz ldquoYielding of red pep-per (Capsicum annuum L) under the influence of varied potas-sium fertilizationrdquo Acta Scientiarum PolonorummdashHortorumCultus vol 11 no 4 pp 3ndash15 2012

[16] J Curry M Aluru M Mendoza J Nevarez M Melendrezand M A OrsquoConnell ldquoTranscripts for possible capsaicinoidbiosynthetic genes are differentially accumulated in pungentand non-pungent Capsicum spprdquo Plant Science vol 148 no 1pp 47ndash57 1999

[17] D T Clarkson and J B Hansonn ldquoThe mineral nutrition ofhigher plantsrdquo Annual Review of Plant Physiology vol 31 pp239ndash298 1980

[18] F Medina-Lara I Echevarrıa-Machado R Pacheco-ArjonaN Ruiz-Lau A Guzman-Antonio and M Martinez-EstevezldquoInfluence of nitrogen and potassium fertilization on fruitingand capsaicin content in habanero pepper (Capsicum chinenseJacq)rdquo HortScience vol 43 no 5 pp 1549ndash1554 2008

[19] C D Johnson and D R Decoteau ldquoNitrogen and potassiumfertility affects Jalapeno pepper plant growth pod yield andpungencyrdquo HortScience vol 31 no 7 pp 1119ndash1123 1996

[20] W L Scoville ldquoNote on capsicumsrdquo Journal of the AmericanPharmaceutical Association vol 1 no 5 pp 453ndash454 1912

[21] H A A Gibbs and L W O OrsquoGarro ldquoCapsaicin content ofWest Indies hot pepper cultivars using colorimetric and chro-matographic techniquesrdquoHortScience vol 39 no 1 pp 132ndash1352004

[22] P H Todd M G Bensinger and T Biftu ldquoDetermination ofpungency due to capsicumby gas-liquid chromatographyrdquo Jour-nal of Food Science vol 42 no 3 pp 660ndash665 1977

[23] K Iwai T Suzuki H Fujiwake and S Oka ldquoSimultane-ous microdetermination of capsaicin and its four analoguesby using high-performance liquid chromatography and gaschromatography-mass spectrometryrdquo Journal of Chromatogra-phy A vol 172 no 1 pp 303ndash311 1979

[24] N T Nyberg H Baumann and L Kenne ldquoApplication of solid-phase extraction coupled to an NMR flow-probe in the analysisof HPLC fractionsrdquo Magnetic Resonance in Chemistry vol 39pp 236ndash240 2001

[25] S-H Choi B-S Suh E Kozukue N Kozukue C E Levin andM Friedman ldquoAnalysis of the contents of pungent compoundsin fresh Korean red peppers and in pepper-containing foodsrdquoJournal of Agricultural and Food Chemistry vol 54 no 24 pp9024ndash9031 2006

[26] AM Krajewska Isolation identification and sensory evaluationof capsaicinoids [PhD thesis] University of Georgia AthensGa USA 1984

[27] J J DiCecco ldquoSpectrophotometric difference method for deter-mination of capsaicinrdquo Journal Association of Official AnalyticalChemistry vol 62 pp 998ndash1000 1979

[28] D A Nikolaeva ldquoSpectrophotometric determination of cap-saicin in peppers (Capsicum annuum L)rdquo Biokhim MetodyAnaliza Plodov Kishinev pp 99ndash102 1984

[29] C BDavis C EMarkeyMA Busch andKW Busch ldquoDeter-mination of capsaicinoids in habanero peppers by chemometricanalysis of UV spectral datardquo Journal of Agricultural and FoodChemistry vol 55 no 15 pp 5925ndash5933 2007

[30] Instituto Nacional de Estadıstica y Geografia (INEGI) Prontu-ario de Informacion geografica municipal de los Estados UnidosMexicanos Edited by Francisco I Madero INEGI Coahuila deZaragoza Mexico 2009

