Journals ITB
JETS Journal of Engineering and Technological Sciences
  1. Home
  2. Archives
  3. Vol 49 (2024) Issue 2
  4. Articles

Isolation and Identification of Bioactive Flavonoid Glycosides from Syzygium Jambos and Their Α-glucosidase Inhibitory Activity

Published: | Vol 49, Issue 2 (2024)

Authors

Abstract

Diabetes has grown into a demanding global public health problem, with generally 537 million individuals afflicted worldwide. Without appropriate intervention, the risk of developing chronic complications increases. One of the practical actions for controlling type 2 diabetes mellitus is suppressing carbohydrate-hydrolyzing enzymes such as α-glucosidase. Syzygium jambos has a long history as a traditional medicine. It

Keywords

⍺-glucosidase; Diabetic; Syzygium jambos; Quercetin 3-O-xylosyl-(1→2)-rhamnoside; myricetin 3-O-xylosyl-(1→2)-rhamnoside

*Correspondence

Muhamad Insanu Email: insanu99@itb.ac.id

Diabetes has grown into a demanding global public health problem, with generally 537 million individuals af<licted worldwide. Without appropriate intervention, the risk of developing chronic complications increases. One of the practical actions for controlling type 2 diabetes mellitus is suppressing carbohydrate-hydrolyzing enzymes such as α-glucosidase. Syzygium jambos has a long history as a traditional medicine. It's commonly used to treat diabetes, rheumatism, in<lammatory pain, and as a diuretic. This study aimed to investigate the α-glucosidase inhibitory properties of S. jambos leaves and their isolated compounds. Ethanol was used as the solvent for maceration, followed by fractionation using liquid-liquid extraction, vacuum liquid chromatography, column chromatography, and radial chromatography. The isolated compounds were identi<ied using thin-layer chromatography (TLC), TLC densitometry, shifting reagents, infrared (IR) spectroscopy, mass spectrometry, and nuclear magnetic resonance (NMR) spectrometry. The ethyl acetate fraction presented the most potent activity of α-glucosidase inhibitory, with an IC50 value of 0.31 ± 0.01 µg/mL, exceeding the acarbose as positive control (IC50 62.86 ± 1.24 µg/mL). Two <lavonoid glycosides, quercetin 3-O-xylosyl-(1→2)-rhamnoside and myricetin 3-O-xylosyl-(1→2)-rhamnoside, were identi<ied inside this fraction, with IC50 values of 65.63 ± 1.36 µg/mL and 188.96 ± 2.85 µg/mL, respectively. These <indings suggest that S. jambos has potential as a natural source for developing dietary supplements to control hyperglycemia af<iliated with type 2 diabetes mellitus.

Keywords: ⍺-glucosidase; Diabetic; Syzygium jambos; Quercetin 3-O-xylosyl-(1→2)-rhamnoside; myricetin 3-O-xylosyl-(1→2)-rhamnoside

ISOLASI DAN IDENTIFIKASI GLIKOSIDA FLAVONOID BIOAKTIF DARI SYZYGIUM JAMBOS DAN AKTIVITAS PENGHAMBAT ENZIM Α-GLUKOSIDASE

ABSTRAK

Diabetes telah berkembang menjadi masalah kesehatan global yang serius, dengan sekitar 537 juta orang di seluruh dunia menderita penyakit ini. Tanpa intervensi yang tepat, risiko berkembangnya komplikasi kronis akan meningkat. Salah satu tindakan yang efektif untuk mengendalikan diabetes melitus tipe 2 adalah dengan menekan enzim penghidrolisis karbohidrat seperti α-glukosidase. Syzygium jambos memiliki sejarah panjang dalam penggunaannya pada pengobatan tradisional, umumnya digunakan untuk mengobati kondisi seperti kencing manis, rematik, nyeri in<lamasi, dan sebagai diuretik. Penelitian ini bertujuan untuk menyelidiki sifat penghambat α-glukosidase dari daun S. jambos dan senyawa yang diisolasi. Etanol digunakan sebagai pelarut untuk maserasi, diikuti oleh fraksinasi menggunakan ekstraksi cair-cair, kromatogra<i cair vakum, kromatogra<i kolom, dan kromatogra<i radial. Senyawa yang diisolasi diidenti<ikasi menggunakan kromatogra<i lapis tipis (KLT), densitometri KLT, analisis pergeseran yang diinduksi reagen, spektroskopi inframerah (IR), spektrometri massa, dan spektrometri resonansi magnetik nuklir (NMR). Fraksi etil asetat menunjukkan aktivitas penghambatan α-glukosidase terkuat, dengan nilai IC50 sebesar 0,31 ± 0,01 µg/mL, melebihi akarbose sebagai kontrol positif (IC50 62,86 ± 1,24 µg/mL). Dua glikosida <lavonoid, quercetin 3-O-xylosyl-(1→2)-rhamnoside dan myricetin 3-O-xylosyl-(1→2)-rhamnoside, diidenti<ikasi di dalam fraksi ini, dengan nilai IC50 masing-masing sebesar 65,63 ± 1,36 µg/mL dan 188,96 ± 2,85 µg/mL. Temuan ini menunjukkan bahwa S. jambos berpotensi sebagai sumber alami untuk mengembangkan suplemen makanan guna mengendalikan hiperglikemia yang terkait dengan diabetes melitus tipe 2.

