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Fulltext Detection of Naegleria species in environmental samples from Peninsular Malaysia

Ithoi I, Ahmad AF, Nissapatorn V, Lau YL, Mahmud R, Mak JW

PLoS One, 2011;6(9):e24327.
PMID: 21915311 DOI: 10.1371/journal.pone.0024327

BACKGROUND: In Malaysia, researchers and medical practitioners are unfamiliar with Naegleria infections. Thus little is known about the existence of pathogenic Naegleria fowleri, and the resultant primary amoebic meningoencephalitis (PAM) is seldom included in the differential diagnosis of central nervous system infections. This study was conducted to detect the presence of Naegleria species in various environmental samples.
METHODS/FINDINGS: A total of 41 Naegleria-like isolates were isolated from water and dust samples. All these isolates were subjected to PCR using two primer sets designed from the ITS1-ITS2 regions. The N. fowleri species-specific primer set failed to produce the expected amplicon. The Naegleria genus-specific primers produced amplicons of 408 bp (35), 450 bp (2), 457 bp (2) or 381 bp (2) from all 41 isolates isolated from aquatic (33) and dust (8) samples. Analysis of the sequences from 10 representative isolates revealed that amplicons with fragments 408, 450 and 457 bp showed homology with non-pathogenic Naegleria species, and 381 bp showed homology with Vahlkampfia species. These results concurred with the morphological observation that all 39 isolates which exhibited flagella were Naegleria, while 2 isolates (AC7, JN034055 and AC8, JN034056) that did not exhibit flagella were Vahlkampfia species.
CONCLUSION: To date, pathogenic species of N. fowleri have not been isolated from Malaysia. All 39 isolates that produced amplicons (408, 450 and 457 bp) from the genus-specific primers were identified as being similar to nonpathogenic Naegleria. Amplicon 408 bp from 5 representative isolates showed 100% and 99.7% identity to Naegleria philippinensis isolate RJTM (AM167890) and is thus believed to be the most common species in our environment. Amplicons 450 bp and 457 bp were respectively believed to be from 2 new species of Naegleria, since representative isolates showed lower homology and had a longer base pair length when compared to the reference species in the Genbank, Naegleria schusteri (AJ566626) and Naegleria laresi (AJ566630), respectively.
Fulltext The Authentication and Grading of Edible Bird's Nest by Metabolite, Nutritional, and Mineral Profiling

Mohamad Ibrahim R, Mohamad Nasir NN, Abu Bakar MZ, Mahmud R, Ab Razak NA

Foods, 2021 Jul 07;10(7).
PMID: 34359444 DOI: 10.3390/foods10071574

Edible bird's nest (EBN) produced by Aerodramus fuciphagus has a high demand for nutritional and medicinal application throughout the world. The present study was to evaluate the authentication of a man-made house EBN, which are half cup and stripe-shaped by FTIR. Next, both samples were compared according to their metabolite, nutritional, and mineral composition. The results indicated that the FTIR spectra of both EBN samples were identical and similar to the reference, suggesting the authenticity of the EBN used. The metabolites that contribute to the possible medicinal properties of EBN were found by using GC-MS. The results of the proximate analysis, followed by the standard AOAC method, inferred that both EBN shapes to be rich in crude protein and carbohydrate contents. However, the proximate composition between the half cup and stripe-shaped EBN showed significant differences. Major mineral elements detected were calcium and sodium, and magnesium contents were significantly different between both EBN. Additionally, the half cup and stripe-shaped EBN had a low level of heavy metal content than the maximum regulatory limit as set by the Malaysian Food Act 1983. This study concludes that the nutritional composition varied between the samples and thus suggests that nutrient content should be considered as criteria for the grading requirement of commercialized EBN.
Fulltext An assessment of plantar fascia with ultrasound findings in patients with plantar fasciitis: a systematic review