[31] C Molina A Morales and A Marquez Tecnicas para elestablecimiento y produccion de chiltepın silvestre bajo un sis-tema agroforestal en Sonora Mexico Capsicum annuum L varglabriusculum (Dunal) Heiser amp Pickersgill CONAFOR ITVYINIFAP Jalisco Mexico 2009

[32] Z A Al Othman Y B H Ahmed M A Habila and AA Ghafar ldquoDetermination of capsaicin and dihydrocapsaicinin Capsicum fruit samples using high performance liquidchromatographyrdquoMolecules vol 16 no 10 pp 8919ndash8929 2011

[33] M Parrish ldquoLiquid chromatographic method of determiningcapsaicinoids in Capsicum and their extractives colaborativestudyrdquo Journal Association of Official Analytical Chemistry vol79 pp 738ndash745 1996

[34] K Sanatombi and G J Sharma ldquoCapsaicin content and pun-gency of different Capsicum spp cultivarsrdquo Notulae BotanicaeHorti Agrobotanici Cluj-Napoca vol 36 pp 89ndash90 2008

[35] M Locatelli G Carlucci S Genovese M Curini and FEpifano ldquoUse of HPLC in the determination of the molarabsorptivity of 41015840-geranyloxyferulic acid and boropinic acidrdquoChromatographia vol 73 no 9-10 pp 889ndash896 2011

[36] T Suzuki and K Iwai ldquoChapter 4 constituents of red pep-per species chemistry biochemistry pharmacology and foodscience of the pungent principle of Capsicum speciesrdquo TheAlkaloids Chemistry and Pharmacology vol 23 pp 227ndash2991984

[37] S Kosuge Y Inagaki and M Nishinura ldquoStudies on pungentprinciples of red pepper IV Paper chromatography of pungentprinciplesrdquo Journal of the Agricultural Chemical Society of Japanvol 33 no 11 pp 915ndash918 1959

[38] B Estrada F Pomar J Dıaz F Merino and M A BernalldquoEffects of mineral fertilizer supplementation on fruit devel-opment and pungency in Padron peppersrdquo The Journal ofHorticultural Science and Biotechnology vol 73 no 4 pp 493ndash497 1998

[39] M Monforte-Gonzalez A Guzman-Antonio F Uuh-Chimand F Vazquez-Flota ldquoCapsaicin accumulation is related tonitrate content in placentas of habanero peppers (Capsicumchinense Jacq)rdquo Journal of the Science of Food and Agriculturevol 90 no 5 pp 764ndash768 2010

[40] C A Edwards and I Burrows ldquoThe potential of earthwormcomposts as plant growth mediardquo in Earthworms in Environ-mental and Waste Management C A Neuhauser Ed pp 211ndash220 SPB Academic Publishing The Hague the Netherlands1988

[41] J J Tewksbury G P Nabhan D Norman H Suzan J Tuxilland J Donovan ldquoIn situ conservation of wild chiles and theirbiotic associatesrdquoConservation Biology vol 13 no 1 pp 98ndash1071999

[42] N Rodrıguez Dimas P Cano Rıos U Figueroa Viramontes etal ldquoTomato production in greenhouse using vermicompost as


substraterdquo Revista Fitotecnia Mexicana vol 31 no 3 pp 265ndash272 2008

[43] A Hernandez H Castillo D Ojeda A Arras J Lopez andE Sanchez ldquoEffect of vermicompost and compost on lettuceproductionrdquo Chilean Journal of Agricultural Research vol 70no 4 pp 583ndash589 2010

[44] P Ramesh M Singh and A Subba Rao ldquoOrganic farming itsrelevance to the Indian contextrdquo Current Science vol 88 no 4pp 561ndash568 2005

[45] N Q Arancon C A Edwards R M Atiyeh and J DMetzger ldquoEffects of vermicomposts produced from food wasteon the growth and yields of greenhouse peppersrdquo BioresourceTechnology vol 93 no 2 pp 139ndash144 2004

[46] S T Lopez-Espinosa A Moreno-Resendez P Cano Rıos NRodrıguez-Dimas V Robledo-Torres and C Marquez-QuirozldquoOrganic fertilization an alternative to produce jalapeno pep-per under greenhouse conditionsrdquo Emirates Journal of Food andAgriculture vol 25 pp 666ndash672 2013