Kata kunci: ⍺-glucosidase; Diabetes; Syzygium jambos; Quercetin 3-O-xylosyl-(1→2) rhamnoside; myricetin 3-O-xylosyl-(1→2)-rhamnoside

INTRODUCTION

Diabetes has appeared as a dominant global epidemiology concern, exerting a potent strain on healthcare systems worldwide, with its prevalence regularly increasing over recent decades (Lin et al. 2020). In agreement with the International Diabetes Federation (IDF), over 537 million people globally are presently affected by diabetes, with projections suggesting this statistic will increase to 643 million by 2030 and 783 million by 2045 (Hossain et al. 2024). In Indonesia presently, there are 18.69 million recorded cases of diabetes, with projections suggesting an increase to 22.99 million by 2025, 27.13 million by 2030, and 40.70 million by 2045 (Wahidin et al. 2024).

Type 2 diabetes mellitus (T2DM) constitutes more than 90% of diabetes cases worldwide, establishing it as a highly prevalent metabolic disorder (Vonia et al. 2022). Without timely intervention, the risk of developing chronic complications increases, including macroangiopathy, retinopathy, nephropathy, neuropathy, diabetic foot, increased vulnerability to infections, myopathy, osteoporosis, arthropathies, and liver damage (Farmaki et al. 2020).

Inflated postprandial blood glucose is a significant risk factor responsible for the onset and progression of type 2 diabetes (Feng et al. 2022). Controlling type 2 diabetes mellitus is affiliated with suppressing carbohydrate hydrolyzing enzymes and α-glucosidase in the digestive system to decrease postprandial glucose levels (Nor et al. 2023). Acarbose, miglitol, and voglibose are known as α-glucosidase inhibitors, but their use usually results in side effects like abdominal bloating, flatulence, diarrhea, and pneumatosis cystoides intestinalis. This has prompted the search for new

plant-based alternatives for α-glucosidase inhibition (Sheliya et al. 2015).

Syzygium jambos (L.) Alston, commonly called the rose apple, belongs to the Myrtaceae family and is extensively found across regions like South Africa, Central America, and Asia (Jiang et al. 2024). S. jambos has a rich history in traditional medicine. Its leaf decoctions are used as diuretics to treat rheumatism, inflammatory pain, and diabetes. Crushed leaf juice is used to manage fever, sore throat, and eye discomfort, while leaf powder is used to cool smallpox patients. Additionally, fruit, flowers, seeds, and bark are traditionally used to manage diarrhea, dysentery, asthma, bronchitis, and dysphonia (Wong et al. 2024). Previous studies have shown that S. jambos leaves exhibit a range of pharmacological activities, in addition to antibacterial, anti-acne, antioxidant, antiinflammatory, anti-arthritic, and anti-quorum sensing activities (Mohanty and Cock 2010; Sharma et al. 2013; Ghareeb et al. 2017; Sobeh et al. 2018; Apaza Ticona et al. 2021). Besides, the leaves are evident to manage hepatoprotective (Sobeh et al. 2018), antifungal (Noé et al. 2019), anticancer (Rocchetti et al. 2019), antihyperglycemic (Nor et al. 2023), and anti-acetylcholinesterase (Amir Rawa et al. 2022).

Previous studies have shown that S. jambos reveals strong potential as an α-glucosidase inhibitor, similar to S. myrtifolium, and comparable to other species like S. aqueum, S. aromaticum, S. cumini, S. malaccense, and S. polyanthum (Nor et al. 2023). Building on these findings, the present study aims to investigate further the α-glucosidase inhibitory activity of S. jambos leaf extract and isolate compounds. This research provides valuable insights into S. jambos as a promising alternative treatment for diabetes mellitus.

Figure 1.(A) Quercetin 3-0-xylosyl-\((1\rightarrow 2)\)-rhamnoside, (B) Myricetin 3-0-xylosyl-\((1\rightarrow 2)\)-rhamnoside

MATERIAL AND METHOD

Chemical and reagents

α-Glucosidase from Saccharomyces cerevisiae (G5003-100UN) and 4-nitrophenyl-α-Dglucopyranoside (pNPG, N1377-1G) purchased from Sigma-Aldrich (St. Louis, MO, USA). Acarbose hydrate was purchased from TCI Chemicals (Tokyo, Japan). Citroborate was used as specific spray reagent for thin-layer chromatography (TLC). Sodium hydroxide (NaOH), aluminum chloride (AlCl<sub>3</sub>), hydrochloric acid (HCl), sodium acetate (NaOAc), and boric acid (H<sub>3</sub>BO<sub>3</sub>) were utilized for chemical shift analysis. Methanol (LiChrosolv hypergrade for LC-MS, Merck Supelco) was used for mass spectrometry analysis.

Sample and extract preparation

The leaves of S. jambos were collected in Dago, Kabupaten Bandung, West Java, Indonesia, on December 30, 2019. The plant material was authenticated under voucher specimen B-151/2021 at the Research Center for Biology, Indonesian Institute of Sciences, Bogor, West Java, Indonesia. The leaves were washed with running water and dried in a drying cabinet at 40-50°C. Once thoroughly dried, the leaves were ground into a fine powder using a mechanical grinder. The powdered leaves were macerated in 96% ethanol at a 1:10 (w/v) ratio for 3x24 hours at room temperature. The resulting extract was filtered

through Whatman filter paper and concentrated using a rotary evaporator at 50°C. All extraction and concentration procedures were performed at the Department of Biological Pharmacy, Bandung Institute of Technology, Bandung, Indonesia.

