CN110308135A - Raman-enhanced rapid detection of capsaicin in capsicum based on gold-silver alloy nano-surface with empty shell - Google Patents

Abstract

The invention discloses a method for rapid detection of capsaicin in capsicum based on nano-surface Raman enhancement of empty shell gold-silver alloy, comprising: step 1, weighing fresh red and dried capsicum to obtain a crude capsaicin extract; step 2, using ethanol to extract capsaicin from capsicum Extracting capsaicin from the crude extract to obtain a capsaicin extract; step 3, taking the capsaicin extract, adding acetonitrile to prepare liquids to be tested with different concentrations; step 4, preparing an empty shell gold-silver alloy nanosol substrate; step 5. After the empty shell gold-silver alloy nanosol substrate, the liquid to be tested and the aqueous sodium chloride solution are mixed uniformly, Raman spectrum collection is performed; Step 6, the content of capsaicin in the dried chili pepper is calculated according to the following formula. The invention applies the surface Raman enhancement technology to the detection of capsaicin in dried chili peppers for the first time, and has high detection accuracy, short time and simple operation.

Description

Raman-enhanced Raman-enhanced Rapid Detection of Capsicum in Capsicum Based on Gold-Silver Alloy Nano-Surface prime method

technical field

The invention relates to the technical field of plant detection, in particular to a method for rapid detection of capsaicin in capsicum based on an empty shell gold-silver alloy nano-surface Raman enhancement.

Background technique

The following background art is provided to assist the reader in understanding the present invention and should not be regarded as prior art.

Capsaicin ((trans)8-methyl-N-vanillyl-6-nonenamide) is the active ingredient in capsicum. It is irritating to mammals including humans and can cause a burning sensation in the mouth. Capsaicin and some related compounds, also known as capsaicin, are secondary metabolites produced by peppers, possibly as a deterrent to herbivores. Birds are generally not sensitive to capsicum elements. Pure capsaicin is a water-repellent, lipophilic, colorless, odorless, crystalline or waxy compound. Capsaicin has several effects: 1. Capsaicin also inhibits the activity of a gene that causes arteries to constrict. Arterial constriction reduces blood flow to the heart and other organs. By inhibiting the activity of this gene, the muscles can relax and expand, which in turn increases blood volume. 2. Studies have found that capsaicin can improve a number of factors related to heart and blood vessel health. However, it is not recommended that people consume chili in excess. A healthy diet needs to be balanced, and people must be aware that chili peppers are not a substitute for clinically tested and proven prescription drugs. 3. The research team found in animal experiments that capsaicin, the main ingredient in peppers, can promote muscle growth and inhibit muscle atrophy. The researchers believe that this discovery may be helpful for the development of drugs for the treatment of muscle atrophy, and it is necessary to confirm the safety of using capsaicin.

Raman spectrum is a kind of scattering spectrum. The Raman spectrum analysis method is based on the Raman scattering effect discovered by Indian scientist C.V. Raman (Raman). Analysis is performed to obtain information on molecular vibrations, rotations. Surface-enhanced Raman spectroscopy (SERS) technology refers to the adsorption of molecules into the surface or sol of some nano-rough metals (such as gold, silver, and copper), so that the Raman signal of the adsorbed species is geometrically multiple. enhanced. SERS technology can realize the rapid detection of trace samples and single molecules. At the same time, it is also a highly sensitive fingerprint spectrum. It has the characteristics of easy portability, simple operation, and convenient pretreatment. At present, there is no report on the rapid detection of capsaicin in pepper by surface-enhanced Raman spectroscopy.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide a method for rapid detection of capsaicin in capsicum based on the Raman enhancement of the nano-surface of empty shell gold-silver alloy.

The technical solution adopted by the present invention to solve the technical problem is: a method for rapid detection of capsaicin in pepper based on the Raman enhancement on the nano-surface of empty shell gold-silver alloy, including:

Step 1. Weigh fresh red dried peppers to obtain capsaicin crude extract;

Step 2, using ethanol to extract capsaicin from the capsaicin crude extract to obtain a capsaicin extract;

Step 3, take the capsaicin extract, add acetonitrile, and prepare different concentrations of the test solution;

Step 4, preparing an empty shell gold-silver alloy nano-sol substrate;

Step 5. After the empty shell gold-silver alloy nano-sol substrate, the liquid to be tested and the sodium chloride aqueous solution are evenly mixed, Raman spectrum collection is performed;

Step 6, calculate the content of capsaicin in the dried peppers according to the following formula:

y=137.55x+9094.4,

In the formula: y is the content of capsaicin in the dried pepper, the unit is mg/L, where L is the volume of the liquid to be tested, and mg is the content of chlorpyrifos pesticide; x is the intensity of the Raman absorption peak at 1065cm ^-1 , and the unit is au.

