Jurnal AGRINIKA.
September-2021.
: 122-128
Antioxidant Activity of Non-Volatile Lime (Citrus aurantifolia Swingl.
Extract Nandang Permadi1.
Euis Julaeha2.
Yudi Rosandi3.
Mohamad Nurzaman1* Department of Biology.
Faculty of Mathematics and Natural Sciences.
Universitas Padjadjaran.
Sumedang 45363.
Indonesia Department of Chemistry.
Faculty of Mathematics and Natural Sciences.
Universitas Padjadjaran.
Sumedang 45363.
Indonesia Department of Geophysics.
Faculty of Mathematics and Natural Sciences.
Universitas Padjadjaran.
Sumedang 45363.
Indonesia *Correspondence: m.
nurzaman@unpad.
Diterima 24 Juli 2021/ Direvisi 07 September 2021/ Disetujui 20 September 2021
ABSTRACT
The paper reports the antioxidant activity of a non-volatile fraction of lime processing byproducts from the lime syrup home industry.
The activity was measured by spectrophotometry to obtain the 50% inhibition concentration (IC .
using the 2,2diphenyl-1-picrylhydrazyl (DPPH) method.
The sample was extracted by the maceration method with n-hexane, ethyl-acetate, and ethyl-ethanol.
The IC50 values of 681 ppm, 458 ppm, and 2,775 ppm were n-hexane, ethyl-acetate, and ethyl-ethanol.
The experiments concluded that the ethyl-acetate extract provides the best inhibition value to scavenge free-radicals DPPH.
The HPLC and mass spectroscopy were measured to find out the content and group of active compounds.
The significant compounds consisted of naringin, poncirin, or neoponcirin, which are known as antioxidant substances.
The result shows the potential application of the lime by-products, its volatile fraction, and the nonvolatile fraction, which is the production residue of lime peel.
This work can be applied as an alternative to zero-waste lime production, which may benefit the industry and the Keywords: Antioxidant.
Citrus aurantifolia.
DPPH.
Zero-waste industry INTRODUCTION Citrus aurantifolia, a member of the Rutaceae family, has been commonly used for food and health In the conventional industry, the main product is obtained from lime The peel is considered as waste and usually trashed without further Some research reports that the peel is rich in substances that can be extracted further to obtain various active compounds for food, medicine, and health applications (Loizzo et al.
Zhuo Zou et al.
, 2.
The biological function of lime peel is originated from the phytochemical Among the biological activities of the peel are antioxidant, antiinflammation, anti-bacterial, anti-viral anti-fungi, anti-diabetic substances (Safdar et al.
, 2017.
Samira Lagha-Benamrouche & Madania, 2013.
Shofinita.
Feng, & Langrish, 2.
Citrus fruits are the primary source of ascorbate acid, containing many bioactive substances, such as coumarin, carotenoid, limonoid, and flavonoid, especially polymethoxy-flavone and Many species of citrus own a Nandang Permadi.
Antioxidant Activity of Non-VolatileA broad spectrum of biological activities.
According to the literature, the plant can also be applied for traditional medical treatment of digestion system disorder.
It is also reported that the polyphenols substances can obtain potent freeradical scavenging (Loizzo et al.
, 2.
due to the ability to act as hydrogen, electron, and chelate transition metals In this work, the antioxidant activity of the Citrus aurantifolia extract was The antioxidant substance has been used for food products additive, which can protect from product quality decrease through the oxidativedegradation.
Although the synthetic antioxidant is commonly used, the natural substance is preferable to avoid side effects, such as liver function disturbance and carcinogenic diseases (Chen et al.
, 2.
Hence, the isolation antioxidant substances are demanding.
The solvent extraction method, which is commonly used in preparing plant extract, was used in this work.
The method is widely applied, efficient, and relatively easy to operate.
The most used solvents are hot water, methanol, ethanol, acetone, and ethyl-acetate (Chen et al.
, 2017.
Samira LaghaBenamrouche & Madania, 2.
contrast, solid-liquid maceration is the most used conventional extraction method to obtain bioactive compounds of plants.
This paper reports the result of the non-volatile extraction process of the C.
aurantifolia, after separating volatile fraction .
ssential oi.
by hydrodistillation technique.
The residue was further extracted by the maceration method using n-hexane, ethyl-acetate, and ethanol solvents, respectively.
The antioxidant activity of each produced extract was evaluated using the 2,2diphenyl-1-picrylhydrazyl (DPPH) The polyphenols profile of the most active extract was studied using HPLC Mass-spectroscopy MATERIALS AND METHODS Materials and characterization The C.
aurantifolia peel was obtained from the residue of the lime home industry in Kuningan.
West Java.
The laboratory standard Sthal distillation equipment was used to extract the volatile fraction.
HPLC apparatus with RF-18 column and photodiode array detector (PDA) was used to analyze the chemical content of the sample.
The mass spectroscopy apparatus uses TOP MS ES .
The used organic solvents are n-hexane, ethyl-acetate, and ethanol, obtained from the local chemical supply.
Preparation The approximately 2 cm wide, weighed, and put in the distillation flask, soaked in distilled water.
Then the distillation process is operated for 3 hours to remove the volatile fraction.
After this process, the residue of citrus peels was taken and blended with n-hexane The first maceration procedure for three days.
The same procedure is performed sequentially using different ethyl-acetate ethanol, as the second and third maceration processes.