Isolation and identification of active compounds

Compound isolation was guided by \(\alpha\)-glucosidase inhibition assay. Fractionation was performed using liquid-liquid extraction with solvents of increasing polarity: n-hexane, ethyl acetate, and methanol. A total of 30 g of the ethyl acetate fraction was further separated by vacuum liquid chromatography (VLC) using a gradient elution system consisting of n-hexane, ethyl acetate, and methanol. This process yielded 21 fractions. Fractions exhibiting similar chromatographic profiles and promising bioactivity were pooled and subjected to further purification via column chromatography, using an isocratic elution system of chloroform/methanol (8:2). The fractions with a significant activity was then purified using radial chromatography (chromatotron) with an isocratic elution system of chloroform/methanol/water (7:2.75:0.25), leading to the isolation of compounds A and B.

Identification isolate

The identification of isolated compounds was carried out using several analytical techniques.

Two-dimensional thin-layer chromatography (2D-TLC) was employed, followed by TLC with three different solvent systems. The spots were visualized using a citroborate spray reagent. TLC densitometry was performed to determine wavelength absorption and assess the purity of the isolates. Reagent-induced shift analysis was conducted using NaOH, AlCl3, AlCl3/HCl, NaOAc, and NaOAc/H3BO3. Further characterization involved infrared (IR), mass spectrometry, and nuclear magnetic resonance (NMR) spectroscopy.

2D-TLC was conducted with two solvent systems: ethyl acetate/methanol/water/acetic acid (8.5:1:0.5:0.25) and chloroform/methanol/water/acetic acid (7:2.75:0.5:0.25). Additionally, TLC analysis with three distinct eluents was performed using ethyl acetate/methanol/water/acetic acid (8.5:1:0.5:0.25), chloroform/methanol/water/acetic acid (7:2.75:0.5:0.25), and toluene/acetone/water/acetic acid (1:8.1:0.25).

TLC densitometry was performed using the chloroform/methanol/water/acetic acid (7:2.75:0.5:0.25) solvent system for the eluents of the isolates, with the densitometric analysis used to assess purity using Camag® TLC scanner 3.

The IR spectra were documented using a Jasco FTIR-4200 instrument. Mass spectra were collected using Waters Xevo TQD, running in negative mode with [M-H]⁻ ionization. The conditions were as follows: capillary voltage of 2.5 kV, source temperature of 150°C, desolvation temperature of 450°C, nitrogen gas flow rate of 900 L/h, cone gas flow rate of 1 L/h, cone voltage of 50 V, and collision energy of 30 V. NMR spectra were recorded on an Agilent 500 MHz system with a DD2 console, operating at 500 MHz for proton (1H) and 125 MHz for carbon (13C) analysis. Isolates A and B were dissolved in CD3OD and analyzed for 1H and 13C NMR.

α-Glucosidase inhibitory activity

The α-glucosidase inhibition assay was tested using a modified version of the method reported before (Nor et al. 2023). In a 96-well microplate, 30 μL of the sample, 36 μL of 0.1 M phosphate buffer (pH 6.8), and 17 μL of 6 mM p-nitrophenyl-α-Dglucopyranoside (pNPG) substrate were added. The mixture was preincubated at 37°C for 5 minutes. Afterward, 17 μL of α-glucosidase enzyme (0.2 U/mL) was added to each well, followed by an additional incubation for 15 minutes at 37°C to complete the reaction. The reaction was terminated by adding 100 μL of 200 mM sodium carbonate (Na2CO3)to each well. The absorbance of the reaction mixture was measured at 400 nm using a microplate reader (Tecan Infinite 200 Pro). All experiments were performed in triplicate. Acarbose was used as the positive control, while the negative control consisted of wells without any inhibitor.

The percentage of inhibition was calculated using the formula:

% Inhibition = \[\left(\frac{B1 - B2}{B1}\right) \times 100\%\]

Where:

  • B1 is the absorbance of the blank (PBS + pNPG + enzyme + Na2CO3),
  • B2 is the absorbance of the sample (sample extract + PBS + pNPG + enzyme + Na2CO3).

Controls included:

  • Blank control: PBS + Na2CO3, excluding the enzyme,
  • Sample control: sample extract + PBS + Na2CO3, excluding the enzyme.

The IC50 value, representing the concentration required to inhibit 50% of α-glucosidase activity, was determined using a linear regression model based on the relationship between sample concentration (x-axis) and percentage inhibition (y-axis).

Statistical analysis

The α-glucosidase inhibitory activity of the plant extracts, fractions, and isolated compounds was expressed as the mean ± standard deviation (SD). The data were analyzed using one-way analysis of variance (ANOVA), followed by the Games-Howell post hoc test to determine significant differences between groups. Statistical analysis was performed using SPSS software, version 26.0, and a p-value of less than 0.05 was considered statistically significant.

