Research article · 2025 · Full text
International Journal of Agriculture and NutritionDetermination of capsaicin content in organically and conventionally grown chili fruits
Volume 7(8) · Pages 108–112 · DOI: 10.33545/26646064.2025.v7.i8b.276 ↗
Research areasAgricultural Practices and Plant Genetics
Abstract
Author-writtenCapsicum annuum L. (chili) is widely consumed for its pungency and is commonly used as a condiment, while also boasting numerous pharmaceutical and medicinal properties. Capsaicin, an odorless alkaloid with an intense, often irritating flavor, is the primary bioactive compound in chili, accompanied by other capsaicinoids like dihydrocapsaicin and nordihydrocapsaicin, which offer high nutritional value. Chromatographic techniques were employed to identify and quantify a broad range of capsaicin and related compounds in four chili fruit samples of Kharpuri variety from both conventional and organic cultivation. Using the HPTLC method, capsaicin content was found to be significantly higher in organic chili fruits grown in Rajasthan, India (0.3635 mg/g), compared to conventionally grown fruits (0.1368 mg/g). The difference in capsaicin levels highlights the impact of cultivation methods, with conventional farming relying on chemical fertilizers and pesticides, while organic farming uses eco-friendly fertilizers and biopesticides. This comparative study underscores the nutritional and bioactive advantages of organic chili fruits grown under natural conditions.
Summary
Source-checkedPlain-language reading of the full paper, prepared with AI and checked against the text. Each point links to the authors' own words.
Why it matters
- Chili (Capsicum annuum L.) is a widely used condiment valued for its pungency and various medicinal and pharmaceutical properties. View evidence ↗
- The primary bioactive compound responsible for the intense flavor and pungency of chili is the odorless alkaloid capsaicin. View evidence ↗
- Capsaicin is noted for its chemopreventive properties, which can reduce the malignant growth of several types of cancer, including lung, breast, and prostate cancers. View evidence ↗
The question
- The study aimed to use High-Performance Thin-Layer Chromatography (HPTLC) to isolate, identify, and quantify capsaicin in chili samples. View evidence ↗
What they did
- The researchers used Kharpuri variety chili fruits collected from fields using either conventional or organic farming practices. View evidence ↗
- Samples were divided into four groups: one conventional (Con) and three organic (Org I, II, and III), with the organic groups representing different durations of chemical-free farming (2-3, 4-5, and 8-9 years). View evidence ↗
- Capsaicin was extracted using a Soxhlet apparatus at 60 °C for 24 hours, utilizing 4 grams of dried chili powder and 300 mL of ethanol. View evidence ↗
- The separated bands were measured using HPTLC densitometric scanning at a wavelength of 282 nm. View evidence ↗
What they found
- The capsaicin standard showed a strong linear relationship in the 1-5 µg/spot range, with R² values of 0.985 for band height and 0.995 for peak area. View evidence ↗
- The retention factor (Rf) values for the samples (0.515 to 0.527) were close to the standard capsaicin Rf of 0.517, confirming the identity of the compound. View evidence ↗
- Organic I chili fruits had the highest capsaicin concentration at 0.3635 mg/g, whereas conventional fruits had the lowest at 0.1368 mg/g. View evidence ↗
- All organic samples (Org I: 0.3635 µg/g, Org II: 0.2672 µg/g, Org III: 0.3135 µg/g) contained higher capsaicin levels than the conventional sample (0.1368 µg/g). View evidence ↗
What it means
- The authors concluded that organic cultivation of the Kharpuri chili variety results in significantly higher capsaicin content compared to conventional cultivation. View evidence ↗
- The authors state that HPTLC is an efficient, quick, and easy method for quantifying capsaicin in both organic and conventional chili fruits. View evidence ↗
Visual summary
2 visualsIllustrations explain the paper; charts are replotted from the paper's own table values.
Experimental Design Overview
AI-generated conceptual illustration (Kharpuri variety chili fruits). It explains the idea and does not show study data.
View source evidence ↗
Content µg/gram dry weight: Conventional chili fruit vs Organic I chili fruit vs Organic II chili fruit vs Organic III chili fruit
Replotted from Table 1 (page 3) of the original paper. Every bar is the paper's own table value.
