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2. Materials and methods2.1. Materi
2. Materials and methods
2.1. Materials and reagents
Fresh red peppers (red ripe fruits of capsicum annuum L) were
purchased in September 25, 2012 from the local vegetable e market
located in Hunan Agricultural University. They were packaged in
polyethylene bags and stored in a refrigerator (4 C) prior to
extraction. Each 100 g pericarp (after removal of seeds and stalk) of
FRP contained 24.7 mg capsaicinoids and 439.2 mg carotenoids.
Capsaicin (98%, Sigma company, America). b-carotene (90%
(HPLC), Chengdu Mansite company, China). Ethanol, n-hexane,
acetone, ethyl acetate were of analytical grade and purchased from
local chemical company (Changsha, China). Re-distilled water was
used for the preparation of all solutions and dilutions.
2.2. Stage extraction process of capsaicinoids and red pigments
from FRPs
2.2.1. The total procedures
The pretreatment of FRPs was carried out as follows. The sarcocarp
was obtained by weeding out the stalk and seeds from FRPs.
Then the sarcocarp of peppers was cut up into small pieces of
2 2 mm approximately.
For optimizing the conditions of stage extraction, 10 g of FRPs
sarcocarp were put into a 100 mL glass flask and were immersed in
extractive solvent for 30 min. Then they were extracted by heat
refluxing extraction. The procedures of stage extraction were
shown in Fig. 1.
2.2.2. Optimizing extraction process
Effect of some extractive conditions on yield of capsaicinoids
and red colorants was firstly investigated by single-factor experiments,
respectively. These conditions, including the ratio of solvent
to material (2, 3, 4, 5, 6 mL/g), the temperature of water bath (50,
60, 70, 80, 90 C) and time of extraction (60, 90, 120, 150, 180 min),
were same at two different stages: the removal stage of capsaicinoids
or extraction stage of red pigments. But, the concentration
of ethanol aqueous was different at above two stages. For
removal of capsaicinoids, the concentration of ethanol was set as
10%, 20%, 30%, 40%, 50%, 60%, 70% (v/v). Meanwhile, it was set as
60%, 70%, 80%, 90%, 95% (v/v) for the stage of extraction of red
pigments.
Then, orthogonal experimental designs were used to optimize
the extractive conditions at both of extraction stages, respectively.
Four different parameters of extraction and three levels for each
were studied by the orthogonal array designs of L9(34) and nine
combinations of extraction conditions with different parameters
were established at both of extraction stages (Table 1).
Every extractive solution was collected and evaporated to dry at
40 C under reduced pressure with a rotary evaporator, respectively.
The residues obtained were dissolved with 20 mL solventand centrifuged for 10 min at 3000 rpm. Each solution was
collected and diluted with homologous solvent to a constant volume
for analysis. The solvent used was methanol for capsaicinoids
analysis, while acetone for red pigments analysis. HPLC analysis
was used for capsaicinoids, while colorimetric analysis for red
pigments.
2.3. Stage extraction of capsaicinoids and red pigments,
respectively, from FRPs
2.3.1. Effect of extraction times on the yield of capsaicinoids and red
pigments
Ten grams of pepper sarcocarp with 40 mL of 50% ethanol
aqueous were refluxed at 90 C of water bath in 2 h. Then the
extracted solution was collected and the residues were again
extracted with the same conditions until the capsaicinoids were
exhausted. Each solution was used to analyze the content of capsaicinoids
by HPLC.
Then red pigments were further extracted from the residues
after extracting capsaicinoids from 10 g starting material with
40 mL of 95% ethanol for 2 h at 90 C of water bath. The extracted
solution was collected and the residues were again extracted with
the same conditions until the red pigments were exhausted. Each
solution was collected, respectively, and used to determine the
absorbance by spectrophotometric analysis.
2.3.2. Thrice replicates for extraction of capsaicinoids and red
pigments
Thrice replicates were carried out as same as the Section 2.3.1,
but the extraction times were twice for each stage.
2.4. Extraction process of capsaicinoids and red pigments from
DRPs
The dried red peppers (DRPs) were obtained by desiccating FRPs
(whole pods) under solar light (the temperature about 20e28 C,
8 h 10 days) or under 50 C (for 8 h 3 days) in an electrothermal
drying oven to constant weight. Then the pericarp were collected
and crushed to a homogeneous size in a mill, sieved through a
No.40 mesh.
The simulative industrial extraction process of red pigmentswas
carried out mainly according to the references (Bae, Jayaprakasha,
Jifon, & Patil, 2012; Sang, Jiang, & Yu, 2008). The detail methods
were as follows. An amount of 0.9 g powders of DRPs (equivalent to
10 g of FRPs) was loaded into a Soxhlet extractor and then was
extracted using hexane 150 mL for 6 h. The extracted solution was
collected and evaporated to dry under reduced pressure with a
rotary evaporator at 35 C. A oleoresin containing red pigments and
pungent substance was obtained.