RESULT AND DISCUSSION

This study investigates the pharmacological activity of extracts, fractions, and active compounds from S. jambos leaves as inhibitors of the α-glucosidase enzyme for managing type 2 diabetes mellitus. The research extends previous work on the potential of S. jambos as an αglucosidase inhibitor. The α-glucosidase inhibitory activity of the extract, fractions, and isolated compounds was assessed using an in vitro assay (Table 1). Among the tested samples, the ethyl acetate fraction, obtained through liquid-liquid extraction, exhibited the greatest inhibitory activity, with an IC50 value of 0.31 ± 0.01 µg/mL. In contrast, Isolate B showed the weakest inhibition, with an IC50 value of 188.96 ± 2.85 µg/mL. Acarbose, used as the positive control, had an IC50 value of 62.86 ± 1.24 µg/mL. At concentrations of 100 µg/mL and 200 µg/mL, the S. jambos leaf extract demonstrated inhibition rates of 83.75 ± 1.37% and 92.27 ± 1.13%, respectively, comparable to S. myrtifolium and outperforming S. cumini, S. aqueum, S. malaccense, S. aromaticum, and S. polyanthum (Nor et al. 2023). The ethanol extract of S. jambosleaves exhibited an IC50 value of 1.22 ± 0.03 µg/mL (Table 1), consistent with earlier findings that reported an IC50 value of 0.90 ± 0.07 µg/mL. Moreover, the most potent inhibition was observed in the 70% ethanol extract with an IC50 value of 0.52 ± 0.01 µg/mL (Wong et al. 2024).

The ethyl acetate fraction demonstrated the strongest α-glucosidase inhibition activity compared to the water and n-hexane fractions, making it the focus for further isolation of active compounds. Using vacuum column chromatography, column chromatography, and radial chromatography, two isolates were obtained.

Isolate A was analyzed using two-dimensionalthinlayer chromatography (2D-TLC) and TLC with three different solvent systems, revealing a single spot. This spot reacted with citroborate, producing a green coloration. TLC densitometry indicated absorption maxima at 350 and 255 nm, with a purity of 100%. The presence of two absorption bands suggests the existence of a flavonoid (Harborne JB and TJ Mabry 1975). Isolate A was further characterized by reagent-induced shifts

using NaOH, AlCl3, AlCl3/HCl, NaOAc, and NaOAc/H3BO3. The addition of 2 M NaOH produced a bathochromic shift of 45 nm in band 1 without a reduction in intensity, indicating the presence of a 4'-OH group. AlCl3 caused an 18 nm bathochromic shift in band 1, suggesting the presence of a 5-OH group with oxygenation at position 6. AlCl3/HCl resulted in a 40 nm bathochromic shift in band 1, indicating an O-dihydroxy substitution on ring B. NaOAc produced a 15 nm bathochromic shift at 0 minutes with an absorbance of 1.584, suggesting a 7-OH group. After 5 minutes, the absorbance slightly decreased to 1.575, indicating the presence of 6,7 or 7,8 or 3',4'-dihydroxy groups (orthodihydroxy). AddingNaOAc/H3BO3 resulted in an 18 nm bathochromic shift in band 1, confirming a dihydroxy substitution on ring B (Markham 1982).

The infrared (IR) spectrum of Isolate A showed absorption bands at 3401.82 cm⁻¹ (O–H), 2927.41 cm⁻¹ (sp³ C–H), 1650.77 cm⁻¹ (C=C aromatic), 1874.47 cm⁻¹ (benzene ring), 1095.37 cm⁻¹ (C–O), 1419.35 cm⁻¹ (CH₃ bending), and 1735.62 cm⁻¹ (C=O). Mass spectrometry (MS) analysis showed a molecular ion peak at m/z 579.10 [M-H]⁻. The 1H NMR spectrum of Isolate A in CD3OD showed two singlet protons at δ 6.15 and 6.30, appropriate to meta-coupled protons at positions 6 and 8 on the flavonoid's A-ring. Chemical shifts at δ 7.35, 6.92, and 7.29 indicated three aromatic proton signals at positions 2', 5', and 6', consistent with a 3',4' disubstituted B-ring. The sugar moiety was identified as a disaccharide based on the two anomeric proton signals at δ 5.35 and 4.23, corresponding to rhamnose and xylose, respectively. The 13C NMR spectrum detected 26 carbon signals. Chemical shifts at δ 116.71 and 116.49 corresponded to carbons at positions 2' and 5', while signals at δ 146.6, 150.12, and 122.63 were assigned to carbons at positions 3', 4', and 6', respectively, confirming that the aglycone is quercetin. Based on the NMR data (Table 2), Isolate A was identified as quercetin 3-O-xylosyl-(1→2) rhamnoside (Fig. 1), consistent with previously reported data (Soicke et al. 1990; Slowing et al. 1994).

