
==== Front
J Adv Pharm Technol Res
J Adv Pharm Technol Res
JAPTR
J Adv Pharm Technol Res
Journal of Advanced Pharmaceutical Technology & Research
2231-4040
0976-2094
Wolters Kluwer - Medknow India

JAPTR-15-144
10.4103/JAPTR.JAPTR_9_24
Original Article
Botanical and pharmacognostic investigation of Strobilanthes kalimantanensis
Prabowo Wisnu Cahyo
Kuncoro Hadi
Irawan Budi 1
Kusuma Sri Agung Fitri 2
Susilawati Yasmiwar 2
Department of Pharmaceutical Sciences, Faculty of Pharmacy, Mulawarman University, Samarinda, East Kalimantan, Indonesia
1 Department of Biology, Faculty of Mathematics and Natural Sciences, Padjadjaran University, Jatinangor, Indonesia
2 Department of Biology Pharmacy, Faculty of Pharmacy, Padjadjaran University, Jatinangor, Indonesia
Address for correspondence: Mr. Wisnu Cahyo Prabowo, Department of Pharmaceutical Sciences, Faculty of Pharmacy, Mulawarman University, Samarinda, East Kalimantan 75119, Indonesia. E-mail: wisnu@farmasi.unmul.ac.id
Jul-Sep 2024
22 7 2024
15 3 144149
04 1 2024
03 6 2024
18 5 2024
Copyright: © 2024 Journal of Advanced Pharmaceutical Technology & Research
2024
https://creativecommons.org/licenses/by-nc-sa/4.0/ This is an open access journal, and articles are distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 License, which allows others to remix, tweak, and build upon the work non-commercially, as long as appropriate credit is given and the new creations are licensed under the identical terms.
Under a hidden waterfall in the interior of the tropical rainforest of East Kalimantan, a new medicinal plant that produces essential oil (EO) was found with the name Strobilanthes kalimantanensis. The aim was to investigate the botanical and evaluate the pharmacognostic characteristics of S. kalimantanensis leaves from East Kalimantan, Indonesia. Pharmacognostic studies can provide recommendations for establishing quality control standards or guidelines for cultivating, harvesting, and processing S. kalimantanensis to ensure the consistent and reliable quality of medicinal products. Characteristic methods of S. kalimantanensis leaves include botanical macroscopic, fluorescence, physicochemical, and phytochemical evaluation. The plant characteristics of this plant are similar to S. kunthia and S. reptans but can be differentiated in the leaves and flowers. Fluorescence assay with sodium hydroxide 5% shows unique characteristics of secondary metabolites based on their ability to form dark green with black precipitate in Ultraviolet 365 nm. The physicochemical characteristics showed yield, water content, water-soluble, ethanol soluble, total ash value, and acid-insoluble ash. Phytochemicals showed the presence of alkaloids, polyphenols, terpenoids, and EO containing 23% trans-anethole. This evaluation report details the chemical composition, identity, and safety of S. kalimantanensis leaves.

Acanthaceae
East Kalimantan
essential oil
new Strobilanthes
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pmcINTRODUCTION

Borneo Island is the largest island in Indonesia. It has the third-largest tropical rainforest in the world, so it has a biodiversity that has the potential to be used in the development of new drugs.[1] Biodiversity in the form of medicinal plants has often been used as traditional medicine for generations. Most people use traditional medicines with proven efficacies empirically to justify their use scientifically and not solely on experience. Drug development efforts are needed to see which components are responsible for providing benefits to society.

Pharmacy standardization refers to a set of criteria, practices, and measurement techniques whose outcomes are components associated with standard requirements (chemical, biological, and pharmaceutical), such as stability assurances for pharmaceutical products. In other words, standardization also means guaranteeing that the final drug product (drug or extract product) has fixed, unchanging values for specific parameters.[2] Identifying botanical species or varieties is very important in the case of new medicinal plants. Documentation and collections can be used to identify medicinal and essential oil (EO) plants from the country of origin.

