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Background
Cannabis, which is a psychoactive plant, refers to the leaves, flowers, stems, and seeds. Cannabis is used globally for its psychoactive properties, with 2.5% of the world’s population consuming it for recreational purposes. But in Indonesia, the plant is classified as a Class 1 narcotic, with a prevalence of use reaching 41.4%. Aceh is one of the largest cannabis-producing regions in Indonesia due to its favorable geographical and climatic conditions. Despite its illegal status, cannabis contains valuable phytocannabinoid compounds and is potentially important in medical applications. Previous studies have shown a correlation between the compound profile of cannabis and its geographical origin. This study aims to develop a classification method based on the cannabinoids compound profiles of dried cannabis samples taken from five regions in Aceh (Aceh Besar, Aceh Tengah, Bireuen, Lhokseumawe, and Pidie Jaya) by microscopy, Raman spectrophotometry, GC–MS, and parametric statistical analysis to assist authorities in tracing the source of cannabis for law enforcement and forensic purposes.
Results
In this study, dried Cannabis sativa from five regions of Aceh, Indonesia, was tested with Raman spectroscopy and GC–MS to produce informative cannabinoid compound profiles as plant profiling. The results obtained 10 cannabinoids quantified in plant samples (Δ9-THC, CBD, THCV, CBL, CBTC, Methoxy-THC, CBC, CBG, Δ9-THCH, and CBN). The cannabinoids compound profile showed Δ9-THC had the highest overall content and was indicated as the most important compound in the cannabis plant clustering profile. Among the various regions, Aceh Besar had the highest cannabis content. Statistical analysis of Raman spectroscopy and GC–MS data found (1) revealed compounds responsible for clustering cultivars between clusters, (2) variation among cannabis chemical profiles as a result of growing environment, and (3) facilitated prediction of cannabis profiles in helping to categorise regions of unknown cannabis origin based on chemical profiles.
Conclusion
Raman spectroscopy and GC–MS have proven reliable and efficient methods for classifying Cannabis sativa based on its cannabinoid profile in Aceh, Indonesia. The findings help reveal the geographical origin of the growing location of cannabis plant specimens. All five cannabis samples contained a major Δ9-THC psychoactive constituent. The highest Δ9-THC content comes from AB due to the influence of environmental factors. Parametric test analysis concluded that there was no significant effect of geographical origin related to the relatively close distance range of samples. Additionally, comparing these methods with other analytical techniques will support defined classification models and improve their application in forensic science, particularly in drug enforcement and quality assessment.
Introduction
Background
Cannabis, which is a psychoactive plant, refers to the leaves, flowers, stems, and seeds (Fig. 1). Cannabis has only one genus with one species (sativa) that is highly variable. However, cannabis can also be classified into several subspecies or varieties of Cannabis sativa, such as C. sativa variety indica, C. sativa variety ruderalis, and C. sativa variety afghanica (Hourfane et al. 2023). Cannabis is a plant that is used globally mainly as a source of psychoactive substances, where 2.5% of the world’s population use cannabis (Farrelly et al. 2023). But in Indonesia, cannabis is a Class I narcotic with a prevalence of use reaching 41.4% and is the most widely used illicit drug as regulated in Law Number 35 of 2009 (Indriana et al., 2024). Aceh is one of the main areas producing quality C. sativa in Indonesia, due to its geographical and climatic conditions that support the optimal growth of this plant (Sukri et al. 2020).
Growth stages of C. sativa from seed to mature plant in Aceh, Indonesia. a C. sativa seeds with variations in size and color. b Juvenile C. sativa plant with serrated leaves growing in soil. c Leaves, stems, and roots of C. sativa at various growth stages. d Mature C. sativa plant in natural setting
Currently, most countries around the world consider cannabis to be an illegal drug that can be abused. Despite this, cannabis has the potential for abuse and its illegal status at the federal level in Indonesia. Research on the content of chemical compounds and pharmacological aspects has shown that cannabis also has medicinal properties. Over its long history, cannabis has been used by humans as a medicinal plant, intoxicant, and recreational substance (Hourfane et al. 2023; Sagar & Gruber 2025). Cannabis can produce unique compounds, namely phytocannabinoids (Smith et al. 2022). When consumed, phytocannabinoids act on the Central Nervous System (CNS) and Peripheral Nervous System (PNS), causing changes in perception and producing euphoric, analgesic, and other effects (Barrales-Cureño et al. 2020). The impact of these key cannabinoid compounds explains why the cultivation and illegal consumption of cannabis-based products are widespread today.