View source evidence ↗Figures & tables
From the original paper
Figure 1
Table 1
Figure 1
Figure 2
Figure 3
Figure 4Introduction
Capsaicin plays a crucial role in determining the pungency and commercial quality of chili fruits globally. Pungency, the sensory perception of heat in chili, is primarily due to alkaloids known as capsaicinoids. The two predominant capsaicinoids, (i) capsaicin (8-methyl-Nvanillyl-trans-6-nonenamide) and (ii) dihydrocapsaicin (N[(4-Hydroxy-3-methoxyphenyl) methyl]-8-methylnonanamide), account for approximately 80-90% of total capsaicinoid content, with capsaicin alone constituting around 71% in typically pungent varieties (Barbero et al., 2014; Kumar et al., 2024) [2,14] . Capsaicin is not only a key factor in chili pungency but also exhibits significant pharmacological, neurological, and dietary benefits (Gurnani et al., 2016) [13] , including anti-cancer properties (Anandakumar et al., 2013; Dawan et al., 2017) [6,11] . Research has demonstrated that capsaicin has chemopreventive properties, effectively reducing the malignant growth of cancers such as lung, colorectal, gastric, pancreatic, breast, and prostate (Pramanik and Srivastava, 2013) [13] . It has also been shown to shrink tumors by inhibiting cancer cell progression (Szallasi, 2023) [16] . Capsaicin was found to be predominantly distributed in the placenta of the chili fruit, compared to the pericarp and seed (Taira et al., 2012) [15] . The amount of capsaicin in a specific variety can differ, based on the light intensity, temperature, and the age of the fruit. With growing consumer demand for vegetables and fruits rich in nutritional content and quality, the development of superior pepper varieties through crop improvement programs has become increasingly important. Understanding the variation in bioactive compounds, such as capsaicin, across different pepper germplasms is essential for this process. This study aimed to quantify capsaicin, a key determinant of pungency and a compound with significant medicinal properties, in chili samples using High-Performance Thin-Layer Chromatography (HPTLC) for its isolation and identification. Capsaicin, known for its intense pungency, is colorless and highly soluble in oils, fats, ethanol, methanol, and ethyl alkali (Ekwere & Udon, 2016) [12] . Previous studies have shown that ethanol extracts yield a high concentration of capsaicin. HPTLC, an advanced form of Thin-Layer Chromatography, is a robust analytical technique that operates on the principles of adsorption and partition, depending on the absorbent materials used on the plates and the solvent system for the mobile phase (Bairy, 2015) [9] . To assess pungency, the Scoville test method and advanced instrumental technologies were employed to determine capsaicin content in the samples.
Material and Methods
Chili fruits of Kharpuri variety were collected from different fields employing conventional and organic farming practices. Fields using chemical fertilizers and pesticides represented conventional farming, while organic fields were free of chemical fertilizers and pesticides. The fruits were thoroughly washed with tap water to remove dust and airdried in the shade. The chili samples were categorized into four groups: Con (conventional farming), Org I, Org II, and Org III. The organic samples were collected from fields practicing chemical-free farming for varying durations: Org I (2-3 years), Org II (4-5 years), and Org III (8-9 years).
• Extraction Method: Capsaicin extraction and quantification followed the method of Collins et al.
(1995) [10] with minor modifications. Dried chili powder was mixed with solvents in a 1:10 ratio (g) and extracted at 60 °C for 24 hours using a Soxhlet apparatus. For the process, 4 grams of chili powder and 300 mL of ethanol were used. The extraction continued until the tissue was decolorized (approximately 16 hours). • Standard preparation: 10 mg standard capsaicin was dissolved in 10 mL of Methanol. Further, 1 ml of the prepared solution was diluted with 10 mL of methanol to get 100 µg/mL solution. This solution was applied on a TLC plate. • Capsaicin identification: Capsaicin peak identification in the extract was done by matching the retention time with that of standard capsaicin, which was further confirmed by co-injecting standard capsaicin together with the extract for quantitative analysis of capsaicin in chosen samples of Capsicum fruits in this study.
• Instrumentation and chromatographic conditions:
Using a Linomat 5 automated TLC applicator, five microliters of ethanolic extract (500 mg/mL) were applied to pre-coated TLC silica gel plates. A standard capsaicin solution (100 µg/mL) was used as a reference.