Strobilanthes kalimantanensis EO plant was found in the Inar waterfall forest, Temula Village, West Kutai Regency, East Kalimantan. The Dayak Tunjung and Dayak Benuaq traditionally use them as a medicine for fever and asthma of breath. Its natural habitat grows on the cliffs around the waterfall, which has a wet area. Based on the source, it is necessary to investigate the pharmacognostic, botanical, and activity of S. kalimantanensis. This information can be essential to guarantee sustainable and high-quality sources, such as pharmaceutical raw materials. To learn more about new species and their advantages, research is required on plant properties, chemical composition, and activity. The need for raw materials and distribution in the world is very high. There must be a guarantee of the quality of the product, and also, the scientific data, mainly pharmacognostic character, is limited. This research aims to evaluate the pharmacognostic characteristics of S. kalimantanensis from the Acanthaceae family to guarantee the marketed plants’ quality and authenticity.[3]

MATERIALS AND METHODS

Material

The equipment used in this study was a rotary evaporator (Buchi®), grinder, D7100 Nikon®, furnace, ultraviolet (UV) light (254 and 365 nm), azeotropic distillation (Phyrex®), and gas chromatography/mass spectrometry (GC-MS) 7890A (Agilent®). The materials used in this study consisted of Fluorescence reagents (NaOH 5%, HCl 5%, NH₄OH 25%, and H2SO4 5%) (Merck®), phytochemical reagent (Dragendorf, Mayer, Wagner, glacial-HCl, magnesium powder, FeCl3 1%, glacial-H2SO4) (Merck®), toluene (Merck®), ethanol 96%, chloroform (Merck®), and aquadest.

Collection and plant preparation

S. kalimantanensis seeds were collected from West Kutai and bred in Samarinda City, East Kalimantan. After 3 months, the fresh leaves are ready to be harvested. Number sample: S.431-BP2TKSD-DISP-9-2021 was collected at the Research and Development Institute for Natural Resources Conservation Technology in Samboja, Indonesia. Leaf samples were dried, powdered, and then macerated with 96% ethanol. EO extraction was performed using the hydrodistillation technique.

Macroscopic evaluation

The botany of S. kalimantanensis leaves was investigated using visual organ observations. Fresh leaf samples were identified visually based on organoleptic characteristics, including color, shape, and specific components inside the leaf samples.

Fluorescence evaluation

In earlier research, a process was followed to evaluate the fluorescence of the dried powder of the leaf sample.[4] The powder and crude ethanol extract were put into a drip plate and dripped with a reagent solution. Color changes were observed using visible light and ultraviolet at 254 and 365 nm.

Physicochemical evaluation

Physicochemical analysis, including water content, water-soluble, alcohol-soluble, ash value, and acid-insoluble ash value. The water content of azeotropic distillation using toluene. Ash content analysis uses temperatures at 500°C to remove organic matter. The acid-insoluble ash content can be used to calculate the inorganic content.[5]

Phytochemical evaluation

Phytochemical investigation of ethanol extract of S. kalimantanensis leaves was performed using chemical reagents based on the literature to identify phytoconstituents such as alkaloids, flavonoids, polyphenols, terpenoids, and saponin.[6] Trans-anethole investigation of EO content, diluted with chloroform (1/100). It was analyzed utilizing GC-MS (Agilent®). The EO (1 μL) using 30 m open tubular column, inner diameter 2 mm, gradient injector temperature (120°C, 200°C, 250°C, and 270°C), temperature in 8°C/min, elution with helium, and flow rate in 0.93 mL/min, comparing compound with 1% trans-anethole standard.

RESULTS

Authentication and macroscopic investigation

The authentication of S. kalimantanensis through the identification process at the Research Institute for Natural Resources Conservation Technology (BKSDA-Wanariset) Samboja, East Kalimantan, Indonesia. Extraordinary institution for the conservation of tropical rainforest plants in East Kalimantan. The classification is shown in Table 1.