[47] J A Barrera M S Hernandez L M Melgarejo O Martınezand J P Fernandez-Trujillo ldquoPhysiological behavior and qualitytraits during fruit growth and ripening of four Amazonichot pepper accessionsrdquo Journal of the Science of Food andAgriculture vol 88 no 5 pp 847ndash857 2008

[48] E Mueller-Seitz C Hiepler and M Petz ldquoChili pepper fruitscontent and pattern of capsaicinoids in single fruits of differentagesrdquo Journal of Agricultural and Food Chemistry vol 56 no 24pp 12114ndash12121 2008

[49] B Estrada M A Bernal J Dıaz F Pomar and F Merino ldquoFruitdevelopment inCapsicumannuum changes in capsaicin ligninfree phenolics and peroxidase patternsrdquo Journal of Agriculturaland Food Chemistry vol 48 no 12 pp 6234ndash6239 2000

[50] N Deepa C Kaur B George B Singh and H C KapoorldquoAntioxidant constituents in some sweet pepper (Capsicumannuum L) genotypes duringmaturityrdquo LWT Food Science andTechnology vol 40 no 1 pp 121ndash129 2007

[51] S Hernandez-Verdugo R G Guevara-Gonzalez R F Rivera-Bustamante and K Oyama ldquoScreening wild plants of Cap-sicum annuum for resistance to pepper huasteco virus (PHV)presence of viral DNA and differentiation among populationsrdquoEuphytica vol 122 no 1 pp 31ndash36 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Inorganic ChemistryInternational Journal of

Hindawi Publishing Corporation httpwwwhindawicom Volume 2014

International Journal ofPhotoenergy


Carbohydrate Chemistry

International Journal of


Journal of

Chemistry


Advances in

Physical Chemistry

Hindawi Publishing Corporationhttpwwwhindawicom

Analytical Methods in Chemistry

Journal of

Volume 2014

Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

SpectroscopyInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Medicinal ChemistryInternational Journal of


Chromatography Research International


Applied ChemistryJournal of



Theoretical ChemistryJournal of


Journal of

Spectroscopy

Analytical ChemistryInternational Journal of


Journal of


Quantum Chemistry


Organic Chemistry International

ElectrochemistryInternational Journal of



CatalystsJournal of


recognized for the quality flavor is in the mountains in thestate of Sonora Mexico which rarely exceed 1000m altitude[5]

Montoya-Ballesteros et al [6] observed that Chiltepinfruits in green and red stage ofmaturity showed differences incolor pungency and capsaicin and dihydrocapsaicin contentThese results suggest that during fruit ripening several bio-chemical physiological and structural modifications occurand these changes determine the attributes of fruit quality

In relation to the importance of phytochemicals for func-tional aspects of pepper the capsaicinoids are a specific classof compounds that causes the spicy sensation (pungency) ofchili pepper fruit The main capsaicinoid is capsaicin fol-lowed by dihydrocapsaicin nordihydrocapsaicin homodihy-drocapsaicin and homocapsaicin Of these compounds cap-saicin and dihydrocapsaicin account for approximately 90of capsaicinoids in chili pepper fruit These compounds arethe twomost potent capsaicinoids theirmolecules differ onlyin the saturation of an acyl group [7] and their amount rep-resents a quality parameter for different kind of products Inaddition capsaicin has been used traditionally for muscularpain and headache and to improve circulation and also com-monly added to herbal formulations because it acts as a cata-lyst for other herbs and aids in their absorption Moreovercapsaicin and other capsaicinoids have shown strong evi-dence that have promising potential in the fight against manytypes of cancer [8] High concentrations of capsaicin occur inChiltepin (C annuum L var glabriusculum (Dunal) Heiser ampPickersgill) [9] Bhut Jolokia (C chinense Jacq) [10] and theamounts vary with genotype fruit maturity and conditionsof cultivation [11 12]