2.5. Analysis
2.5.1. HPLC analysis of capsaicinoids
The samples of capsaicinoids were analyzed by highperformance
liquid chromatography (HPLC) on Agilent 1200 HPLC
system composed of a quaternary pump with a degasser, a thermostatted
column compartment, a detector of a UV-detector (a
variable wavelength detector, VWD) and an autosampler and 1200
ChemStation software. The column used was an Agilent Eclipse
XDB-C18 column (250 mm 4.6 mm I.D., 5 mm), the mobile phase
was methanolewater (65:35, v/v) with flowrate at 0.8 mL/min. The
wavelength of UV detector was set at 280 nm and column temperaturewas
set at 35 C. All samples were filtered through 0.45 mm
membrane before HPLC analysis and the injection volume of sample
was 20 mL.Preparation of standard solution: An amount of 2.7 mg of
capsaicinwas accurately weighed and dissolved in 10 mL methanol.
The standard stock solution was diluted with methanol to appropriate
concentrations for the construction of calibration curves.
2.5.2. Quantification of red pigments
Test solution of pigment samples was determined by spectrophotometry
(National standards of People’s Republic of China,
2008; Tundis et al., 2012). The details were as follows: the
amount of 0.1 (0.0001) g samples was dissolved and diluted into
100 mL with acetone as a stock solution. Then the stock solution
was diluted into an appropriate concentration with acetone and the
absorbance of the diluted solutionwas measured against acetone at
460 nm by a spectrophotometer (Shimadzu UV1600). The content
of red pigments was expressed as the total content of carotenoids
using b-carotene equivalents in mg per g pericarp.
Standard solutionwas prepared as follows: An amount of 5.6 mg
of b-carotene was accurately weighed and dissolved in 50 mL
acetone. The standard stock solution was diluted with acetone to
appropriate concentrations for the construction of calibration
curves.
For determining the content of red colorant in FRP, the pericarp
after removal of seed and stalk were extracted with acetone until
the color was exhausted. The combined extracts were evaporated
to remove acetone and water (from FRP) under reduced pressure
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
2.1. Materials and reagents
Fresh red peppers (red ripe fruits of capsicum annuum L) were
purchased in September 25, 2012 from the local vegetable e market
located in Hunan Agricultural University. They were packaged in
polyethylene bags and stored in a refrigerator (4 C) prior to
extraction. Each 100 g pericarp (after removal of seeds and stalk) of
FRP contained 24.7 mg capsaicinoids and 439.2 mg carotenoids.
Capsaicin (98%, Sigma company, America). b-carotene (90%
(HPLC), Chengdu Mansite company, China). Ethanol, n-hexane,
acetone, ethyl acetate were of analytical grade and purchased from
local chemical company (Changsha, China). Re-distilled water was
used for the preparation of all solutions and dilutions.
2.2. Stage extraction process of capsaicinoids and red pigments
from FRPs
2.2.1. The total procedures
The pretreatment of FRPs was carried out as follows. The sarcocarp
was obtained by weeding out the stalk and seeds from FRPs.
Then the sarcocarp of peppers was cut up into small pieces of
2 2 mm approximately.
For optimizing the conditions of stage extraction, 10 g of FRPs
sarcocarp were put into a 100 mL glass flask and were immersed in
extractive solvent for 30 min. Then they were extracted by heat
refluxing extraction. The procedures of stage extraction were
shown in Fig. 1.
2.2.2. Optimizing extraction process
Effect of some extractive conditions on yield of capsaicinoids
and red colorants was firstly investigated by single-factor experiments,
respectively. These conditions, including the ratio of solvent
to material (2, 3, 4, 5, 6 mL/g), the temperature of water bath (50,
60, 70, 80, 90 C) and time of extraction (60, 90, 120, 150, 180 min),
were same at two different stages: the removal stage of capsaicinoids
or extraction stage of red pigments. But, the concentration
of ethanol aqueous was different at above two stages. For
removal of capsaicinoids, the concentration of ethanol was set as
10%, 20%, 30%, 40%, 50%, 60%, 70% (v/v). Meanwhile, it was set as
60%, 70%, 80%, 90%, 95% (v/v) for the stage of extraction of red
pigments.
Then, orthogonal experimental designs were used to optimize
the extractive conditions at both of extraction stages, respectively.
Four different parameters of extraction and three levels for each
were studied by the orthogonal array designs of L9(34) and nine
combinations of extraction conditions with different parameters
were established at both of extraction stages (Table 1).
Every extractive solution was collected and evaporated to dry at
40 C under reduced pressure with a rotary evaporator, respectively.
The residues obtained were dissolved with 20 mL solventand centrifuged for 10 min at 3000 rpm. Each solution was
collected and diluted with homologous solvent to a constant volume
for analysis. The solvent used was methanol for capsaicinoids
analysis, while acetone for red pigments analysis. HPLC analysis
was used for capsaicinoids, while colorimetric analysis for red
pigments.
2.3. Stage extraction of capsaicinoids and red pigments,
respectively, from FRPs
2.3.1. Effect of extraction times on the yield of capsaicinoids and red
pigments
Ten grams of pepper sarcocarp with 40 mL of 50% ethanol
aqueous were refluxed at 90 C of water bath in 2 h. Then the
extracted solution was collected and the residues were again
extracted with the same conditions until the capsaicinoids were
exhausted. Each solution was used to analyze the content of capsaicinoids
by HPLC.
Then red pigments were further extracted from the residues
after extracting capsaicinoids from 10 g starting material with
40 mL of 95% ethanol for 2 h at 90 C of water bath. The extracted
solution was collected and the residues were again extracted with
the same conditions until the red pigments were exhausted. Each
solution was collected, respectively, and used to determine the
absorbance by spectrophotometric analysis.