One of the commonly used cannabis profiling methods is classification based on physical appearance, external environmental aspects, and tetrahydrocannabinol (THC) content in the sample. However, other cannabinoid compounds such as cannabigerol (CBG), cannabinol (CBN), and tetrahydrocannabivarin (THCV) also play a role in cannabis differentiation, especially for medical and therapeutic applications (Blebea et al. 2024; Curran et al. 2020). The principle of classification refers to the existence of morphological and physiological differences depending on external factors and the origin of the cultivated plant (Hesami et al. 2023). Novotny et al. (1976) stated that different chromatograms were obtained from Cannabis samples with different regional origins, so there may be a correlation between changes in compound content and geographic origin. Tzimas et al. (2022) also reported the impact of changing environmental conditions on the chemical composition and variability of cannabinoids in C. sativa.
A wide variety of analytical techniques have been used to perform cannabis profiling, including thin layer chromatography (Radosavljević-Stevanović et al. 2023), profiling with HPLC (Lynch et al. 2023), GC–MS (Arena et al. 2022), and H-NMR to fingerprint aqueous extracts and cannabis tinctures (Politi et al. 2008). However, GC is the specific and most commonly used instrument to analyse cannabinoid content (Baron 2015). Raman spectroscopy has also been successfully used to investigate THC content in samples with the advantage of its non-destructive nature (Sanchez et al. 2020). However, questions remain as to whether Raman spectroscopy combined with GC–MS and statistical analysis using parametric tests can differentiate cannabis species from different geographical origins to determine their origin. Moreover, there is no comprehensive classification available for the chemical profiles for samples in the Aceh region of Indonesia, which requires exploration of the similarities/differences that may exist among cannabis samples from the region.
In this study, a comprehensive investigation was conducted to identify important compounds and determine the presence or absence of variations in the chemical profile of C. sativa as a result of cultivating the plants in different environments from five regions in Aceh: Aceh Besar, Aceh Tengah, Bireuen, Lhokseumawe, and Pidie Jaya. Therefore, this research aims to develop a classification method based on the chemical profile analysis of cannabis from various regions in Indonesia, particularly Aceh, by comparing the main content of A9-THC through a combination of tests using microscopy, Raman spectroscopy, GC–MS, and ANOVA analysis, with the hope of assisting authorities in tracing the source of cannabis plants in Aceh based on their chemical composition and gaining further insight into the regions producing high-quality cannabis plants in Aceh as a strategy for law enforcement and forensic purposes.
Materials and methods
Materials and samples
Five samples of cannabis plants (Cannabis sativa) were confiscated from five different regions in Aceh Province, Indonesia, namely Aceh Besar (AB), Lhokseumawe (LS), Pidie Jaya (PJ), Bireuen (BR), and Aceh Tengah (AT). Samples were obtained from the Forensic Field Laboratory of the North Sumatra Regional Police, Indonesia. All samples collected were confirmed as C. sativa, with possible variations in phenotype and cannabinoid content due to differences in geographic origin as well as post-harvest morphological conditions. Samples were received in dry condition, consisting of buds, leaves, stems, seeds, and flowers. Methanol and Sodium Hydroxide were obtained from Merck (Darmstadt, Germany), Fast Blue B (FBB) salt was obtained from Sigma-Aldrich (Missouri, American), and the standard used was Δ9-THC Cerilliant Supelco 1.0 mg/L. Location details can be seen in Table 1.
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https://link.springer.com/article/10.1186/s41935-025-00480-y