The fertilization regime also affects the concentrationsof many secondary metabolites such as capsaicinoids andphenols [13 14] Golcz et al [15] stated that chili pepper hasthe greatest requirement for potassium (40) and nitrogen(31) in relation to the total amount of absorbed nutrientsLittle is currently known about the effect of soil nutrients onpepper fruit pungency Nitrogen availability may have amoredirect effect on capsaicin accumulation since the synthesis ofa single capsaicin molecule requires 3mol of nitrogen to beformed [16] Hence nitrogen availability may affect pepperpungency through its content in the fruit tissues On the otherhand potassium may also affect pepper pungency given itspositive effect on fruit development [17] Previously somestudies have documented N and K effect on hot pepper [1819] However the results obtained by researchers suggest thatK apparently does not notably interfere in capsaicin metabo-lization

In the past organoleptic tests such as the Scoville heattest were used to determine the pungency in peppers [20]These methods have been replaced by analytical methodsas colorimetry [21] gas chromatography [22] liquid chro-matography GCLC-MS [23] and NMR-flow probe analysis[24] Quantification of capsaicinoids often employs separa-tion methods combined with ultraviolet (UV) detection [25]Among these the most widely used is high performance liq-uid chromatography (HPLC) which has become one of themost important tools in the separation and identification ofcapsaicinoids from raw pepper fruit extracts In this method

a 46mm times 250mm C-18 column (10 120583m packing) is usedwith mobile phase consisting of a mixture of acetonitriledioxane water methanol and perchloric acid dependingon the sample concentration For samples with more than700 ppm absorption UV detection at 280 nm is used Forsamples with less than 700 ppm fluorescence detection isemployed with excitation at 288 nm and emission monitoredat 320 nm [26]

At present evaluation of capsaicinoids content in chilipeppers is of great interest to plant breeders food technol-ogists and nutritionists and also to assess breeding lines forgenetic selection purposes or monitor levels during storageor processing This interest has underscored the need forrapid and efficient quantification methods HPLC methodsalthoughmore precise are costly and require a trained opera-tor Simple spectrophotometric assays allowmore rapid anal-ysis [27 28] However ordinary UV-visible spectrophotome-try at a single wavelength generally requires that the analytebe separated from the other absorbing constituents in thecomplex sample matrix before the absorbance measurementsaremadeDavis et al [29] proposed a novel spectrophotomet-ric method for the determination of capsaicinoids in haba-nero pepper extracts that does not require prior analyte sep-aration We therefore took advantage of the enhanced sen-sitivity of currently available spectrophotometer (absorbancemeasured down to a sensitivity of 00001) to develop an assaythat uses capsaicinoids The theory of the calculation of themolar absorptivity is briefly illustrated

We address here the precision of spectrophotometer val-ues as a reflection of total amount of capsaicin and dihydroca-psaicin which represents the majority of capsaicinoids cont-ent in chili peppers and we compared values obtained withsimple spectrophotometer readings with the cumulative am-ounts of individual capsaicinoids identified through HPLCanalysis The presence of correlation between the two meth-ods is disclosed and discussed

2 Experimental

21 Reagents All the reagents standards (capsaicingt95 di-hydrocapsaicin about 90 from Capsicum sp) and solventsused were of high degree of purity or HPLC grade and werepurchased from Sigma-Aldrich (St Louis MO USA)

22 PlantMaterial andGrowthConditions A sample of seedsof three different semidomesticated cultivars (named hereCh1 Ch2 and Ch3) was obtained from the semiarid regionsof the state of Sonora in Northwest Mexico These threecultivars are not significantly different morphologically andgrow in different regions of the Sonoran Desert where peopleharvest the fruits and aid in seed dispersalThe three cultivarswere cultivated in an experimental area within a rural land ofthe municipality of Francisco I Madero Coahuila Mexico(25∘ 461015840 3110158401015840 N 103∘ 161015840 2310158401015840 W) In this region the averagealtitude is 1105m the climate is predominantly semidry theaverage annual temperature is 22∘C with maximum temper-atures of 45∘C in summer and minimum of 4∘C in winterand the average annual rainfall is less than 400mm [30]