Khammas ASA, Mahmud R, Hassan HA, Ibrahim I, Mohammed SS

J Ultrasound, 2023 Mar;26(1):13-38.
PMID: 36040577 DOI: 10.1007/s40477-022-00712-0

INTRODUCTION: Plantar fasciitis (PFS) is described by an intense pain over medial tubercle of calcaneus, increased with the first step after waking up, after rest and during weightbearing activity. It is the most common cause of plantar heel pain in adults with the prevalence estimated 10% of the general population. Ultrasound imaging is commonly being used to measure the PF thickness, evaluate the efficacy of different treatments and a guide therapeutic technique in patients with PFS. The objective of this study was to systematically review the studies that were previously published to evaluate the role of ultrasound in the assessment of PF in patients with PFS.
METHODS: A systematic search was carried out over the last 5 years from 2017 to 2022 on basis the following electronic databases: Science Direct, Scopus, Web of Science, Springer and PubMed. The keywords that used in the searching were: ultrasound, sonography, ultrasonography, plantar fasciitis, imaging of plantar fascia, physiotherapy of plantar fasciitis, interventional treatment of plantar fasciitis, randomized controlled trial of plantar fasciitis and interventional ultrasound. The review focused on the assessment of PF in patients with PFS underwent different interventions using B-mode, shear wave elastography (SWE) and color Doppler ultrasound.
RESULTS: During the search process, 1661 were recorded using the proper keywords from 2017 to 2022 in which 666 original articles were found after removing the review and duplicated articles. Of these, thirty articles met the inclusion criteria and included in this review. The articles have assessed the PF in patients with PFS under different conditions using different ultrasound modes. Twenty-six articles evaluated the effectiveness of different treatment on PF in patients with PFS using different ultrasound modes. In 8 of 26 articles, the ultrasound was used as both an assessment tool of PF and guide therapeutic technique in patients with PFS. In 18 articles, the ultrasound was used as only assessment tool to identify the PF thickness and its observation changes in patients with PFS. Four articles compared the PF thickness and its intrafascial changes between patients with PFS and healthy subjects.
CONCLUSION: The ultrasound can be a reliable tool in assessment the effect of different interventions on PF by evaluating its thickness, echogenicity and stiffness changes in patients with PFS. There were different methods and treatments were used among the studies.
Fulltext Determinants of quality of life among Malaysian cancer patients: a cross-sectional study

Akhtari-Zavare M, Mohd-Sidik S, Periasamy U, Rampal L, Fadhilah SI, Mahmud R

Health Qual Life Outcomes, 2018 Aug 13;16(1):163.
PMID: 30103759 DOI: 10.1186/s12955-018-0989-5

BACKGROUND: Cancer is a serious public health problem not only in Malaysia, also worldwide. The aim of this study was to determine the determinants of quality of life (QOL) among cancer patients in Peninsular Malaysia.
METHODS: A cross sectional study was conducted among 2120 cancer patients in Peninsular Malaysia, between April 2016 to January 2017. All cancer patients aged 18 years old and above, Malaysian citizens and undergoing cancer treatment at government hospitals were approached to participate in this study and requested to complete a set of validated questionnaires. Inferential statistical tests such as t-test and one-way ANOVA were used to determine the differences between demographic variables, physical effects, clinical factors, psychological effects and self-esteem with the quality of life of cancer patients. Predictor(s) of quality of life were determined by using Multivariate linear regression models.
RESULT: A total 1620 out of 2120 cancer patients participated in this study, giving a response rate of 92%. The majority of cancer patients were female 922 (56.9%), Malays 1031 (63.6%), Muslim 1031 (63.6%), received chemotherapy treatment 1483 (91.5%). Overall, 1138 (70.2%) of the patients had depression and 1500 (92.6%) had anxiety. Statistically significant associations were found between QOL and clinical factors, physical side effects of cancer, psychological effects and self-esteem (p
Fulltext Protective Effects of Nigella sativa on Metabolic Syndrome in Menopausal Women

Ibrahim RM, Hamdan NS, Ismail M, Saini SM, Abd Rashid SN, Abd Latiff L, et al.