Table 1: Classification of Strobilanthes kalimantanensis

Division	Magnoliophyta	
Class	Magnoliopsida	
Sub-class	Asteridae	
Ordo	Scrophulariales	
Family	Acanthaceae	
Genus	Strobilanthes	
Species	Strobilanthes kalimantanensis	

Fluorescence evaluation

The fluorescence test shows the characteristics of metabolites in the sample that can fluoresce in UV light after adding reagents. Table 2 displays the findings of fluorescence experiments on leaf extracts and powders using various reagents.

Table 2: Fluorescence character of extract

Treatment	Visible	UV light	
		254 nm	365 nm	
Ethanol extract	Light green	Light green	Black with bright-red	
Aquadest	Dark black	Light green	Dark green	
+ NaOH 5%	Dark green	Light green	Dark green with black precipitate*	
+ HCl 5%	Yellowish green	Light green	Reddish black	
+ H2SO4 5%	Dark green	Light green	Reddish green	
+ NH4OH 25%	Yellowish-light green	Light green	Reddish black	
*Special characteristic. UV: Ultraviolet light

Physicochemical evaluation

The physiochemical parameters of simplicia powder are essential indicators for determining the quality and purity of herbal medicines. The physiochemical analysis includes [Table 3] water content, water-soluble, ethanol-soluble, total ash value, and acid-insoluble ash.

Table 3: Physiochemical parameters of simplicia

Parameter	Average (% w/w)	
Water content	1.41±0.050	
Water soluble	4.60±0.748	
Ethanol soluble	11.00±1.414	
Total ash value	7.83±0.001	
Acid insoluble ash	3.04±1.247	

Phytochemical evaluation

A sample of S. kalimantanensis EO extracted by hydrodistillation contains Trans-anethole as a marker in Figure 1. The results of GC-MS analysis of trans-anethole content in EO was 23%.

Figure 1 Gas chromatography mass spectrometry analysis of trans-anethole from essential oil sample

DISCUSSION

The Acanthaceae family of Strobilanthes is widely distributed from South to East Asia. Botanists have identified plants from tropical rainforests and examined their distribution, origins, and specific characteristics. S. kalimantanensis has never been found anywhere else, based on the reports of this botanical investigation of the new species. Confirmation has been strengthened by registration and storage of the Wanariset Herbarium collection (Number: WCP-01), which was discovered at the coordinates map as shown in Figure 2. Authentication of S. kalimantanensis is based on literature studies of several of its related species. The macroscopic study was carried out based on similarities to S. kunthia and S. reptans. Table 4 and Figure 3 show the complete physical observations-based morphology characteristics of the S. kalimantanensis plant. The habitat of this plant is in East Kalimantan, Indonesia, like many other Strobilanthes genera spread across Indonesia and Southeast Asia.[6] Based on a comparison of organ morphology against S. kunthia and S. reptans. S. kalimantanensis plant has smaller plant, leaf, and flower sizes. Its main characteristics can be seen in the abaxial leaf.

Figure 2 Map coordinates of Strobilanthes kalimantanensis

Figure 3 (a) Type opposite decussate leaf, the lower midrib, and the veins are red; (b) Purple axillaries flos with four lobes; (c) Calyx segments fused into a tube; (e) New leaves and brace roots at nodes; (d) Leaves and stems in old plants; (f) Pollen grains are oval from young to mature (green-yellow-black)

Table 4: Comparison of characteristics of Strobilanthes kalimantanensis with Strobilanthes kunthia and Strobilanthes reptans