Total SHU



(1)






200 250 300 3500

05

1

15

2

25

Abso

rban

ce

120582 (nm)

000020040060080100

0 20 40 60 80 100 120Ab

sorb

ance

Concentration (mgL)

y = 0009x + 00384

R2 = 09942

100mgL80mgL60mgL

40mgL20mgL10mgL

(a)

200 250 300 350120582 (nm)

100mgL80mgL60mgL

40mgL20mgL10mgL

0

05

1

15

2

25

Abso

rban

ce 000020040060080100120

0 20 40 60 80 100 120

Abso

rban

ce

Concentration (mgL)

y = 00106x + 00331

R2 = 0999

(b)



119860 = 120576 sdot 119887 sdot 119888 (2)




is (120576280




0

119865 sdotMW (3)












0

02

04

06

08

1

12

14

16

200 250 300 350

Abso

rban

ce

120582 (nm)


10mgL10mgL1 1000) +

1 1000







Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(


)andverm

icom

postsupp

lement(Vc

)no


HC)

capsaicin


(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of










Total SHU



(1)






200 250 300 3500

05

1

15

2

25

Abso

rban

ce

120582 (nm)

000020040060080100

0 20 40 60 80 100 120Ab

sorb

ance

Concentration (mgL)

y = 0009x + 00384

R2 = 09942

100mgL80mgL60mgL

40mgL20mgL10mgL

(a)

200 250 300 350120582 (nm)

100mgL80mgL60mgL

40mgL20mgL10mgL

0

05

1

15

2

25

Abso

rban

ce 000020040060080100120

0 20 40 60 80 100 120

Abso

rban

ce

Concentration (mgL)

y = 00106x + 00331

R2 = 0999

(b)



119860 = 120576 sdot 119887 sdot 119888 (2)




is (120576280




0

119865 sdotMW (3)












0

02

04

06

08

1

12

14

16

200 250 300 350

Abso

rban

ce

120582 (nm)


10mgL10mgL1 1000) +

1 1000







Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(


)andverm

icom

postsupp

lement(Vc

)no


HC)

capsaicin


(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


200 250 300 3500

05

1

15

2

25

Abso

rban

ce

120582 (nm)

000020040060080100

0 20 40 60 80 100 120Ab

sorb

ance

Concentration (mgL)

y = 0009x + 00384

R2 = 09942

100mgL80mgL60mgL

40mgL20mgL10mgL

(a)

200 250 300 350120582 (nm)

100mgL80mgL60mgL

40mgL20mgL10mgL

0

05

1

15

2

25

Abso

rban

ce 000020040060080100120

0 20 40 60 80 100 120

Abso

rban

ce

Concentration (mgL)

y = 00106x + 00331

R2 = 0999

(b)



119860 = 120576 sdot 119887 sdot 119888 (2)




is (120576280




0

119865 sdotMW (3)












0

02

04

06

08

1

12

14

16

200 250 300 350

Abso

rban

ce

120582 (nm)


10mgL10mgL1 1000) +

1 1000







Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(


)andverm

icom

postsupp

lement(Vc

)no


HC)

capsaicin


(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of






0

02

04

06

08

1

12

14

16

200 250 300 350

Abso

rban

ce

120582 (nm)


10mgL10mgL1 1000) +

1 1000







Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(


)andverm

icom

postsupp

lement(Vc

)no


HC)

capsaicin


(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


Table1Con

tent

ofcapsaicino

idsa

ndpu

ngency

levelsin

Chiltepin

chili

samples

cultivatedin

Lagunera

Region

Mexico

Sample

(maturationsta

ge-fe

rtilizatio

n)