Isolate B was analyzed using two-dimensionalthinlayer chromatography (2D-TLC) and TLC with three different solvent systems, revealing a single

spot. This spot reacted with citroborate, producing a green coloration. TLC densitometry indicated absorption maxima at approximately 353 nm and 258 nm, with a purity of 91.71%. Similar to Isolate A, two absorption bands indicated the existence of a flavonoid, specifically a flavonol with three free or substituted hydroxyl (OH) groups (Harborne JB and TJ Mabry 1975). Isolate B was further analyzed through reagent-induced shifts using NaOH, AlCl3, AlCl3/HCl, NaOAc, and NaOAc/H3BO3. The addition of 2 M NaOH decreased intensity, indicating the presence of 3',4'-OH groups, O-dihydroxy groups on ring A or B, or adjacent 3-OH groups. AlCl3 produced a bathochromic shift of 40 nm in band 1, suggesting the presence of a 5-OH group. NaOAc induced a 13 nm bathochromic shift in band 2, with an absorbance of 1.494, indicating a 7-OH group. After 5 minutes, the absorbance decreased to 1.452, supporting the presence of 3',4'-dihydroxy groups (ortho-dihydroxy). Following the addition of NaOAc/H3BO3, band 1 exhibited a 19 nm bathochromic shift, confirming the presence of ortho-dihydroxy groups on ring B (Markham 1982).

The infrared (IR) spectrum of Isolate B showed absorption bands at 3401.82 cm⁻¹ (O–H), 2923.56 cm⁻¹ (sp³ C–H), 1600.63 and 1650.77 cm⁻¹ (C=C aromatic), 1801 cm⁻¹ (benzene ring), 1083.08 cm⁻¹ (C–O), 1376.93 cm⁻¹ (CH₃ bending), and 1716.34 cm⁻¹ (C=O). Mass spectrometry (MS) analysis displayed a molecular ion peak at m/z 595.10 [M-H]⁻. The 1H NMR spectrum of Isolate B in CD3OD revealed two singlet protons on the A-ring of the flavonoid at δ 6.15 and 6.30, corresponding to meta-coupled protons at positions 6 and 8. A chemical shift at δ 6.95 indicated two protons at positions 2' and 6' with identical chemical environments. As with Isolate A, the sugar moiety in Isolate B was identified as a disaccharide, based on the two anomeric proton signals at δ 5.28 and 4.46, corresponding to rhamnose and xylose, respectively. The 13C NMR spectrum detected 26 carbon signals. A chemical shift at δ 109.21 was assigned to carbons at positions 2' and 6', while a signal at δ 147.02 corresponded to carbons at positions 3' and 5'. A signal at δ 138.15 indicated the carbon at position 4'. This chemical shift data confirmed that the aglycone was myricetin. Based on the NMR spectrum (Table 2), Isolate B was identified as myricetin 3-O-xylosyl-(1→2) rhamnoside (Fig. 1), consistent with previously reported data (Soicke et al. 1990; Slowing et al. 1994).

Quercetin 3-O-xylosyl-(1→2)-rhamnoside and myricetin 3-O-xylosyl-(1→2)-rhamnoside are flavonol diglycosides previously reported in S. jambos leaves (Slowing et al. 1994; Nawwar et al. 2016; Sobeh et al. 2018). In addition to being present in S. jambos, these two compounds have also been identified in Moghania faginea (Soicke et al. 1990). There are few reports on these compounds. However, previous studies have noted that quercetin 3-O-xylosyl-(1→2)-rhamnoside is also found in Erythrospermum monoticolum (Harborne and Williams 2000) and Schizophragma hydrangeoides (Oh et al. 2022). In this study, both compounds were assessed for their α-glucosidase inhibitory properties. There are no prior reports on the α-glucosidase inhibition evaluation of these two compounds. The IC50 values for α-glucosidase inhibition were 65.63 ± 1.36 µg/mL for quercetin 3-O-xylosyl-(1→2)-rhamnoside and 188.96 ± 2.85 µg/mL for myricetin 3-O-xylosyl-(1→2) rhamnoside (Table 1). Previous research has reported that quercetin 3-O-xylosyl-(1→2) rhamnoside is active against acute inflammation (Harborne and Williams 2000) and exhibits protective effects against UV-induced skin aging (Oh et al. 2022).

Many reports have documented various compounds isolated from S. jambos leaves, aside from quercetin 3-O-xylosyl-(1→2)-rhamnoside and myricetin 3-O-xylosyl-(1→2)-rhamnoside. These include anacardic acid analogue; myricitrin; myricetin; ursolic acid; gallic acid; squalene (Sharma et al. 2013); 7-phloroglucinol(Jambone A-G); oleanic acid; betulinic acid; 3β-O-trans-pcoumaroylalphitolic acid; 3β-O-cis-pcoumaroylalphitolic acid; 5,7-dihydroxy-6,8 dimethyl-4'-methoxyflavone; 6-desmethylsideroxylin (Li et al. 2015); quercetin 3-O-xylosyl- (1→2)-xyloside; quercetin 3-O-glucuronide; myricetin 3-O-glucoside; myricetin 7-methylether 3-O-xylosyl-(1→2)-rhamnoside; myricetin 3',5' dimethyl ether 3-O-xylosyl-(1→2)-rhamnoside; myricetin 3',5'-dimethyl ether 3-O-rhamnoside; 1,2,3-tri-O-galloyl-β-glucose; ellagic acid (Nawwar