The TLC plates were pre-washed with methanol, dried, and developed using a mobile phase in a CAMAG HPTLC chamber. After plate saturation and equilibrium, the plates were developed to 85 mm thickness and dried. HPTLC densitometric scanning at 282 nm was performed to measure the separated bands.
Comparative densitometric analysis of peak height and area allowed quantitative analysis of the extracts.
Observations and results
Capsaicin was extracted from chili fruits grown under conventional (Con) and organic (Org I, Org II, and Org III) cultivation methods (Figure 1), followed by densitometric HPTLC analysis. Capsaicin identification was confirmed by comparing band heights of the extracts with external standards, using retention factor (Rf) values for validation. Linear regression analysis of the capsaicin standard demonstrated a strong linear relationship, with a regression coefficient (R²) of 0.985 for band height and 0.995 for peak area, within the concentration range of 1-5 µg/spot (Figure 2; A). Capsaicin isolated from conventional and organic (Org I, Org II, Org III) chili samples was analyzed by comparing their Rf values with a standard capsaicin reference (Figure 2;B). The recorded Rf values were 0.527 for Org I, 0.523 for Org II, 0.515 for Org III, and 0.523 for the conventional sample, compared to the standard capsaicin Rf of 0.517. The mobile phase of methanol provided optimal resolution for capsaicin, with an Rf value around 0.52, enabling simultaneous quantification. The peak area for standard capsaicin was 0.00379, while the peak areas for Org I, Org II, Org III, and conventional samples were 0.00644, 0.00537, 0.00589, and 0.00392, respectively (Figure 3). Capsaicin content was quantified using the HPTLC method, revealing that organic chili fruit samples contained higher amounts of capsaicin compared to conventional samples. The highest capsaicin concentration was observed in organic I chili fruits (0.3635 mg/g), while the lowest was found in conventional chili fruits (0.1368 mg/g). The detailed results are presented in Table 1 and Figure 4.
| S. No. | Sample | Rf | Maximum Height | Area | Content µg/gram dry weight |
|---|---|---|---|---|---|
| 1 | Standard capsaicin | 0.517 | 0.216 | 0.00379 | 14.545 |
| 2 | Conventional chili fruit | 0.523 | 0.119 | 0.00392 | 0.1368 |
| 3 | Organic I chili fruit | 0.527 | 0.180 | 0.00644 | 0.3635 |
| 4 | Organic II chili fruit | 0.523 | 0.153 | 0.00537 | 0.2672 |
| 5 | Organic III chili fruit | 0.515 | 0.148 | 0.00589 | 0.3135 |
Table text extracted from the original PDF; superscript letters are shown inline. The original table image is below.
View original table image
Discussion
The concentration of capsaicin and dihydrocapsaicin is a major determinant of pepper pungency (Sweat et al.,
. These two capsaicinoids are responsible for the majority of the heat sensation in chili peppers, with capsaicin typically contributing the most-were analysed using GCMS (Choudhary et al., 2022) [4] . The growing system (organic farming), ripening stage, and genotype are key factors that contribute to the variation in capsaicinoid content in chili fruits (Guijarro-Real et al., 2023) [1] . Bicikliski et al., 2017 [8] observed capsaicin levels in peppers grown organically tended to have higher capsaicin levels compared to those grown conventionally, across all six genotypes in Republic of Macedonia. However, the difference in capsaicin content between the two cultivation methods was not statistically significant, indicating that while organic farming may impact capsaicin levels, the variation is not enough to clearly distinguish between organic and conventional methods. A mesocosm experiment found that using a combination of organic and mineral fertilizers (OMF) significantly increased capsaicin content in Capsicum chinense cv Trinidad Scorpion compared to mineral fertilizer alone using HPLC. Capsaicin levels were five times higher with the OMF treatment, highlighting the effectiveness of this combined fertilization approach (Pampuro et al., 2017) [7] .
Conclusion
Quantification of capsaicin and associated capsaicinoids using HPTLC to compare organic and conventional fruits is first of its kind. The HPTLC proved to be a quick and easy approach for the analysis and quantification of capsaicin in conventional and organically grown chili fruits. Organic samples of Kharpuri variety of chili showed much higher capsaicin in comparison to conventionally chili fruits of same variety.