HPL

CSpectro

photom

eter

n-DHC

CDHC

CD

HCratio

Total

capsaicino

ids

SHU

Total

capsaicino

ids

SHUlowast

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

mggminus1

dryweight

Ch1R

R-Cr

396plusmn025

bc7

1779plusmn10

4a30

3721plusmn

219

e63

121

5895plusmn346

b922233plusmn54074

bc5338plusmn200

b8594

04plusmn32248

b

Ch1G

M-C

r460plusmn019

cd6

2378plusmn087

b33

4167plusmn14

8f59

118

7004plusmn254

c10

9639

3plusmn39631

cd5947plusmn211b

9575

17plusmn34022

b

Ch1R

R-Vc

463plusmn036

cd8

3124plusmn212

c52

2408plusmn16

1c40

108

5995plusmn409

b933736plusmn6333

3bc

ND

ND

Ch1G

M-Vc

611plusmn033

ef8

4202plusmn19

3d54

2949plusmn13

2d38

107

7762plusmn358

cd12

08086plusmn55466

de7995plusmn099

c12

87262plusmn16010

c

Ch2RR

-Cr

541plusmn029

de7

4308plusmn12

6de

592337plusmn050

bc32

105

7187plusmn204

c1120242plusmn30824

cd8183plusmn254

c13

17507plusmn40

947

c

Ch2GM-C

r855plusmn033

h9

4014plusmn025

d43

4377plusmn024

f47

111

9245plusmn061

e14

3031

5plusmn8629fg

1096

4plusmn15

9e17

65283plusmn25650

e

Ch2RR

-Vc

643plusmn022

fg7

4306plusmn072

de48

3739plusmn067

e42

109

8688plusmn14

5e13

55038plusmn2233

8ef

9773plusmn437

d15

73485plusmn70367

d

Ch2GM-Vc

838plusmn027

h8

5668plusmn14

9f53

3963plusmn10

3ef

37107

10469plusmn277

f16

2852

4plusmn42886

g118

93plusmn347

f246

8807plusmn55855

f

Ch3RR

-Cr

163plusmn004

a5

1596plusmn040

a52

1196plusmn026

a39

108

2955plusmn07a

464591plusmn

11055

a2018plusmn021

a324924plusmn3381a

Ch3GM-C

r597plusmn014

ef8

3992plusmn066

d55

2524plusmn039

c35

106

7113plusmn116c

1104602plusmn1774

3cd

7679plusmn052

c12

36362plusmn8379c

Ch3RR

-Vc

323plusmn018

b6

2675plusmn111bc

541915plusmn18

8b39

107

4912plusmn269

b768946plusmn42380

b6218plusmn219

b10

01040plusmn35293

b

Ch3GM-Vc

708plusmn070

g8

4753plusmn354

e54

3193plusmn283

d37

107

8653plusmn70

2de

1345094plusmn108385e

f8977plusmn257

c14

4531

2plusmn41422

c

Redrip

ening(


)andverm

icom

postsupp

lement(Vc

)no


HC)

capsaicin


(DHC)

Scoville

heatun

its=SH

UCon

tent

ofn-DHC

was

calculated

asequivalentso

fDHC

Means

inthes

amec

olum

nfollo

wed

bydifferent

lette

rs(andashh

)weresignificantly

different

byTu

keyrsquos

testat119875le005N

D=no

tdetectedlowastCa

psaicincontentcon

verted

toSH

Uusingequatio

nfro

m[21]


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


20 40 60 80 100 120 140minus25

minus20

minus15

minus10

minus5

0

5

10

15

20


HPL

C-SP

T

Meanminus37

minus218

143

minus196 SD

+196 SD

(a)

20 40 60 80 100 120 14020

40

60

80

100

120

140


Mea

n of

HPL

C an

d H

PLC1

(b)










0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


0 20 40 60 80 100 120 140

Spec

troph

otom

eter

20

40

60

80

100

120


tota

l cap

saic

inoi

ds (m

gg d

wt)



4 Conclusion




Acknowledgments



References
































































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of




















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of





















Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of










Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of

Documents

Research Article Measurement of Capsaicinoids in Chiltepin Hot …downloads.hindawi.com/journals/jchem/2015/709150.pdf · AlbertoGonzález-Zamora, 1,2 ErickSierra-Campos, 3 RebecaPérez-Morales,