et al. 2016); malic acid; citric acid; hexahydroxydiphenoyl-hexoside; castalagin/vescalagin isomer; galloyl-HHDP-DHHDP-hexoside; bis-HHDP-hexoside; galloyl-bis-HHDP-hexoside (casuarinin); ellagic acid pentoside; ellagic acid rhamnoside; myricetin rhamnoside; rosmarinic acid rhamnoside (Sobeh et al. 2018); quercetin-3-O-rutinoside; prenylbenzoic acid 4-β-D-glucoside; morolic acid 3-O-caffeate; 5,4'-dihydroxy, 7-methoxy, 6-methyl-flavone; 3,4,5-trihydroxybenzoic acid; quercetin; isoetin-7- O-β-D-glucopyranoside; 4'-hydroxy-3' methoxyphenol-β-D-[6-O-(4"-hydroxy-3",5" dimethoxybenzoate)] glucopyranoside (Ghareeb et al. 2017); 5-hydroxymethyl-2-furfural; betulin (Rajkumari et al. 2018); stigmasterol; lupeol; syringic acid; beta-amyrin; bergenin derivatives; myricetin derivatives; kaempferol derivatives; friedelin; stigmasterol glucoside; myrigalone G; galloyl castalagin; tellimagrandin II; coriariin A; coriariin B; praecoxin A; praecoxin B; vescalagin; vescalin; trigalloyl glucose; di-HHDP glucose; pomolic acid (Wong et al. 2024).

The α-glucosidase inhibition of S. jambos leaf extracts and fractions exceeds that of the isolated compounds (Table 1). Plant extracts often contain various bioactive compounds, and the therapeutic efficacy of medicinal plants can be attributed to the synergistic interactions between these compounds (Vaou et al. 2022). Compared to isolated compounds, the enhanced enzyme inhibition observed in extracts and fractions suggests a synergistic or potentiating effect where the combined activity exceeds that of individual components (Rasoanaivo et al. 2011). This finding supports further exploration of S. jambos to isolate additional compounds and evaluate their αglucosidase inhibitory effects, individually and in combination. The potent inhibition shown by the extracts and fractions demonstrates significant potential for further development. Future research on S. jambos should focus on detailed evaluations, including toxicity assessments, preclinical studies, and clinical trials, to advance its potential as an alternative α-glucosidase inhibitor alongside current options like acarbose, miglitol, and voglibose.

Table 1. The IC50 Value of Extract, Fraction, and Isolate Compound of S. jambos against ⍺-Glucosidase Enzyme

SamplesIC50 (µg/mL)
Ethanol extract1.22 ± 0.03a
n-hexane fraction41.53 ± 0.56
Ethyl Acetate fraction0.31 ± 0.01
Water fraction1.21 ± 0.08a
Isolate A65.63 ± 1.36b
Isolate B188.96 ± 2.85
Acarbose62.86 ± 1.24b

Note: Data are expressed as the mean ± standard deviation (n = 3). Means within the same column followed by identical letters are not statistically different at p < 0.05.

Table 2. NMR Data of Isolate A and Isolate B

PositionIsolate A
(Quercetin 3-O-xylosyl-(1→2)-rhamnoside)		Isolate B
(Myricetin 3-O-xylosyl-(1→2)-rhamnoside)
13C
1H
δC
δH (J in Hz)		13C1H
δCδH (J in Hz)
1
2158.73158.69
3136.53136.67
4179.43179.51
5159.84158.84
6100.956.15 (1H, s)100.786.15 (1H, d, 2.05)
7166.16168.5
895.516.30 (1H, s)94.986.30 (1H, d, 2)
9163.04163.05
10104.93104.96
1'122.8121.73
PositionIsolate A
(Quercetin 3-O-xylosyl-(1→2)-rhamnoside)		Isolate B
(Myricetin 3-O-xylosyl-(1→2)-rhamnoside)
13C
δC		1H13C
δC		1H
δH (J in Hz)
δH (J in Hz)
2'116.717.35 (1H, d, 2.1)109.216.95 (2H, s)
3'146.6147.03
4'150.12138.15
5'116.496.92 (1H, d, 8.25)147.03
6'122.637.29 (1H, dd, 2.05, 8.2)109.216.95 (2H, s)
1"103.175.35 (1H, s)103.235.28 (1H, d, 1.55)
2"82.733.05 - 3.8982.823.05 - 3.84
3"70.963.05 - 3.8970.973.05 - 3.84
4"73.73.05 - 3.8973.803.05 - 3.84
5"71.93.05 - 3.8971.913.05 - 3.84
6"17.671.02 (3H, d, 6.2)17.71.02 (3H, d, 6.2)
1'''107.724.23 (1H, d, 7.65)107.814.46 (1H, d, 7.65)
2'''75.253.05 - 3.8975.283.05 - 3.84
3'''77.753.05 - 3.8977.763.05 - 3.84
4'''71.773.05 - 3.8971.743.05 - 3.84
5'''67.023.05 - 3.8966.973.05 - 3.84

d, duplet; dd, duplet of duplet; s, singlet

CONCLUSION

S. jambos exhibits significant α-glucosidase inhibitory activity. The ethyl acetate fraction demonstrated the highest inhibitory effect among the extracts and fractions tested. Quercetin 3-Oxylosyl-(1→2)-rhamnoside and myricetin 3-Oxylosyl-(1→2)-rhamnoside, both flavonoid glycosides isolated from S. jambos, showed αglucosidase inhibition, though less potent compared to the extract and its fractions. Quercetin 3-O-xylosyl-(1→2)-rhamnoside exhibited inhibitory activity comparable to acarbose, with no statistically significant difference. These findings highlight the potential of S. jambos extracts and fractions as effective α-glucosidase inhibitors, offering promise for further development into dietary supplements to manage hyperglycemia associated with type 2 diabetes mellitus.