Declarations
• Ethics approval and consent to participate: Not
Acknowledgement
AcknowledgementsWe are highly grateful to Krishi Vigyan Kendra Jhunjhunu to provide insights and identification of diseases. We acknowledge Department of Botany, University of Rajasthan to provide facilities to carry out research.
References
16 in the original paper- Guijarro-Real C, Adalid-Martínez AM, Pires CK, Ribes-Moya AM, Fita A, Rodríguez-Burruezo A. The effect of the varietal type, ripening stage, and growing conditions on the content and profile of sugars and capsaicinoids in Capsicum peppers. Plants. 2023;12(2):231.
- Barbero GF, Ruiz AG, Liazid A, Palma M, Vera JC, Barroso CG. Evolution of total and individual capsaicinoids in peppers during ripening of the Cayenne pepper plant (Capsicum annuum L.). Food Chem. 2014;153:200-206.
- Pramanik KC, Srivastava SK. Role of capsaicin in cancer prevention. Role of Capsaicin in Oxidative Stress and Cancer. 2013;1-18.
- Choudhary K, Mishra P, Dayanand M, Patni V. Influence of organic farming on volatile compounds in methanolic fruit extracts of chili (Capsicum annuum L.). J Microbiol Biotechnol Food Sci. 2022;12(3):e3545-e3545.
- Sweat KG, Broatch J, Borror C, Hagan K, Cahill TM. Variability in capsaicinoid content and Scoville heat ratings of commercially grown Jalapeño, Habanero and Bhut Jolokia peppers. Food Chem. 2016;210:606-612.
- Anandakumar P, Kamaraj S, Jagan S, Ramakrishnan G, Devaki T. Capsaicin provokes apoptosis and restricts benzo(a)pyrene induced lung tumorigenesis in Swiss albino mice. Int Immunopharmacol. 2013;17(2):254259.
- Pampuro N, Tebaldo V, Fabbri D, Calza P, Faga MG, Cavallo E. Effect of organic fertilization on capsaicin content in Trinidad Scorpion (Capsicum chinense) peppers: preliminary results. Chem Eng Trans. 2017;58:253-258.
- Bicikliski O, Tasev K, Trajkova F, Mihajlov L, Koleva Gudeva L. Comparative analysis of capsaicin content in peppers (Capsicum annuum L.) grown in conventional and organic agricultural systems. J Agric Plant Sci. 2017;15(1/2):27-35.
- Bairy PS. A comparison study of HPLC and HPTLC: principles, instrumentations and applications. ASIO J Anal Chem. 2015;1(1):20-28.
- Collins MD, Wasmund LM, Bosland PW. Improved method for quantifying capsaicinoids in Capsicum using high-performance liquid chromatography. 1995;137-139.
- Dawan P, Satarpai T, Tuchinda P, Shiowatana J, Siripinyanond A. A simple analytical platform based on thin-layer chromatography coupled with paper-based analytical device for determination of total capsaicinoids in chili samples. Talanta. 2017;162:460465.
- Ekwere MR, Udoh DE. Extraction and comparative analysis of moisture and capsaicin contents of capsicum peppers. J Pain Relief. 2016;5(268):2167-0846.
- Gurnani N, Gupta M, Mehta D, Mehta BK. Chemical composition, total phenolic and flavonoid contents, and in vitro antimicrobial and antioxidant activities of crude extracts from red chili seeds (Capsicum frutescens L.). J Taibah Univ Sci. 2016;10(4):462-470.
- Kumar S, Sarpras M, Mushtaq F, Singh S, Thattantavide A, Kumar A. Capsaicin-producing plants: species diversity, distribution and botanical details. In: Capsaicinoids: From Natural Sources to Biosynthesis and their Clinical Applications. Singapore: Springer Nature Singapore; 2024. p. 1-24.
- Taira S, Shimma S, Osaka I, Kaneko D, Ichiyanagi Y, Ikeda R, et al. Mass spectrometry imaging of the capsaicin localization in the Capsicum fruits. Int J Biotechnol. 2012.
- Szallasi A. Capsaicin and cancer: guilty as charged or innocent until proven guilty?. Temperature. 2023;10(1):35-49.