2.3.2. Thrice replicates for extraction of capsaicinoids and red
pigments
Thrice replicates were carried out as same as the Section 2.3.1,
but the extraction times were twice for each stage.
2.4. Extraction process of capsaicinoids and red pigments from
DRPs
The dried red peppers (DRPs) were obtained by desiccating FRPs
(whole pods) under solar light (the temperature about 20e28 C,
8 h 10 days) or under 50 C (for 8 h 3 days) in an electrothermal
drying oven to constant weight. Then the pericarp were collected
and crushed to a homogeneous size in a mill, sieved through a
No.40 mesh.
The simulative industrial extraction process of red pigmentswas
carried out mainly according to the references (Bae, Jayaprakasha,
Jifon, & Patil, 2012; Sang, Jiang, & Yu, 2008). The detail methods
were as follows. An amount of 0.9 g powders of DRPs (equivalent to
10 g of FRPs) was loaded into a Soxhlet extractor and then was
extracted using hexane 150 mL for 6 h. The extracted solution was
collected and evaporated to dry under reduced pressure with a
rotary evaporator at 35 C. A oleoresin containing red pigments and
pungent substance was obtained.
2.5. Analysis
2.5.1. HPLC analysis of capsaicinoids
The samples of capsaicinoids were analyzed by highperformance
liquid chromatography (HPLC) on Agilent 1200 HPLC
system composed of a quaternary pump with a degasser, a thermostatted
column compartment, a detector of a UV-detector (a
variable wavelength detector, VWD) and an autosampler and 1200
ChemStation software. The column used was an Agilent Eclipse
XDB-C18 column (250 mm 4.6 mm I.D., 5 mm), the mobile phase
was methanolewater (65:35, v/v) with flowrate at 0.8 mL/min. The
wavelength of UV detector was set at 280 nm and column temperaturewas
set at 35 C. All samples were filtered through 0.45 mm
membrane before HPLC analysis and the injection volume of sample
was 20 mL.Preparation of standard solution: An amount of 2.7 mg of
capsaicinwas accurately weighed and dissolved in 10 mL methanol.
The standard stock solution was diluted with methanol to appropriate
concentrations for the construction of calibration curves.
2.5.2. Quantification of red pigments
Test solution of pigment samples was determined by spectrophotometry
(National standards of People’s Republic of China,
2008; Tundis et al., 2012). The details were as follows: the
amount of 0.1 (0.0001) g samples was dissolved and diluted into
100 mL with acetone as a stock solution. Then the stock solution
was diluted into an appropriate concentration with acetone and the
absorbance of the diluted solutionwas measured against acetone at
460 nm by a spectrophotometer (Shimadzu UV1600). The content
of red pigments was expressed as the total content of carotenoids
using b-carotene equivalents in mg per g pericarp.
Standard solutionwas prepared as follows: An amount of 5.6 mg
of b-carotene was accurately weighed and dissolved in 50 mL
acetone. The standard stock solution was diluted with acetone to
appropriate concentrations for the construction of calibration
curves.
For determining the content of red colorant in FRP, the pericarp
after removal of seed and stalk were extracted with acetone until
the color was exhausted. The combined extracts were evaporated
to remove acetone and water (from FRP) under reduced pressure
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
0/5000
2. Materials and methods2.1. Materials and reagentsFresh red peppers (red ripe fruits of capsicum annuum L) werepurchased in September 25, 2012 from the local vegetable e marketlocated in Hunan Agricultural University. They were packaged inpolyethylene bags and stored in a refrigerator (4 C) prior toextraction. Each 100 g pericarp (after removal of seeds and stalk) ofFRP contained 24.7 mg capsaicinoids and 439.2 mg carotenoids.Capsaicin (98%, Sigma company, America). b-carotene (90%(HPLC), Chengdu Mansite company, China). Ethanol, n-hexane,acetone, ethyl acetate were of analytical grade and purchased fromlocal chemical company (Changsha, China). Re-distilled water wasused for the preparation of all solutions and dilutions.2.2. Stage extraction process of capsaicinoids and red pigmentsfrom FRPs2.2.1. The total proceduresThe pretreatment of FRPs was carried out as follows. The sarcocarpwas obtained by weeding out the stalk and seeds from FRPs.Then the sarcocarp of peppers was cut up into small pieces of2 2 mm approximately.For optimizing the conditions of stage extraction, 10 g of FRPssarcocarp were put into a 100 mL glass flask and were immersed inextractive solvent for 30 min. Then they were extracted by heatrefluxing extraction. The procedures of stage extraction wereshown in Fig. 1.2.2.2. Optimizing extraction processEffect of some extractive conditions on yield of capsaicinoidsand red colorants was firstly investigated by single-factor experiments,respectively. These conditions, including the ratio of solventto material (2, 3, 4, 5, 6 mL/g), the temperature of water bath (50,60, 70, 80, 90 C) and time of extraction (60, 90, 120, 150, 180 min),were same at two different stages: the removal stage of capsaicinoidsor extraction stage of red pigments. But, the concentrationof ethanol aqueous was different at above two stages. Forremoval of capsaicinoids, the concentration of ethanol was set as10%, 20%, 30%, 40%, 50%, 60%, 70% (v/v). Meanwhile, it was set as60%, 70%, 80%, 90%, 95% (v/v) for the stage of extraction of redpigments.Then, orthogonal experimental designs were used to