Preferably, in step (2), the method for obtaining the capsaicin extract comprises: weighing 5g of the capsaicin crude extract, placing it in a 50mL centrifuge tube, adding 60mL ethanol into the centrifuge tube, ultrasonically vibrating for 2 minutes, at 4500 rpm Centrifuge for 10 min at a high speed, extract the supernatant, and then filter through a 0.45 μm organic filter to obtain a capsaicin extract.

Preferably, in step (3), 9 concentration gradients of the liquid to be tested are prepared, 0.5, 1, 5, 10, 15, 20, 50, 70 and 100 mg/L respectively, and 3 parallel samples are prepared for each concentration.

Preferably, in step (4), the method for preparing empty shell gold-silver alloy nanosol is: first, in the presence of phytic acid, reducing silver nitrate with trisodium citrate to prepare silver nanoparticles; Take out 10 mL, dilute it to 25 mL with deionized water, add 5 mL of 10 ^-3 mol/L chloroauric acid under constant stirring, and stir and heat at 50 °C for 30 min to prepare an empty shell gold-silver alloy nanosol solution.

Preferably, in step (5), the volumes of the liquid to be tested, the empty shell gold-silver alloy nanosol substrate, and the aqueous sodium chloride solution are 100 μL, 500 μL, and 100 μL, respectively, wherein the mass fraction of the aqueous sodium chloride solution is 1%.

Preferably, the Raman spectrum parameters are set as: excitation wavelength 785nm, power 200mW, scanning range 300-3200cm ^-1 , exposure times 2 times, smoothing coefficient 1, integration time 10s, parallel sample detection 3 times.

Compared with the prior art, the present invention has the following advantages:

1. The surface Raman enhancement technology is applied to the detection of capsaicin in dried pepper for the first time, with high detection accuracy, short time and simple operation.

2. Establish a model of Raman absorption peak intensity and the true value of capsaicin in dried pepper, and quantitatively detect the content of capsaicin in dried pepper.

3. Using ethanol to extract capsaicin in dried peppers, the capsaicin has high color value, high extraction efficiency and low economic cost.

4. The core-shell gold-silver alloy nano-substrate has the best enhancement effect, and can effectively suppress the fluorescence interference, thereby effectively preventing the baseline drift of the Raman spectrum.

Description of drawings

Figure 1 is a schematic diagram of the molecular structure of capsaicin.

Figure 2 shows the spectrum of capsaicin standard SERS at 1007cm ^-1 at different concentrations: (a) 100mg/L, (b) 70mg/L, (c) 50mg/L, (d) 20mg/L, (e) 15mg /L, (f) 10mg/L, (g) 5mg/L, (h) 1mg/L, (i) 0.5mg/L.

Figure 3 is a standard curve of capsaicin concentration in capsicum.

Detailed ways

The structures involved in the present invention or the used technical terms are further described below. These descriptions only illustrate how the present invention is implemented by way of examples, and do not constitute any limitation to the present invention.

Figure 1 is a schematic diagram of the molecular structure of capsaicin. A Raman-enhanced Raman-enhanced method for rapid detection of capsaicin in peppers based on empty shell gold-silver alloy nanosurfaces includes the following steps:

Step 1. Select fresh red dried chili peppers, select secondary or tertiary chili peppers according to the national standard GB-10465-89, the water content is less than or equal to 14%, and dry them in an oven at 50° to constant weight, remove bad pepper impurities, and pulverize the dried chili peppers. Pulverized by machine to obtain dry pepper samples, which were extracted, filtered and distilled to obtain capsaicin crude extract.