Adv Pharm Bull, 2014;4(1):29-33.
PMID: 24409406 DOI: 10.5681/apb.2014.005

PURPOSE: This study was conducted in menopausal women to determine the metabolic impact of Nigella sativa.
METHODS: Thirty subjects who were menopausal women within the age limit of 45-60 were participated in this study and randomly allotted into two experimental groups. The treatment group was orally administered with N. sativa seeds powder in the form of capsules at a dose of 1g per day after breakfast for period of two months and compared to control group given placebo. Anthropometric and biochemical parameters were measured at baseline, 1st month, 2nd month and a month after treatment completed to determine their body weight, serum lipid profile and fasting blood glucose (FBG).
RESULTS: The treatment group showed slight reduction with no significant difference in body weight changes of the respondents. However, significant (p<0.05) improvement was observed in total cholesterol (TC), triglycerides (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), and blood glucose (p<0.05).
CONCLUSION: These results suggested that treatment with N. sativa exert a protective effect by improving lipid profile and blood glucose which are in higher risk to be elevated during menopausal period.
Fulltext Antimalarial activity of methanolic leaf extract of Piper betle L

Al-Adhroey AH, Nor ZM, Al-Mekhlafi HM, Amran AA, Mahmud R

Molecules, 2010 Dec 28;16(1):107-18.
PMID: 21189459 DOI: 10.3390/molecules16010107

The need for new compounds active against malaria parasites is made more urgent by the rapid spread of drug-resistance to available antimalarial drugs. The crude methanol extract of Piper betle leaves (50-400 mg/kg) was investigated for its antimalarial activity against Plasmodium berghei (NK65) during early and established infections. The phytochemical and antioxidant potentials of the crude extract were evaluated to elucidate the possibilities of its antimalarial effects. The safety of the extract was also investigated in ICR mice of both sexes by the acute oral toxicity limit test. The leaf extract demonstrated significant (P < 0.05) schizonticidal activity in all three antimalarial evaluation models. Phytochemical screening showed that the leaf extract contains some vital antiplasmodial chemical constituents. The extract also exhibited a potent ability to scavenge the free radicals. The results of acute toxicity showed that the methanol extract of Piper betle leaves is toxicologically safe by oral administration. The results suggest that the Malaysian folklorical medicinal application of the extract of Piper betle leaf has a pharmacological basis.
Fulltext Hexaplex PCR detection system for identification of five human Plasmodium species with an internal control

Chew CH, Lim YA, Lee PC, Mahmud R, Chua KH

J Clin Microbiol, 2012 Dec;50(12):4012-9.
PMID: 23035191 DOI: 10.1128/JCM.06454-11

Malaria remains one of the major killers of humankind and persists to threaten the lives of more than one-third of the world's population. Given that human malaria can now be caused by five species of Plasmodium, i.e., Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, Plasmodium ovale, and the recently included Plasmodium knowlesi, there is a critical need not only to augment global health efforts in malaria control but also, more importantly, to develop a rapid, accurate, species-sensitive/species-specific, and economically effective diagnostic method for malaria caused by these five species. Therefore, in the present study, a straightforward single-step hexaplex PCR system targeting five human Plasmodium 18S small-subunit rRNAs (ssu rRNAs) was designed, and the system successfully detected all five human malaria parasites. In addition, this system enables the differentiation of single infection as well as mixed infections up to the two-species level. This assay was validated with 50 randomly blinded test and 184 clinical samples suspected to indicate malaria. This hexaplex PCR system is not only an ideal alternative for routine malaria diagnosis in laboratories with conventional PCR machines but also adds value to diagnoses when there is a lack of an experienced microscopist or/and when the parasite morphology is confusing. Indeed, this system will definitely enhance the accuracy and accelerate the speed in the diagnosis of malaria, as well as improve the efficacy of malaria treatment and control, in addition to providing reliable data from epidemiological surveillance studies.
Fulltext Code-mixing unveiled: Enhancing the hate speech detection in Arabic dialect tweets using machine learning models