optimizethe extractive conditions at both of extraction stages, respectively.Four different parameters of extraction and three levels for eachwere studied by the orthogonal array designs of L9(34) and ninecombinations of extraction conditions with different parameterswere established at both of extraction stages (Table 1).Every extractive solution was collected and evaporated to dry at40 C under reduced pressure with a rotary evaporator, respectively.The residues obtained were dissolved with 20 mL solventand centrifuged for 10 min at 3000 rpm. Each solution wascollected and diluted with homologous solvent to a constant volumefor analysis. The solvent used was methanol for capsaicinoidsanalysis, while acetone for red pigments analysis. HPLC analysiswas used for capsaicinoids, while colorimetric analysis for redpigments.2.3. Stage extraction of capsaicinoids and red pigments,respectively, from FRPs2.3.1. Effect of extraction times on the yield of capsaicinoids and redpigmentsTen grams of pepper sarcocarp with 40 mL of 50% ethanolaqueous were refluxed at 90 C of water bath in 2 h. Then theextracted solution was collected and the residues were againextracted with the same conditions until the capsaicinoids wereexhausted. Each solution was used to analyze the content of capsaicinoidsby HPLC.Then red pigments were further extracted from the residuesafter extracting capsaicinoids from 10 g starting material with40 mL of 95% ethanol for 2 h at 90 C of water bath. The extractedsolution was collected and the residues were again extracted withthe same conditions until the red pigments were exhausted. Eachsolution was collected, respectively, and used to determine theabsorbance by spectrophotometric analysis.2.3.2. Thrice replicates for extraction of capsaicinoids and redpigmentsThrice replicates were carried out as same as the Section 2.3.1,but the extraction times were twice for each stage.2.4. Extraction process of capsaicinoids and red pigments fromDRPsThe dried red peppers (DRPs) were obtained by desiccating FRPs(whole pods) under solar light (the temperature about 20e28 C,8 h 10 days) or under 50 C (for 8 h 3 days) in an electrothermaldrying oven to constant weight. Then the pericarp were collectedand crushed to a homogeneous size in a mill, sieved through aNo.40 mesh.The simulative industrial extraction process of red pigmentswascarried out mainly according to the references (Bae, Jayaprakasha,Jifon, & Patil, 2012; Sang, Jiang, & Yu, 2008). The detail methodswere as follows. An amount of 0.9 g powders of DRPs (equivalent to10 g of FRPs) was loaded into a Soxhlet extractor and then wasextracted using hexane 150 mL for 6 h. The extracted solution wascollected and evaporated to dry under reduced pressure with arotary evaporator at 35 C. A oleoresin containing red pigments andpungent substance was obtained.2.5. Analysis2.5.1. HPLC analysis of capsaicinoidsThe samples of capsaicinoids were analyzed by highperformanceliquid chromatography (HPLC) on Agilent 1200 HPLCsystem composed of a quaternary pump with a degasser, a thermostattedcolumn compartment, a detector of a UV-detector (avariable wavelength detector, VWD) and an autosampler and 1200ChemStation software. The column used was an Agilent EclipseXDB-C18 column (250 mm 4.6 mm I.D., 5 mm), the mobile phasewas methanolewater (65:35, v/v) with flowrate at 0.8 mL/min. Thewavelength of UV detector was set at 280 nm and column temperaturewasset at 35 C. All samples were filtered through 0.45 mmmembrane before HPLC analysis and the injection volume of samplewas 20 mL.Preparation of standard solution: An amount of 2.7 mg ofcapsaicinwas accurately weighed and dissolved in 10 mL methanol.The standard stock solution was diluted with methanol to appropriateconcentrations for the construction of calibration curves.2.5.2. Quantification of red pigmentsTest solution of pigment samples was determined by spectrophotometry(National standards of People’s Republic of China,2008; Tundis et al., 2012). The details were as follows: theamount of 0.1 (0.0001) g samples was dissolved and diluted into100 mL with acetone as a stock solution. Then the stock solutionwas diluted into an appropriate concentration with acetone and theabsorbance of the diluted solutionwas measured against acetone at460 nm by a spectrophotometer (Shimadzu UV1600). The contentof red pigments was expressed as the total content of carotenoidsusing b-carotene equivalents in mg per g pericarp.Standard solutionwas prepared as follows: An amount of 5.6 mgof b-carotene was accurately weighed and dissolved in 50 mLacetone. The standard stock solution was diluted with acetone toappropriate concentrations for the construction of calibrationcurves.For determining the content of red colorant in FRP, the pericarpafter removal of seed and stalk were extracted with acetone untilthe color was exhausted. The combined extracts were evaporatedto remove acetone and water (from FRP) under reduced pressure
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
การแปล กรุณารอสักครู่..
2. Materials and methods
2.1. Materials and reagents
Fresh red peppers (red ripe fruits of capsicum annuum L) were
purchased in September 25, 2012 from the local vegetable e market
located in Hunan Agricultural University. They were packaged in
polyethylene bags and stored in a refrigerator (4 C) prior to
extraction. Each 100 g pericarp (after removal of seeds and stalk) of
FRP contained 24.7 mg capsaicinoids and 439.2 mg carotenoids.