FUNDING

The author(s) received no financial support for the research, authorship, and/or publication of this article.

ACKNOWLEDGEMENT

The researchers would like to thank the NMR Laboratory, Faculty of Natural Science and Mathematics, Institut Teknologi Bandung.

DECLARATION OF COMPETING INTEREST

The authors declare that there are no conflicts of interest, financial or personal, that could have influenced the work presented in this paper.

References

  1. Amir Rawa MS, Nurul Azman NA, Mohamad S, Nogawa T, Wahab HA, 2022, In Vitro and In Silico Anti-Acetylcholinesterase Activity from Macaranga tanarius and Syzygium jambos, Molecules 27, doi: 10.3390/molecules27092648. DOI: 10.3390/molecules27092648
  2. Apaza Ticona L, Souto Pérez B, Martín Alejano V, Slowing K, 2021, Anti-inflammatory and Anti-arthritic Activities of Glycosylated Flavonoids from Syzygium jambos in Edematogenic Agent-Induced Paw Edema in Mice, Revista Brasileira de Farmacognosia 31: 429–441, doi: 10.1007/s43450-021-00167-0. DOI: 10.1007/s43450-021-00167-0
  3. Farmaki P, Damaskos C, Garmpis N, Garmpi A, Savvanis S, Diamantis E, 2020, Complications of the Type 2 Diabetes Mellitus, Perspective Current Cardiology Reviews 16: 249-251, doi: 10.7326/0003-4819-152-5-201003020-01003. DOI: 10.7326/0003-4819-152-5-201003020-01003
  4. Feng Y, Nan H, Zhou H, Xi P, Li B, 2022, Mechanism of inhibition of α-glucosidase activity by bavachalcone, Food Science and Technology (Brazil) 42, doi: 10.1590/fst.123421. DOI: 10.1590/fst.123421
  5. Ghareeb MA, Hamed MM, Abdel-Aleem AAH, Saad AM, Abdel-Aziz MS, Hadad AH, 2017, Extraction, isolation, and characterization of bioactive compounds and essential oil from Syzygium jambos, Asian Journal of Pharmaceutical and Clinical Research 10: 194–200, doi: 10.22159/ajpcr.2017.v10i8.18849. DOI: 10.22159/ajpcr.2017.v10i8.18849
  6. Harborne JB, TJ Mabry HM, 1975, The Flavonoids. Springer Science+Business Media Dordrecht.
  7. Harborne JB, Williams CA, 2000, Advances in flavonoid research since 1992, Phytochemistry 55: 481–504, doi: 10.1016/S0031-9422(00)00235-1. DOI: 10.1016/s0031-9422(00
  8. Hossain MJ, Al-Mamun M, Islam MR, 2024, Diabetes mellitus, the fastest growing global public health concern: Early detection should be focused, Health Sci Rep 7, doi: 10.1002/hsr2.2004. DOI: 10.1002/hsr2.2004
  9. Jiang Y, Liu F, Zhang D, Fu X, Chen S, Wei M, Li Z, Lei H, Niu H, 2024, Identification of Novel -Glucosidase Inhibitors from Syzygium jambos (L.) Using Spectroscopy and Molecular Docking, Plant Foods for Human Nutrition 79: 73–82, doi: 10.1007/s11130-023-01123-0. DOI: 10.1007/s11130-023-01123-0
  10. Li GQ, Zhang YB, Wu P, Chen NH, Wu ZN, Yang L, Qiu RX, Wang GC, Li YL, 2015, New Phloroglucinol Derivatives from the Fruit Tree Syzygium jambos and Their Cytotoxic and Antioxidant Activities, J Agric Food Chem 63: 10257–10262, doi: 10.1021/acs.jafc.5b04293. DOI: 10.1021/acs.jafc.5b04293
  11. Lin X, Xu Y, Pan X, Xu J, Ding Y, Sun X, Song X, Ren Y, Shan PF, 2020, Global, regional, and national burden and trend of diabetes in 195 countries and territories: an analysis from 1990 to 2025. Sci Rep 10, doi: 10.1038/s41598-020-71908-9. DOI: 10.1038/s41598-020-71908-9
  12. Markham KR, 1982, Techniques of Flavonoid Identification, Academic Press.
  13. Mohanty S, Cock IE, 2010, Bioactivity of Syzygium jambos methanolic extracts: Antibacterial activity and toxicity, Pharmacognosy Res 2: 4–9, doi: 10.4103/0974-8490.60577. DOI: 10.4103/0974-8490.60577
  14. Nawwar MA, Hashem AN, Hussein SA, Swilam NF, Becker A, Haertel B, Lindequist U, El-Khatib A, Linscheid MW, 2016, Phenolic profiling of an extract from Eugenia jambos L. (Alston) - The structure of three flavonoid glycosides - Antioxidant and cytotoxic activities, Pharmazie 71: 162–168, doi: 10.1691/ph.2016.5747. DOI: 10.1691/ph.2016.5747