Character	Strobilanthes kalimantanensis	Strobilanthes kunthia[7]	Strobilanthes reptans[8]	
Habit	Grassy; ca 1 m high, hydrophytic	Shrubs; ca 2 m high, branches stout	Grassy; ca 1 m high	
Distribution	Indonesia (East Borneo)	India, South East Asia	South East Asia	
Size of leaf (mm)	30×50	39×65	35×60	
Leaf shape	Elliptic oblong-ovate; apex obtuse or obtusely acuminate	Elliptic oblong-ovate; apex very obtusely acuminate	Elliptic oblong-ovate; very obtuse or obtusely acuminate	
Leaf margin	Entire to distinctly serrate	Entire to distinctly serrate	Sub-entire to repand or shallowly crenate	
Abaxial leaf	Dense white woolly indumentum; ventrally light red	Sparsely to densely white farinate	Sparsely pubescent; ventrally red	
Adaxial leaf	Green; glabrous	Green; glabrous or sparsely covered with short	Dark green; glabrous, ventrally red	
Inflorescence	Broad uninterrupted spikes, densely covered in short yellow hairs; 4–5 mm × 8–10 mm	Broad uninterrupted spikes, densely covered in short white hairs; 6–12 mm × 12–43 mm	Broad uninterrupted spikes, dense woolly indumentum; 6–7 mm × 13–15 mm	
Calyx fusion	Fused from the base for 0.2–0.3 of total length at anthesis; 4 lobes	Fused from the base for 0.3–0.4 of total length at anthesis; 5 lobes	Fused from the base for 0.5–0.6 of total length at anthesis; 5 lobed	
Bract: Calyx	Equal or shorter	Longer	Equal	
Corolla	Ovate equally divided; single lipped; 4–6 mm	Orbicular or suborbicular; clear 2-lipped; 20–30 mm	Equally divided; slightly 2-lipped; sub-orbicular, 11–12 mm	
Stamens	Present	Present	Present	
Ovary apex	Pubescence	Pubescence	Pubescence	

Inflorescence with four purple lobes in the form of broad uninterrupted spikes, densely covered with short yellow hairs. The base of the flower petals has a distinctive pattern of a combination of red and white lines. Wet soil is the S. kalimantanensis plant’s natural habitat. Hydrophyte plants have roots and stems in the water, and their leaves grow upward. It can be planted in tropical climates with lots of sunlight, as shown in Figure 4. Powder characteristics such as color and physical texture are essential in delivering raw materials. The physiochemical parameters of simplicia are important indicators for determining the quality and purity of herbal medicines.

Figure 4 (a) Strobilanthes kalimantanensis plant can be cultivated in wet soil, producing many branches with dense growth. (b) Simplicia leaf powder has a soft texture and light green color

Physiochemical values can later help determine the authenticity, purity, and contamination of natural medicinal plants or EO plants. Fluorescence is a phenomenon of chemical elements in secondary metabolites owned by plant material. Special characteristics of secondary metabolites occur in the fluorescence test using NaOH 5%. A dark green precipitate with black is formed when exposed to UV 365 nm, as shown in Table 2. The color resulting from the reaction with sodium hydroxide is related to the content of secondary metabolites such as anthocyanins, quinones, or coumarins. This compound is known to have antioxidant, antimicrobial, and anti-inflammatory activity.[910] This method can be an essential parameter of pharmacognostic evaluation in the field.[6] A more measurable analysis can also use Fourier-transform infrared spectroscopy for metabolomic profiling.[11] Phytochemical and physicochemical values are essential for agriculture quantitative standards.

Physicochemical values [Table 3] are important for evaluating crude drugs. The water content of the S. kalimantanensis extract is essential to maintain stability from the threat of contamination. The solubility of the extract in ethanol is better than water, making it easier to use. The acid-insoluble ash content with a value of 3.04% ± 1.247% indicates the presence of small silica or silicate contamination in the soil. A balanced silica content in the soil will help the absorption of beneficial minerals into the roots, which are used in metabolic processes.[12]

This EO plant is known to have a content of 0.547%.[13] GC-MS compounds identified in EO are octenol (attractants and traps),[14] linalool (antidepressant),[15] estragole, humulene (antioxidant, anti-inflammatory, and antimicrobial),[1617] threo-anethole glycol, neophytadiene (anxiolytic and anticonvulsant),[18] and the majority compound of trans-anethole (antibacterial).[19] Major compounds can be markers for the activity of EO as pharmaceutical raw materials. Trans-anethole in S. kalimantanensis leaves can be an alternative to the primary commodity of this component, which is currently sourced from the flowers of the Pimpinella anisum and Illicium verum.[20]