Capsaicin (98%, Sigma company, America). b-carotene (90%
(HPLC), Chengdu Mansite company, China). Ethanol, n-hexane,
acetone, ethyl acetate were of analytical grade and purchased from
local chemical company (Changsha, China). Re-distilled water was
used for the preparation of all solutions and dilutions.
2.2. Stage extraction process of capsaicinoids and red pigments
from FRPs
2.2.1. The total procedures
The pretreatment of FRPs was carried out as follows. The sarcocarp
was obtained by weeding out the stalk and seeds from FRPs.
Then the sarcocarp of peppers was cut up into small pieces of
2 2 mm approximately.
For optimizing the conditions of stage extraction, 10 g of FRPs
sarcocarp were put into a 100 mL glass flask and were immersed in
extractive solvent for 30 min. Then they were extracted by heat
refluxing extraction. The procedures of stage extraction were
shown in Fig. 1.
2.2.2. Optimizing extraction process
Effect of some extractive conditions on yield of capsaicinoids
and red colorants was firstly investigated by single-factor experiments,
respectively. These conditions, including the ratio of solvent
to material (2, 3, 4, 5, 6 mL/g), the temperature of water bath (50,
60, 70, 80, 90 C) and time of extraction (60, 90, 120, 150, 180 min),
were same at two different stages: the removal stage of capsaicinoids
or extraction stage of red pigments. But, the concentration
of ethanol aqueous was different at above two stages. For
removal of capsaicinoids, the concentration of ethanol was set as
10%, 20%, 30%, 40%, 50%, 60%, 70% (v/v). Meanwhile, it was set as
60%, 70%, 80%, 90%, 95% (v/v) for the stage of extraction of red
pigments.
Then, orthogonal experimental designs were used to optimize
the extractive conditions at both of extraction stages, respectively.
Four different parameters of extraction and three levels for each
were studied by the orthogonal array designs of L9(34) and nine
combinations of extraction conditions with different parameters
were established at both of extraction stages (Table 1).
Every extractive solution was collected and evaporated to dry at
40 C under reduced pressure with a rotary evaporator, respectively.
The residues obtained were dissolved with 20 mL solventand centrifuged for 10 min at 3000 rpm. Each solution was
collected and diluted with homologous solvent to a constant volume
for analysis. The solvent used was methanol for capsaicinoids
analysis, while acetone for red pigments analysis. HPLC analysis
was used for capsaicinoids, while colorimetric analysis for red
pigments.
2.3. Stage extraction of capsaicinoids and red pigments,
respectively, from FRPs
2.3.1. Effect of extraction times on the yield of capsaicinoids and red
pigments
Ten grams of pepper sarcocarp with 40 mL of 50% ethanol
aqueous were refluxed at 90 C of water bath in 2 h. Then the
extracted solution was collected and the residues were again
extracted with the same conditions until the capsaicinoids were
exhausted. Each solution was used to analyze the content of capsaicinoids
by HPLC.
Then red pigments were further extracted from the residues
after extracting capsaicinoids from 10 g starting material with
40 mL of 95% ethanol for 2 h at 90 C of water bath. The extracted
solution was collected and the residues were again extracted with
the same conditions until the red pigments were exhausted. Each
solution was collected, respectively, and used to determine the
absorbance by spectrophotometric analysis.
2.3.2. Thrice replicates for extraction of capsaicinoids and red
pigments
Thrice replicates were carried out as same as the Section 2.3.1,
but the extraction times were twice for each stage.
2.4. Extraction process of capsaicinoids and red pigments from
DRPs
The dried red peppers (DRPs) were obtained by desiccating FRPs
(whole pods) under solar light (the temperature about 20e28 C,
8 h 10 days) or under 50 C (for 8 h 3 days) in an electrothermal
drying oven to constant weight. Then the pericarp were collected
and crushed to a homogeneous size in a mill, sieved through a
No.40 mesh.
The simulative industrial extraction process of red pigmentswas
carried out mainly according to the references (Bae, Jayaprakasha,
Jifon, & Patil, 2012; Sang, Jiang, & Yu, 2008). The detail methods
were as follows. An amount of 0.9 g powders of DRPs (equivalent to
10 g of FRPs) was loaded into a Soxhlet extractor and then was
extracted using hexane 150 mL for 6 h. The extracted solution was
collected and evaporated to dry under reduced pressure with a
rotary evaporator at 35 C. A oleoresin containing red pigments and
pungent substance was obtained.
2.5. Analysis
2.5.1. HPLC analysis of capsaicinoids
The samples of capsaicinoids were analyzed by highperformance
liquid chromatography (HPLC) on Agilent 1200 HPLC
system composed of a quaternary pump with a degasser, a thermostatted
column compartment, a detector of a UV-detector (a
variable wavelength detector, VWD) and an autosampler and 1200
ChemStation software. The column used was an Agilent Eclipse
XDB-C18 column (250 mm 4.6 mm I.D., 5 mm), the mobile phase
was methanolewater (65:35, v/v) with flowrate at 0.8 mL/min. The
wavelength of UV detector was set at 280 nm and column temperaturewas
set at 35 C. All samples were filtered through 0.45 mm
membrane before HPLC analysis and the injection volume of sample
was 20 mL.Preparation of standard solution: An amount of 2.7 mg of
capsaicinwas accurately weighed and dissolved in 10 mL methanol.
The standard stock solution was diluted with methanol to appropriate
concentrations for the construction of calibration curves.