Alhazmi A, Mahmud R, Idris N, Mohamed Abo ME, Eke CI

PLoS One, 2024;19(7):e0305657.
PMID: 39018339 DOI: 10.1371/journal.pone.0305657

Technological developments over the past few decades have changed the way people communicate, with platforms like social media and blogs becoming vital channels for international conversation. Even though hate speech is vigorously suppressed on social media, it is still a concern that needs to be constantly recognized and observed. The Arabic language poses particular difficulties in the detection of hate speech, despite the considerable efforts made in this area for English-language social media content. Arabic calls for particular consideration when it comes to hate speech detection because of its many dialects and linguistic nuances. Another degree of complication is added by the widespread practice of "code-mixing," in which users merge various languages smoothly. Recognizing this research vacuum, the study aims to close it by examining how well machine learning models containing variation features can detect hate speech, especially when it comes to Arabic tweets featuring code-mixing. Therefore, the objective of this study is to assess and compare the effectiveness of different features and machine learning models for hate speech detection on Arabic hate speech and code-mixing hate speech datasets. To achieve the objectives, the methodology used includes data collection, data pre-processing, feature extraction, the construction of classification models, and the evaluation of the constructed classification models. The findings from the analysis revealed that the TF-IDF feature, when employed with the SGD model, attained the highest accuracy, reaching 98.21%. Subsequently, these results were contrasted with outcomes from three existing studies, and the proposed method outperformed them, underscoring the significance of the proposed method. Consequently, our study carries practical implications and serves as a foundational exploration in the realm of automated hate speech detection in text.
Fulltext Effects of Counselling on Quality of Life among Cancer Patients in Malaysia: A Randomized Controlled Trial

Periasamy U, Mohd-Sidik S, Akhtari-Zavare M, Rampal L, Ismail SIF, Mahmud R

Iran J Public Health, 2020 Oct;49(10):1902-1911.
PMID: 33346212 DOI: 10.18502/ijph.v49i10.4693

Background: We aimed to assess whether "Managing Patients on Chemotherapy" book is effective to improve quality of life (QOL) of cancer patient via counselling by pharmacist.
Methods: A randomized control trial study was run among 2120 cancer patients in public hospitals in Peninsular Malaysia, from Apr 2016-Jan 2018. The treatment group received counselling regarding chemotherapy by using developed module. The data were collected at three time-points: baseline, 1st, 2nd and 3rd follow-ups after counseling by Validated Malay version of the WHOQOL-BREF of questionnaire. Data analyses were done using χ2 and two-way repeated measure ANOVA.
Results: The treatment group improved significantly as compared to control group in physical health, psychological health, social relationship, environment and overall QOL (P<0.00).
Conclusion: The "Managing Patients on Chemotherapy" book along with repetitive counselling by pharmacists is a useful intervention for improving QOL of cancer patients undergoing treatment.
Sonographic and clinical effects of manual physical therapy for plantar fasciitis: randomized prospective controlled trial

Khammas ASA, Mahmud R, Hassan HA, Ibrahim I, Mohammed SS

J Ultrasound, 2024 Sep;27(3):487-500.
PMID: 37917407 DOI: 10.1007/s40477-023-00833-0

OBJECTIVE: This study aimed to evaluate the efficacy of manual physiotherapy on clinical outcomes, morphology of plantar fascia (PF), thicknesses of calcaneal fat pad (CFP) and Kager's fat pad (KFP) with ultrasound imaging in plantar fasciitis (PFS) patients. Also, to evaluate the PF thickness, pain and foot functional outcomes among PFS phases.
METHODS: A randomized controlled trial was conducted on 122 subjects divided into three groups: group A (40 patients with PFS) underwent manual physiotherapy, group B (42 patients with PFS) without any intervention and group C (40 healthy subjects) were matched by age, gender and BMI with each patient in group A and B. The following outcomes were evaluated at baseline and one-month of follow-ups: morphology of PF and thicknesses of CFP and KFP, pain, foot functional limitation.
RESULTS: PF thickness was significantly thickened in group A and B compared to group C (P
Anisakiasis causing acute dysentery in Malaysia

Amir A, Ngui R, Ismail WH, Wong KT, Ong JS, Lim YA, et al.