Step 2. Weigh 5 g of capsaicin crude extract, place it in a 50 mL centrifuge tube, add 60 mL of ethanol to the centrifuge tube, ultrasonically vibrate for 2 minutes, centrifuge at 4500 rpm for 10 minutes, extract the supernatant, and then use 0.45 μm organic Filter membrane to obtain capsaicin extract.

The color value of capsaicin in the capsaicin extracted by organic solvents such as acetone is the highest, and the color value of capsaicin obtained by non-continuous enzymatic extraction is in the middle, but the content of its by-product-capsaicin is increased by 30% compared with the acetone extraction method. The capsaicin obtained by the extraction method contains more impurities and lower color value, and the capsaicin content is not high. Therefore, ethanol is used for extraction, and the color value of capsaicin is higher, the extraction efficiency is high, and the economic cost is low.

Step 3, take the capsaicin extract, add acetonitrile, prepare the test solution with capsaicin concentration in capsicum of 0.5, 1, 5, 10, 15, 20, 50, 70 and 100 mg/L, and prepare 3 parallel samples for each concentration. sample.

Step 4, prepare empty shell gold-silver alloy nanosol, the specific operation is as follows:

First, in the presence of phytic acid, use trisodium citrate to reduce silver nitrate to prepare silver nanoparticles, take out 10mL of the prepared silver nanoparticles, dilute to 25mL with deionized water, and add 10 ^- Empty-shell gold-silver alloy nanosol can be prepared by adding 5 mL of ³ mol/L chloroauric acid, stirring and heating at 50 °C for 30 min; after the solution is cooled, pour the above appropriate amount of empty-shell gold-silver alloy nanosol solution into In the centrifuge tube, pour off a small amount of supernatant after centrifugation, add an appropriate amount of ultrapure water to the centrifuge tube, and mix with ultrasonic vibration.

Step 5. Add 100 μL of the test solution prepared in step 2, 500 μL of the empty shell gold-silver alloy nanosol substrate prepared in step 3, and 100 μL of 1% sodium chloride aqueous solution into a 2 mL quartz bottle in turn, and mix them evenly. , put it into the matching liquid sample cell to collect surface-enhanced Raman spectrum. The parameters of Raman spectrum acquisition were set as: excitation wavelength 785nm, power 200mW, scanning range 300-3200cm ^-1 , exposure times 2 times, smoothing coefficient 1, integration time 10s, parallel sample detection 3 times.

Step 6, calculate the content of capsaicin in the dried peppers according to the following formula:

y=137.55x+9094.4,

In the formula: y is the content of capsaicin in the dried pepper, the unit is mg/L, where L is the volume of the liquid to be tested, and mg is the content of capsaicin in the dried pepper; x is the intensity of the Raman absorption peak at 1065cm ^-1 , the unit is au.

The process of establishing the formula is as follows:

(1) Prepare the sample to be tested

Use the methods of steps 1, 2 and 3 to obtain the liquid to be tested.

(2) Using the method of step 2 to prepare an empty shell gold-silver alloy nanosol substrate, and using the parameters and methods of step 4 to test to obtain a surface-enhanced Raman spectrum, the surface-enhanced Raman spectrum is shown in Figure 2, and the x-axis in Figure 2 is the wave number, and the y-axis is the intensity of the Raman absorption peak.

(3) Spectral preprocessing

The original surface-enhanced Raman spectrum collected in step (2) is subjected to preprocessing such as smoothing and baseline pulling by using the spectral preprocessing method to remove the influence of noise.

(4) The 1065cm ^-1 spectral peak can be found in Fig. 2 with a strong signal, so the peak intensity of this characteristic peak at 1065 cm ^-1 is selected for the formulation of the standard curve.

(5) The model relationship between the absorption peaks of Raman spectra and the true value of capsaicin in dried pepper leaves was established by using the single-variable linear regression method.

The model relationship is the formula used in step 6, the linear regression diagram of the formula is shown in Figure 3, and the correlation coefficient of the formula R ² =0.9966.

Precision To verify the reliability of this experiment, the test solutions with capsaicin concentrations of 5, 60 and 90 mg/L in capsicum were prepared, and three parallel samples were prepared for each concentration. Table 1 is the accuracy and precision analysis results. It can be concluded from the table that the recovery rate of the method for detecting capsaicin in peppers is 88.21-96.83%, and the relative standard deviation (RSD) is 5.72-8.97%. The experimental results obtained above basically meet the requirements, indicating the precision and accuracy of the method. high degree.