2.5.2. Quantification of red pigments
Test solution of pigment samples was determined by spectrophotometry
(National standards of People’s Republic of China,
2008; Tundis et al., 2012). The details were as follows: the
amount of 0.1 (0.0001) g samples was dissolved and diluted into
100 mL with acetone as a stock solution. Then the stock solution
was diluted into an appropriate concentration with acetone and the
absorbance of the diluted solutionwas measured against acetone at
460 nm by a spectrophotometer (Shimadzu UV1600). The content
of red pigments was expressed as the total content of carotenoids
using b-carotene equivalents in mg per g pericarp.
Standard solutionwas prepared as follows: An amount of 5.6 mg
of b-carotene was accurately weighed and dissolved in 50 mL
acetone. The standard stock solution was diluted with acetone to
appropriate concentrations for the construction of calibration
curves.
For determining the content of red colorant in FRP, the pericarp
after removal of seed and stalk were extracted with acetone until
the color was exhausted. The combined extracts were evaporated
to remove acetone and water (from FRP) under reduced pressure
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
2.1. Materials and reagents
Fresh red peppers (red ripe fruits of capsicum annuum L) were
purchased in September 25, 2012 from the local vegetable e market
located in Hunan Agricultural University. They were packaged in
polyethylene bags and stored in a refrigerator (4 C) prior to
extraction. Each 100 g pericarp (after removal of seeds and stalk) of
FRP contained 24.7 mg capsaicinoids and 439.2 mg carotenoids.
Capsaicin (98%, Sigma company, America). b-carotene (90%
(HPLC), Chengdu Mansite company, China). Ethanol, n-hexane,
acetone, ethyl acetate were of analytical grade and purchased from
local chemical company (Changsha, China). Re-distilled water was
used for the preparation of all solutions and dilutions.
2.2. Stage extraction process of capsaicinoids and red pigments
from FRPs
2.2.1. The total procedures
The pretreatment of FRPs was carried out as follows. The sarcocarp
was obtained by weeding out the stalk and seeds from FRPs.
Then the sarcocarp of peppers was cut up into small pieces of
2 2 mm approximately.
For optimizing the conditions of stage extraction, 10 g of FRPs
sarcocarp were put into a 100 mL glass flask and were immersed in
extractive solvent for 30 min. Then they were extracted by heat
refluxing extraction. The procedures of stage extraction were
shown in Fig. 1.
2.2.2. Optimizing extraction process
Effect of some extractive conditions on yield of capsaicinoids
and red colorants was firstly investigated by single-factor experiments,
respectively. These conditions, including the ratio of solvent
to material (2, 3, 4, 5, 6 mL/g), the temperature of water bath (50,
60, 70, 80, 90 C) and time of extraction (60, 90, 120, 150, 180 min),
were same at two different stages: the removal stage of capsaicinoids
or extraction stage of red pigments. But, the concentration
of ethanol aqueous was different at above two stages. For
removal of capsaicinoids, the concentration of ethanol was set as
10%, 20%, 30%, 40%, 50%, 60%, 70% (v/v). Meanwhile, it was set as
60%, 70%, 80%, 90%, 95% (v/v) for the stage of extraction of red
pigments.
Then, orthogonal experimental designs were used to optimize
the extractive conditions at both of extraction stages, respectively.
Four different parameters of extraction and three levels for each
were studied by the orthogonal array designs of L9(34) and nine
combinations of extraction conditions with different parameters
were established at both of extraction stages (Table 1).
Every extractive solution was collected and evaporated to dry at
40 C under reduced pressure with a rotary evaporator, respectively.
The residues obtained were dissolved with 20 mL solventand centrifuged for 10 min at 3000 rpm. Each solution was
collected and diluted with homologous solvent to a constant volume
for analysis. The solvent used was methanol for capsaicinoids
analysis, while acetone for red pigments analysis. HPLC analysis
was used for capsaicinoids, while colorimetric analysis for red
pigments.
2.3. Stage extraction of capsaicinoids and red pigments,
respectively, from FRPs
2.3.1. Effect of extraction times on the yield of capsaicinoids and red
pigments
Ten grams of pepper sarcocarp with 40 mL of 50% ethanol
aqueous were refluxed at 90 C of water bath in 2 h. Then the
extracted solution was collected and the residues were again
extracted with the same conditions until the capsaicinoids were
exhausted. Each solution was used to analyze the content of capsaicinoids
by HPLC.
Then red pigments were further extracted from the residues
after extracting capsaicinoids from 10 g starting material with
40 mL of 95% ethanol for 2 h at 90 C of water bath. The extracted
solution was collected and the residues were again extracted with
the same conditions until the red pigments were exhausted. Each
solution was collected, respectively, and used to determine the
absorbance by spectrophotometric analysis.
2.3.2. Thrice replicates for extraction of capsaicinoids and red
pigments
Thrice replicates were carried out as same as the Section 2.3.1,
but the extraction times were twice for each stage.
2.4. Extraction process of capsaicinoids and red pigments from
DRPs
The dried red peppers (DRPs) were obtained by desiccating FRPs
(whole pods) under solar light (the temperature about 20e28 C,
8 h 10 days) or under 50 C (for 8 h 3 days) in an electrothermal
drying oven to constant weight. Then the pericarp were collected
and crushed to a homogeneous size in a mill, sieved through a
No.40 mesh.