The ethanol extract of S. kalimantanensis leaves was detected to contain alkaloids, polyphenols, and terpenoids [Table 5], which are used therapeutically to treat hypertension, antimicrobial, and anti-inflammatory.[21] In the Acanthaceae family, several genera of Strobilanthes are known to have anticancer activity (S. crispus),[22] antimicrobial (S. glutinous),[23] antioxidant (S. erectus),[24] anti-inflammatory (S. formosanus),[7] antidiabetic (S. cordifolia),[25] antidepressant (S. japonici),[16] and analgesic (S. ciliatus).[26] This activity proves the potential for activity in the genus Strobilanthes, including the leaves of S. kalimantanensis. Physiochemical content and value are helpful in determining the authenticity, activity, purity, and contamination of natural medicinal plants or EO plants.

Table 5: Phytochemical screening of the extract

Constituent	Test	Result	Information	
Alkaloid	Dragendorff	+	Orange-red precipitate	
	Mayer	+	White precipitate	
	Wagner	+	Brown	
Flavonoid	HCl + Mg	–	Yellowish green	
Polyphenolic	FeCl3 1%	+	Dark green	
Terpenoid	H2SO4	+	Purplish red	
Saponin	Water with shaken + HCl	–	Green without foam	
+: Detected, –: Not detected

CONCLUSION

The preliminary phytochemical, physicochemical, and botanical investigation confirmed S. kalimantanensis as the new species. The assessment offers details on the botanical investigation, chemical ingredients of this simplicia and ethanol crude extract, and quantity of trans-anethole in EO for marker compound.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

Acknowledgments

The authors would like to appreciate the assistance of the Laboratory of Development of Farmaka Tropis, Mulawarman University, Central Laboratory of Padjajaran University, and BKSDA-Wanariset Samboja, Indonesia.
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REFERENCES