Table 1 Accuracy and Precision Analysis

The invention shown and described herein may be practiced in the absence of any element, limitation, specifically disclosed herein. The terms and expressions employed are to be used as terms of description rather than limitation and it is not intended that the use of these terms and expressions exclude any equivalents of the features shown and described, or parts thereof, and it should be recognized that each Both modifications are possible within the scope of the present invention. It should therefore be understood that while the invention has been specifically disclosed by various embodiments and optional features, modifications and variations of the concepts described herein may be employed by those of ordinary skill in the art and are considered to fall within the scope of It is within the scope of the invention as defined in the appended claims.

The contents of articles, patents, patent applications, and all other documents and electronically available information described or recorded herein are hereby incorporated by reference in their entirety to the same extent as if each individual publication was specifically As if indicated separately for reference. Applicants reserve the right to incorporate into this application any and all materials and information from any such articles, patents, patent applications or other documents.

Claims (7)

1. enhancing the method for capsaicine in quickly detection capsicum based on ghost gold-silver alloy Nanosurface Raman, comprising:

Step 1 weighs scarlet chilli, obtains capsaicine crude extract；

Step 2 extracts capsaicine using ethyl alcohol from capsaicine crude extract, obtains capsaicine extracting solution；

Step 3 takes capsaicine extracting solution, and acetonitrile is added, prepares the prepare liquid of multiple concentration；

Step 4, preparation ghost gold-silver alloy Nano sol substrate；

Step 5, by ghost gold-silver alloy Nano sol substrate, prepare liquid and sodium-chloride water solution after mixing, carry out Raman Spectra collection；

Step 6, the content that capsaicine in chilli is calculated according to following formula:

Y=137.55x+9094.4,

In formula: y is the content of capsaicine in chilli, unit mg/L, wherein L is the volume of prepare liquid, and mg is chlopyrifos agriculture The content of medicine；X is 1065cm^-1Locate Raman absorption peak intensity, unit a.u..

2. as described in claim 1 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (1), the method for obtaining capsaicine crude extract includes: to select scarlet chilli, according to state Mark GB-10465-89 selects second level or three-level capsicum, and water content is less than or equal to 14%, and with 50 ° of baking oven, drying to constant weight, removes bad Green pepper impurity crushes chilli through pulverizer, obtains chilli sample, obtains capsaicine crude extract through extraction, filtering, distillation.

3. as claimed in claim 2 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (2), obtain capsaicine extracting solution method include: to weigh 5g capsaicine crude extract, put It is placed in 50mL centrifuge tube, 60mL ethyl alcohol is added in centrifuge tube, sonic oscillation 2 minutes, be centrifuged with 4500 turns of revolving speed 10min extracts supernatant, then obtains capsaicine extracting solution with 0.45 μm of organic membrane filtration.

4. as claimed in claim 3 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (3), prepare the prepare liquid of 9 concentration gradients, respectively 0.5,1,5,10,15,20, 50,70 and 100mg/L, each concentration prepare 3 parts of parallel samples.

5. as claimed in claim 4 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (4), prepare ghost gold-silver alloy Nano sol method are as follows: first the item existing for phytic acid Under part, Nano silver grain is prepared with trisodium citrate reduction silver nitrate；By the Nano silver grain prepared take out 10mL, spend from Sub- water is diluted to 25mL, is keeping stirring lower addition 10^-3The gold chloride 5mL of mol/L, agitating and heating 30min under the conditions of 50 DEG C, system Standby ghost gold-silver alloy Nano sol solution out.

6. as claimed in claim 5 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (5), prepare liquid, ghost gold-silver alloy Nano sol substrate, sodium-chloride water solution body Product is respectively 100 μ L, 500 μ L, 100 μ L, wherein the mass fraction of sodium-chloride water solution is 1%.

7. as claimed in claim 6 based on capsaicine in the enhancing of ghost gold-silver alloy Nanosurface Raman quickly detection capsicum Method, which is characterized in that in step (5), Raman spectrum parameter setting are as follows: excitation wavelength 785nm, power 200mW, scan model Enclose 300~3200cm^-1, exposure frequency 2 times, smoothing factor 1, time of integration 10s, parallel sample detection 3 times.