HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology

Abstract The mechanical extraction of various phytochemicals from Phlomis stewartii (P. stewartii) was modeled using response surface methodology (RSM). The Box-Behnken design (BBD) was used to optimize the three independent variables comprising of 17 experimental runs, with the experiments randomly...

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Main Authors: Mamoon Ur Rasheed, Haroon Rashid, Shifa Iman, Sadaf Oranab, Muhammad Abdul Rahim, Mohamed H. Mahmoud, Roberto Castro-Muñoz, Eliasse Zongo
Format: Article
Language:English
Published: BMC 2025-08-01
Series:BMC Plant Biology
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Online Access:https://doi.org/10.1186/s12870-025-06997-7
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author Mamoon Ur Rasheed
Haroon Rashid
Shifa Iman
Sadaf Oranab
Muhammad Abdul Rahim
Mohamed H. Mahmoud
Roberto Castro-Muñoz
Eliasse Zongo
author_facet Mamoon Ur Rasheed
Haroon Rashid
Shifa Iman
Sadaf Oranab
Muhammad Abdul Rahim
Mohamed H. Mahmoud
Roberto Castro-Muñoz
Eliasse Zongo
author_sort Mamoon Ur Rasheed
collection DOAJ
description Abstract The mechanical extraction of various phytochemicals from Phlomis stewartii (P. stewartii) was modeled using response surface methodology (RSM). The Box-Behnken design (BBD) was used to optimize the three independent variables comprising of 17 experimental runs, with the experiments randomly arranged to minimize the effects of unexpected variation in the observed dependent variables {extraction yield, total phenolic content (TPC), and total flavonoid content (TFC)} due to systematic errors. The speed (X1, 100, 150 and 200 rpm), solvent volume (X2, 100 and 150 and 200 mL), and extraction time (X3, 2, 5 and 100 h) at 3-levels (high and low coded levels) were used to evaluate BBD of RSM. Results showed that the maximum value of yield, TPC and TFC in leaves ethanol extracts (LEE); flower ethanol extract (FEE), and whole plant ethanol extracts (WPEE) were observed at the following extraction conditions: A X1 of 150 rpm, X2 of 200 mL and X3 at 8 h. The minimum values were observed at an X1 of 150 rpm, X2 of 100 mL and X3 of 2 h. Analysis of variance (ANOVA) and the interaction effects of independent factors showed that X2 and X3 significantly positively influenced the response variables. These values closely matched the predicted yield, TPC and TFC. High-performance liquid chromatography (HPLC) analysis reveals that hydroxybenzoic acid, p-coumaric acid, gallic acid, chlorogenic acid, and salicylic acid, are the major secondary metabolites in the extracts. Gas chromatography-mass spectrometry (GC-MS) analysis showed the presence of different compounds such as thiazole, 2-ethylacridine, silicic acid, arsenous acid, 3,5-ethanoquinolin-10-one, and hexahydropyridine. All these identified compounds exhibit a wide range of various biological potential, including anti-viral, anti-inflammatory, anti-diabetic and anti-bacterial activities. The highest α-amylase inhibitory potential IC50 (53.33 ± 0.21 µg/mL), and α-glucosidase inhibitory potential IC50 (51.07 ± 0.17 µg/mL) were shown by LEE extracts. The study concludes that LEE extracts may have anti-inflammatory, immunomodulatory, antioxidant, hepatoprotective, and nephroprotective potential, suggesting potential for further investigation in diabetes management. Graphical Abstract