The simulative industrial extraction process of red pigmentswas
carried out mainly according to the references (Bae, Jayaprakasha,
Jifon, & Patil, 2012; Sang, Jiang, & Yu, 2008). The detail methods
were as follows. An amount of 0.9 g powders of DRPs (equivalent to
10 g of FRPs) was loaded into a Soxhlet extractor and then was
extracted using hexane 150 mL for 6 h. The extracted solution was
collected and evaporated to dry under reduced pressure with a
rotary evaporator at 35 C. A oleoresin containing red pigments and
pungent substance was obtained.
2.5. Analysis
2.5.1. HPLC analysis of capsaicinoids
The samples of capsaicinoids were analyzed by highperformance
liquid chromatography (HPLC) on Agilent 1200 HPLC
system composed of a quaternary pump with a degasser, a thermostatted
column compartment, a detector of a UV-detector (a
variable wavelength detector, VWD) and an autosampler and 1200
ChemStation software. The column used was an Agilent Eclipse
XDB-C18 column (250 mm 4.6 mm I.D., 5 mm), the mobile phase
was methanolewater (65:35, v/v) with flowrate at 0.8 mL/min. The
wavelength of UV detector was set at 280 nm and column temperaturewas
set at 35 C. All samples were filtered through 0.45 mm
membrane before HPLC analysis and the injection volume of sample
was 20 mL.Preparation of standard solution: An amount of 2.7 mg of
capsaicinwas accurately weighed and dissolved in 10 mL methanol.
The standard stock solution was diluted with methanol to appropriate
concentrations for the construction of calibration curves.
2.5.2. Quantification of red pigments
Test solution of pigment samples was determined by spectrophotometry
(National standards of People’s Republic of China,
2008; Tundis et al., 2012). The details were as follows: the
amount of 0.1 (0.0001) g samples was dissolved and diluted into
100 mL with acetone as a stock solution. Then the stock solution
was diluted into an appropriate concentration with acetone and the
absorbance of the diluted solutionwas measured against acetone at
460 nm by a spectrophotometer (Shimadzu UV1600). The content
of red pigments was expressed as the total content of carotenoids
using b-carotene equivalents in mg per g pericarp.
Standard solutionwas prepared as follows: An amount of 5.6 mg
of b-carotene was accurately weighed and dissolved in 50 mL
acetone. The standard stock solution was diluted with acetone to
appropriate concentrations for the construction of calibration
curves.
For determining the content of red colorant in FRP, the pericarp
after removal of seed and stalk were extracted with acetone until
the color was exhausted. The combined extracts were evaporated
to remove acetone and water (from FRP) under reduced pressure
with a rotary evaporator at 45 C. A residual cream was obtained.
Then it was dissolved with acetone into a determinate volume for
absorbance determination.
3. Results and discussion
3.1. Analysis
3.1.1. HPLC analysis of capsaicinoids
The contents of capsaicinoids were analyzed by HPLC to evaluate
the extracted effect. The HPLC separation conditions including
the ratio of mobile phase (methanolewater) and flow rate were
firstly investigated and the satisfactory chromatographic conditions
were summarized in Section 2.5.1. The HPLC chromatograms
of standard compound (capsaicin) and the crude extract were
shown in Fig. 2, respectively.
การแปล กรุณารอสักครู่..
2 . วัสดุและวิธีการ
2.1 . วัสดุและสารเคมี
พริกแดงสด ( ผลไม้สีแดงสุกพริก annuum L )
ซื้อในวันที่ 25 กันยายน 2012 จาก ตลาดผักพื้นบ้าน E
ตั้งอยู่ในมหาวิทยาลัยการเกษตรมณฑลหูหนาน . พวกเขาถูกบรรจุในถุงโพลีเอทธิลีน
และเก็บไว้ในตู้เย็น ( 4 C ) ก่อน
การสกัด แต่ละ 100 กรัม ( หลังจากที่เอาเปลือกของเมล็ดและลำต้น )
FRP ประกอบด้วย 24และ 7 มิลลิกรัมแคปไซซินอยด์ 439.2 มก. แคโรทีน .
แคปไซซิน ( 98% , บริษัท , Sigma อเมริกา ) เบต้าแคโรทีน ( 90 %
( HPLC ) บริษัท mansite เฉิงตู , จีน ) เอทานอล , บีบ
, อะซิโตน , เอทิลอะซิเตท และวิเคราะห์เกรดและซื้อจาก
เคมี บริษัทท้องถิ่น ( ฉางชา , จีน ) เรื่องน้ำกลั่น
ใช้สำหรับการเตรียมการของโซลูชั่นทั้งหมดและเจือจาง .
2.2 .ขั้นตอนของกระบวนการสกัดแคปไซซินอยด์และสีแดง
จาก frps 2.2.1 . ขั้นตอนทั้งหมด
frps ได้ดําเนินการดังนี้ การ sarcocarp
โดยนำวัชพืชออกก้านและเมล็ดจาก frps .
แล้ว sarcocarp ของพริกก็หั่นเป็นชิ้นเล็ก ๆประมาณ 2 mm
2 .
สำหรับปรับสภาพของการสกัดขั้น frps
10 กรัมsarcocarp ถูกใส่ลงไปในแก้วขวด 100 มล. และแช่ในตัวทำละลายการสกัด
เป็นเวลา 30 นาที จากนั้นนำไปสกัดด้วยความร้อน
กรดสกัด ขั้นตอนของการสกัดระยะที่แสดงในรูปที่ 1
.