The extracts are collected daily, and the same type of solvent is refilled to the flask.
The resulted extracts from this three-stage Page 123 of 128 Nandang Permadi.
Antioxidant Activity of Non-VolatileA procedure are collected for (Maemulyati & A.
Prihadi, 2016.
Safdar et al.
, 2017.
Samira LaghaBenamrouche & Madania, 2.
The antioxidant activity was tested using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method, a very reactive freeradical chromogen.
The substance absorbed visible light at 517 nm, detected by spectroscopy apparatus.
The concentration of the tested substance is varied according to the ability to scavenge DPPH free radicals, which is shown by the color change from purple to yellow.
The method is widely used due to its easiness, measurement speed, and sensitivity.
The amount of 0.
1 mL n-hexane, ethyl-acetate, and ethanol extracts with around 100 to 4,000 ppm were added in 2 mL DPPH .
21 mM in 95% ethano.
The mix was shaken and kept for 60 minutes in a dark room and ambient A spectrometer measured the light absorbance at 517 nm.
The DPPH inhibition, %I, is calculated by .
Antioxidant activity test where Ac and As is the absorbance control and sample, respectively.
The IC50 value is measure by, .
a and b were taken as the line fitting function parameter from the plotted measurement data of %I for each extract.
RESULTS AND DISCUSSION
Extraction yield The yield of the extraction process can be seen in Table 1.
Using the maceration process was obtained 0.
2% yield from the total peel weight, and it strongly depends on the used Table 1.
The yield of extracts.
The initial peel weight is 400 g Solvent Extract weight .
n-hexane ethyl-acetate 80% ethanol The extraction sequence is arranged in such a way depending on the solvent polarity, n-hexane as a non-polar, ethylacetate as a semi-polar, and ethanol as a polar solvent.
The ethanol gave an enormous yield value due to its polarity, which indicates that polar components dominate the crude extract.
Antioxidant test The inhibition value was evaluated using the DPPH method, which %Yield measured the ability of the substance to scavenge free radicals.
The DPPH captures antioxidant substances through reaction, which changes the substance The reaction is accompanied by the decrease of light absorption at 517 nm wavelength.
The decrease is due to the disappearance of induced by the electron dynamics of the free radicals.
Hence, the decrease is Page 124 of 128 Nandang Permadi.
Antioxidant Activity of Non-VolatileA proportional to the number of free radicals stabilized by free radicals.
The phenomenon is observed using the In order to quantize the activities, the IC50 is defined, which stands for the amount of concentration of the substance that can inhibit 50% of oxidation reaction by DPPH.
Figure 1 .
n percentag.
as a function of substance Figure 1.
The correlation between the concentration and the inhibition percentage for n-hexane, ethyl-acetate, and ethanol extract of lime peel.
The red horizontal line is the 50% inhibition value (IC.
The black cross symbols denote the values from line regression approximation.
The dashed lines are to guide the eye The plot shows a positive correlation between the concentration of the n-Hexane extract with the inhibition The demonstrates the effectiveness of the inhibition mechanism induced by the number of polyphenols contained in the The result agrees with the data previously reported in (Samira LaghaBenamrouche & Madania, 2.
The obtained IC50 number extracted by nhexane, ethyl-acetate, and 80% ethanol 2 ppm, 457.
6 ppm, dan 2,775.
ppm, respectively.
The result shows that the ethyl-acetate gives the lowest value.
Hence, the extract is the most active compared to other extracts.
The results of HPLC and spectroscopy measurement The most active extract .
is to be analysed further by HPLC and mass spectroscopy.
Figure 2
shows the chromatogram profile taken
HPLC
Page 125 of 128 Nandang Permadi.
Antioxidant Activity of Non-VolatileA Figure 2.
The chromatogram of ethyl-acetate extract of the lime peel with RP-18 PDA Figures 3 and 4 show the mass spectrum obtained from the experiment.
The Beynon table shows two dominant masses, 594 amu and 580 amu.
C28H34O14
C27H32O14
Figure 3.
The mass spectrum of the main chemical component of the ethyl-acetate .
mass spectrum of the C28H34O14 molecule.
Page 126 of 128 Nandang Permadi.
Antioxidant Activity of Non-VolatileA Figure 4.
The mass spectrum of the main chemical component of the ethyl-acetate extract indicates the C27H32O14 molecule.
According to the phytochemistry tracing of the citrus family, this molecular formulae leads to the poncirin or neoponcirin, and naringin compounds.
These compounds are members of the flavonoid group, which are the effective antioxidants (Chen et , 2017.
Safdar et al.
, 2017.
Xia.
Fanga.
Zhaob.
Jiaob, & Zhoua, 2.
CONCLUSION
From this research, an ethylacetate extract is the most active antioxidant extract, having 457.
6 ppm IC50.
Compared to the value of n-hexane and ethanol extract, this value is Hence this paper concludes that ethyl-acetate is the preferable solvent to extract C.
aurantifolia peels.
The main components for antioxidant activity, poncirin or neoponcirin and naringin, are identified in the resulted The result confirms that the lime peel has the potential to be used as an antioxidant product.
ACKNOWLEDGEMENT
The Ministry of Research and Technology/National.
Research and Innovation Agency, supports this work, which has provided research funding through the Master Thesis Research Grant scheme in 2020, contract number:
1827/UN6.
1/LT/2020.
REFERENCES