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spelling doaj-art-d6874ec855a84db4bf9d7ffa324b478e2025-08-20T03:45:56ZengBMCBMC Plant Biology1471-22292025-08-0125112010.1186/s12870-025-06997-7HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodologyMamoon Ur Rasheed0Haroon Rashid1Shifa Iman2Sadaf Oranab3Muhammad Abdul Rahim4Mohamed H. Mahmoud5Roberto Castro-Muñoz6Eliasse Zongo7Department of Chemistry, Government College UniversityDepartment of Physiology, Faculty of Veterinary and Animal Sciences, The Islamia University of BahawalpurDepartment of Pharmacology, Government College UniversityDepartment of Chemistry, Government College UniversityDepartment of Food Science & Nutrition, Faculty of Medicine and Allied Health Sciences, Times UniversityDepartment of Biochemistry, College of Science, King Saud UniversityDepartment of Sanitary Engineering, Faculty of Civil and Environmental Engineering, Gdansk University of TechnologyLaboratory of Research and Teaching in Animal Health and Biotechnology, Nazi Boni UniversityAbstract The mechanical extraction of various phytochemicals from Phlomis stewartii (P. stewartii) was modeled using response surface methodology (RSM). The Box-Behnken design (BBD) was used to optimize the three independent variables comprising of 17 experimental runs, with the experiments randomly arranged to minimize the effects of unexpected variation in the observed dependent variables {extraction yield, total phenolic content (TPC), and total flavonoid content (TFC)} due to systematic errors. The speed (X1, 100, 150 and 200 rpm), solvent volume (X2, 100 and 150 and 200 mL), and extraction time (X3, 2, 5 and 100 h) at 3-levels (high and low coded levels) were used to evaluate BBD of RSM. Results showed that the maximum value of yield, TPC and TFC in leaves ethanol extracts (LEE); flower ethanol extract (FEE), and whole plant ethanol extracts (WPEE) were observed at the following extraction conditions: A X1 of 150 rpm, X2 of 200 mL and X3 at 8 h. The minimum values were observed at an X1 of 150 rpm, X2 of 100 mL and X3 of 2 h. Analysis of variance (ANOVA) and the interaction effects of independent factors showed that X2 and X3 significantly positively influenced the response variables. These values closely matched the predicted yield, TPC and TFC. High-performance liquid chromatography (HPLC) analysis reveals that hydroxybenzoic acid, p-coumaric acid, gallic acid, chlorogenic acid, and salicylic acid, are the major secondary metabolites in the extracts. Gas chromatography-mass spectrometry (GC-MS) analysis showed the presence of different compounds such as thiazole, 2-ethylacridine, silicic acid, arsenous acid, 3,5-ethanoquinolin-10-one, and hexahydropyridine. All these identified compounds exhibit a wide range of various biological potential, including anti-viral, anti-inflammatory, anti-diabetic and anti-bacterial activities. The highest α-amylase inhibitory potential IC50 (53.33 ± 0.21 µg/mL), and α-glucosidase inhibitory potential IC50 (51.07 ± 0.17 µg/mL) were shown by LEE extracts. The study concludes that LEE extracts may have anti-inflammatory, immunomodulatory, antioxidant, hepatoprotective, and nephroprotective potential, suggesting potential for further investigation in diabetes management. Graphical Abstracthttps://doi.org/10.1186/s12870-025-06997-7P. stewartiiPhytochemicalsEnzyme inhibitionGC-MSHPLC
spellingShingle Mamoon Ur Rasheed
Haroon Rashid
Shifa Iman
Sadaf Oranab
Muhammad Abdul Rahim
Mohamed H. Mahmoud
Roberto Castro-Muñoz
Eliasse Zongo
HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
BMC Plant Biology
P. stewartii
Phytochemicals
Enzyme inhibition
GC-MS
HPLC
title HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
title_full HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
title_fullStr HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
title_full_unstemmed HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
title_short HPLC, GC-MS, XRD profiling, enzyme inhibition and cytotoxicity potential of Phlomis Stewartii extracts using response surface methodology
title_sort hplc gc ms xrd profiling enzyme inhibition and cytotoxicity potential of phlomis stewartii extracts using response surface methodology
topic P. stewartii
Phytochemicals
Enzyme inhibition
GC-MS
HPLC
url https://doi.org/10.1186/s12870-025-06997-7
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