2.2.2 . การเพิ่มประสิทธิภาพกระบวนการและผลกระทบของเงื่อนไข
การสกัดการสกัดผลผลิตของแคปไซซินอยด์
สีแดงและเป็นครั้งแรก โดยปัจจัยเดียว (
)
2.1 . วัสดุและสารเคมี
พริกแดงสด ( ผลไม้สีแดงสุกพริก annuum L )
ซื้อในวันที่ 25 กันยายน 2012 จาก ตลาดผักพื้นบ้าน E
ตั้งอยู่ในมหาวิทยาลัยการเกษตรมณฑลหูหนาน . พวกเขาถูกบรรจุในถุงโพลีเอทธิลีน
และเก็บไว้ในตู้เย็น ( 4 C ) ก่อน
การสกัด แต่ละ 100 กรัม ( หลังจากที่เอาเปลือกของเมล็ดและลำต้น )
FRP ประกอบด้วย 24และ 7 มิลลิกรัมแคปไซซินอยด์ 439.2 มก. แคโรทีน .
แคปไซซิน ( 98% , บริษัท , Sigma อเมริกา ) เบต้าแคโรทีน ( 90 %
( HPLC ) บริษัท mansite เฉิงตู , จีน ) เอทานอล , บีบ
, อะซิโตน , เอทิลอะซิเตท และวิเคราะห์เกรดและซื้อจาก
เคมี บริษัทท้องถิ่น ( ฉางชา , จีน ) เรื่องน้ำกลั่น
ใช้สำหรับการเตรียมการของโซลูชั่นทั้งหมดและเจือจาง .
2.2 .ขั้นตอนของกระบวนการสกัดแคปไซซินอยด์และสีแดง
จาก frps 2.2.1 . ขั้นตอนทั้งหมด
frps ได้ดําเนินการดังนี้ การ sarcocarp
โดยนำวัชพืชออกก้านและเมล็ดจาก frps .
แล้ว sarcocarp ของพริกก็หั่นเป็นชิ้นเล็ก ๆประมาณ 2 mm
2 .
สำหรับปรับสภาพของการสกัดขั้น frps
10 กรัมsarcocarp ถูกใส่ลงไปในแก้วขวด 100 มล. และแช่ในตัวทำละลายการสกัด
เป็นเวลา 30 นาที จากนั้นนำไปสกัดด้วยความร้อน
กรดสกัด ขั้นตอนของการสกัดระยะที่แสดงในรูปที่ 1
.
2.2.2 . การเพิ่มประสิทธิภาพกระบวนการและผลกระทบของเงื่อนไข
การสกัดการสกัดผลผลิตของแคปไซซินอยด์
สีแดงและเป็นครั้งแรก โดยปัจจัยเดียว (
)
การแปล กรุณารอสักครู่..
ภาษาอื่น ๆ
การสนับสนุนเครื่องมือแปลภาษา: กรีก, กันนาดา, กาลิเชียน, คลิงออน, คอร์สิกา, คาซัค, คาตาลัน, คินยารวันดา, คีร์กิซ, คุชราต, จอร์เจีย, จีน, จีนดั้งเดิม, ชวา, ชิเชวา, ซามัว, ซีบัวโน, ซุนดา, ซูลู, ญี่ปุ่น, ดัตช์, ตรวจหาภาษา, ตุรกี, ทมิฬ, ทาจิก, ทาทาร์, นอร์เวย์, บอสเนีย, บัลแกเรีย, บาสก์, ปัญจาป, ฝรั่งเศส, พาชตู, ฟริเชียน, ฟินแลนด์, ฟิลิปปินส์, ภาษาอินโดนีเซี, มองโกเลีย, มัลทีส, มาซีโดเนีย, มาราฐี, มาลากาซี, มาลายาลัม, มาเลย์, ม้ง, ยิดดิช, ยูเครน, รัสเซีย, ละติน, ลักเซมเบิร์ก, ลัตเวีย, ลาว, ลิทัวเนีย, สวาฮิลี, สวีเดน, สิงหล, สินธี, สเปน, สโลวัก, สโลวีเนีย, อังกฤษ, อัมฮาริก, อาร์เซอร์ไบจัน, อาร์เมเนีย, อาหรับ, อิกโบ, อิตาลี, อุยกูร์, อุสเบกิสถาน, อูรดู, ฮังการี, ฮัวซา, ฮาวาย, ฮินดี, ฮีบรู, เกลิกสกอต, เกาหลี, เขมร, เคิร์ด, เช็ก, เซอร์เบียน, เซโซโท, เดนมาร์ก, เตลูกู, เติร์กเมน, เนปาล, เบงกอล, เบลารุส, เปอร์เซีย, เมารี, เมียนมา (พม่า), เยอรมัน, เวลส์, เวียดนาม, เอสเปอแรนโต, เอสโทเนีย, เฮติครีโอล, แอฟริกา, แอลเบเนีย, โคซา, โครเอเชีย, โชนา, โซมาลี, โปรตุเกส, โปแลนด์, โยรูบา, โรมาเนีย, โอเดีย (โอริยา), ไทย, ไอซ์แลนด์, ไอร์แลนด์, การแปลภาษา.
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