1. Misztal PK Owen SM Guenther AB Rasmussen R Geron C Harley P , Large estragole fluxes from oil palms in borneo Atmos Chem Phys 2010 10 4343 58
2. Ismaiel OA Abdelghani E Mousa H I.Eldahmy S Bayoumy BE Determination of estragole in pharmaceutical products, herbal teas and herbal extracts using GC-FID J Appl Pharm Sci 2016 6 144 50
3. Huang YC Wang JC Strobilanthes wallichii (Acanthaceae), a newly recorded species in Taiwan Taiwania 2009 54 93 6
4. Ahmad I Ambarwati NSS Arifuddin M Rijai L Mun’im A Pharmacognostic and cytotoxicity evaluation of Indonesia native plant of piper acre blume leaves (Piperaceae) Pharmacogn J 2017 9 400 4
5. Yuniarti R Nadia S Alamanda A Zubir M Syahputra RA Nizam M Characterization, phytochemical screenings and antioxidant activity test of Kratom Leaf ethanol extract (Mitragyna speciosa Korth) using DPPH method J Phys Conf Ser 2020 1462 1 7
6. Farnsworth NR Biological and phytochemical screening of plants J Pharm Sci 1966 55 225 76 5335471
7. Zhu X Xu Y Sun D Li H Chen L A new species of Strobilanthes (Acanthaceae) from the Western Ghats, India Taiwania 2020 65 167 71
8. Sarma J Barbhuiya HA Dey S Salunkhe CK Strobilanthes reptans (Acanthaceae): A new addition to the flora of India Rheedea 2021 31 58 61
9. Zhu X Xu Y Sun D Li H Chen L The genus Strobilanthes: Phytochemistry and pharmacology TMR Mod Herb Med 2022 5 15
10. Singh D Aeri V Phytochemical and pharmacological potential of Acanthus ilicifolius J Pharm Bioallied Sci 2013 5 17 20 23559819
11. Fonseca TA Von Rekowski CP Araújo R Oliveira MC Justino GC Bento L , The impact of the serum extraction protocol on metabolomic profiling using UPLC-MS/MS and FTIR Spectroscopy ACS Omega 2023 8 20755 66 37323376
12. Khan I Awan SA Rizwan M Ali S Hassan MJ Brestic M , Effects of silicon on heavy metal uptake at the soil-plant interphase: A review Ecotoxicol Environ Saf 2021 222 112510 34273846
13. Prabowo WC Narsa AC Andarista DB Kuncoro H Kusuma SAF Susilawati Y Gc-Ms profile and antibacterial activity of essential oil from Strobilanthes kalimantanensis leaves: A new species from East Kalimantan, Indonesia Int J Appl Pharm 2022 14 143 7
14. Bohbot JD Durand NF Vinyard BT Dickens JC Functional development of the octenol response in Aedes aegypti Front Physiol 2013 4 39 23471139
15. Dos Santos ÉR Maia JG Fontes-Júnior EA do Socorro Ferraz Maia C Linalool as a therapeutic and medicinal tool in depression treatment: A review Curr Neuropharmacol 2022 20 1073 92 34544345
16. Mahendra MY Purba RA Dadi TB Pertiwi H Estragole: A review of its pharmacology, effect on animal health and performance, toxicology, and market regulatory issues Iraqi J Vet Sci 2023 37 537 46
17. Rogerio AP Andrade EL Leite DFP Figueiredo CP Calixto JB Preventive and therapeutic anti-inflammatory properties of the Sesquiterpene α-humulene in experimental airways allergic inflammation Br J Pharmacol 2009 158 1074 87 19438512
18. Gonzalez-Rivera ML Barragan-Galvez JC Gasca-Martínez D Hidalgo-Figueroa S Isiordia-Espinoza M Alonso-Castro AJ In vivo neuropharmacological effects of neophytadiene Molecules 2023 28 3457 37110691
19. Kubo I Fujita KI Nihei KI Antimicrobial activity of anethole and related compounds from aniseed J Sci Food Agric 2008 88 242 7
20. Ahmad W Amir M Ahamad SR Alam P Alshehri S Ghoneim MM , Simultaneous determination of fenchone and trans-anethole in essential oils and methanolic extracts of Foeniculum vulgare Mill. Fruits obtained from different geographical regions using GC-MS approach Separations 2022 9 132
21. Monnerat CS Freitas MSM Vieira IJC Martins MA Carvalho AJC de Santos PC dos , Ajmalicine Bioproduction in Catharanthus roseus (L) G. Don inoculated with arbuscular mycorrhiza and fertilized with nitrogen Rev Bras Cienc do Solo 2018 42 1 12
22. Guan M Hee C Yen K Chye SM Ling APK Koh RY Review anticancer properties of Strobilanthes crispus: A review Processes 2021 9 1 19
23. Sedeek MS Kirollos FN Michel CG Kawy MAA Botanical and genetic characterization of Citrus maxima (Burm.) Merrill. F. Rutaceae Int J Pharm Pharm Sci 2016 9 260
24. Ajaib M Ishtiaq M Shafi F Bhatti KH Zahid MT Antimicrobial and antioxidant analysis of Strobilanthes glutinous: An unexplored medicinal plant Biosci Res 2020 17 1521 34
25. Kirubakaran D Selvam K Prakash P Shivakumar MS Rajkumar M In-vitro antioxidant, antidiabetic, anticholinergic activity of Iron/copper nanoparticles synthesized using Strobilanthes cordifolia leaf extract OpenNano 2023 14 4 16
26. Sebastian R Venkatachalam G Umamaheswari D Abirami MS Venkateswarlu B Evaluation of anticancer activity of Ethanolic Extract of Strobilanthes ciliatus leaves on HT–29 cell lines Asian J Biol Life Sci 2023 12 73 9