  15. Noé W, Murhekar S, White A, Davis C, Cock IE, 2019, Inhibition of the growth of human dermatophytic pathogens by selected australian and asian plants traditionally used to treat fungal infections, J Mycol Med 29: 331–344, doi: 10.1016/j.mycmed.2019.05.003. DOI: 10.1016/j.mycmed.2019.05.003
  16. Nor I, Wirasutisna KR, Hartati R, Insanu M, 2023, The α-glucosidase inhibitory activity of avicularin and 4-O-methyl gallic acid isolated from Syzygium myrtifolium leaves, Saudi Pharmaceutical Journal 31: 101677, doi: 10.1016/j.jsps.2023.06.010. DOI: 10.1016/j.jsps.2023.06.010
  17. Oh SY, Kim SC, Hyun HB, Hyeon H, Go B, Jung YH, Ham YM, 2022, Protective effects of quercetin-3-glucosyl-(1-2)-rhamnoside from Schizophragma hydrangeoides leaves on ultraviolet A-induced photoaging in human dermal fibroblasts, J Appl Biol Chem 65: 277–286, doi: 10.3839/jabc.2022.036. DOI: 10.3839/jabc.2022.036
  18. Rajkumari J, Borkotoky S, Murali A, Busi S, 2018, Anti-quorum sensing activity of Syzygium jambos (L.) Alston against Pseudomonas aeruginosa PAO1 and identification of its bioactive components, South African Journal of Botany 118: 151–157, doi: 10.1016/j.sajb.2018.07.004. DOI: 10.1016/j.sajb.2018.07.004
  19. Rasoanaivo P, Wright CW, Willcox ML, Gilbert B, 2011, Whole plant extracts versus single compounds for the treatment of malaria: Synergy and positive interactions, Malar J 10.
  20. Rocchetti G, Lucini L, Ahmed SR, Saber FR, 2019, In vitro cytotoxic activity of six Syzygium leaf extracts as related to their phenolic profiles: An untargeted UHPLC-QTOF-MS approach, Food Research International 126, doi: 10.1016/j.foodres.2019.108715. DOI: 10.1016/j.foodres.2019.108715
  21. Sharma R, Kishore N, Hussein A, Lall N, 2013, Antibacterial and anti-inflammatory effects of Syzygium jambos L. (Alston) and isolated compounds on acne vulgaris, BMC Complement Altern Med 13: 1–10, doi: 10.1186/1472-6882-13-292. DOI: 10.1186/1472-6882-13-292
  22. Sheliya MA, Rayhana B, Ali A, Pillai KK, Aeri V, Sharma M, Mir SR, 2015 Inhibition of α-glucosidase by new prenylated flavonoids from euphorbia hirta L. herb, J Ethnopharmacol 176: 1–8, doi: 10.1016/j.jep.2015.10.018. DOI: 10.1016/j.jep.2015.10.018
  23. Slowing K, Söllhuber M, Carretero E, Villar A, 1994, Flavonoid glycosides from Eugenia jambos, Phytochemistry 37: 255–258, doi: 10.1016/0031-9422(94)85036-4. DOI: 10.1016/0031-9422(94
  24. Sobeh M, Esmat A, Petruk G, Abdelfattah MAO, Dmirieh M, Monti DM, Abdel-Naim AB, Wink M, 2018, Phenolic compounds from Syzygium jambos (Myrtaceae) exhibit distinct antioxidant and hepatoprotective activities in vivo, J Funct Foods 41: 223–231, doi: 10.1016/j.jff.2017.12.055. DOI: 10.1016/j.jff.2017.12.055
  25. Soicke H, Goner’ K, Waring’ H, 1990, Flavonol Glykoskie aus Moghaniafaginea Flavonol Glycosides from Moghania faginea, Planta Med 56, doi: 10.1055/s-2006-960996. DOI: 10.1055/s-2006-960996
  26. Vaou N, Stavropoulou E, Voidarou C, Tsakris Z, Rozos G, Tsigalou C, Bezirtzoglou E, 2022, Interactions between Medical Plant-Derived Bioactive Compounds: Focus on Antimicrobial Combination Effects, Antibiotics 11, doi: 10.3390/antibiotics11081014. DOI: 10.3390/antibiotics11081014
  27. Vonia S, Hartati R, Insanu M, 2022, In Vitro Alpha-Glucosidase Inhibitory Activity and the Isolation of Luteolin from the Flower of Gymnanthemum amygdalinum (Delile) Sch. Bip ex Walp, Molecules 27, doi: 10.3390/molecules27072132. DOI: 10.3390/molecules27072132
  28. Wahidin M, Achadi A, Besral B, Kosen S, Nadjib M, Nurwahyuni A, Ronoatmodjo S, Rahajeng E, Pane M, Kusuma D, 2024, Projection of diabetes morbidity and mortality till 2045 in Indonesia based on risk factors and NCD prevention and control programs, Sci Rep 14, doi: 10.1038/s41598-024-54563-2. DOI: 10.1038/s41598-024-54563-2
  29. Wong PL, Ramli NS, Tan CP, Azlan A, Abas F, 2024, NMR-based metabolomics and UHPLC-ESI-MS/MS profiling of Syzygium jambos in relation to their antioxidant and anti-hyperglycemic activities, Arabian Journal of Chemistry 17, doi: 10.1016/j.arabjc.2023.105546. DOI: 10.1016/j.arabjc.2023.105546