Am J Trop Med Hyg, 2016 Aug 03;95(2):410-2.
PMID: 27325803 DOI: 10.4269/ajtmh.16-0007

Human anisakiasis is a zoonosis acquired by eating raw or undercooked infected seafood. Herein, we report a case of acute dysentery caused by anisakiasis in a 64-year-old man in Malaysia. A colonoscopy was performed and a nematode larva was found penetrating the mucosa of the ascending colon. Bleeding was observed at the site of penetration. Y-shaped lateral epidermal cords were seen from the cross section of the worm, which is a prominent feature of Anisakis larva. Molecular analysis using polymerase chain reaction of cytochrome oxidase 2 (cox2) gene confirmed the specimen to be larva of Anisakis simplex.
Fulltext High seroprevalence of echinococossis, schistosomiasis and toxoplasmosis among the populations in Babati and Monduli districts, Tanzania

Khan MB, Sonaimuthu P, Lau YL, Al-Mekhlafi HM, Mahmud R, Kavana N, et al.

Parasit Vectors, 2014;7:505.
PMID: 25388913 DOI: 10.1186/s13071-014-0505-7

The neglected tropical diseases, echinococcosis, schistosomiasis and toxoplasmosis are all globally widespread zoonotic diseases with potentially harmful consequences. There is very limited data available on the prevalence of these infections, except for schistosmiasis, in underdeveloped countries. This study aimed to determine the seroprevalence of Echinococcus multilocularis, Schistosoma mansoni, and Toxoplasma gondii antibodies in populations from the Monduli and Babati districts in Tanzania.
Fulltext Developing and evaluating health education learning package (HELP) to control soil-transmitted helminth infections among Orang Asli children in Malaysia

Al-Delaimy AK, Al-Mekhlafi HM, Lim YA, Nasr NA, Sady H, Atroosh WM, et al.

Parasit Vectors, 2014;7:416.
PMID: 25179100 DOI: 10.1186/1756-3305-7-416

This study was carried out to develop a health education learning package (HELP) about soil-transmitted helminth (STH) infections, and to evaluate what impact such a package could have in terms of reducing the incidence and intensity of STH infections among Orang Asli schoolchildren in Pahang, Malaysia.
Fulltext High proportion of knowlesi malaria in recent malaria cases in Malaysia

Yusof R, Lau YL, Mahmud R, Fong MY, Jelip J, Ngian HU, et al.

Malar J, 2014;13:168.
PMID: 24886266 DOI: 10.1186/1475-2875-13-168

Plasmodium knowlesi is a simian parasite that has been recognized as the fifth species causing human malaria. Naturally-acquired P. knowlesi infection is widespread among human populations in Southeast Asia. The aim of this epidemiological study was to determine the incidence and distribution of malaria parasites, with a particular focus on human P. knowlesi infection in Malaysia.
Mapping and modelling the geographical distribution of soil-transmitted helminthiases in Peninsular Malaysia: implications for control approaches

Ngui R, Shafie A, Chua KH, Mistam MS, Al-Mekhlafi HM, Sulaiman WW, et al.

Geospat Health, 2014 May;8(2):365-76.
PMID: 24893014

Soil-transmitted helminth (STH) infections in Malaysia are still highly prevalent, especially in rural and remote communities. Complete estimations of the total disease burden in the country has not been performed, since available data are not easily accessible in the public domain. The current study utilised geographical information system (GIS) to collate and map the distribution of STH infections from available empirical survey data in Peninsular Malaysia, highlighting areas where information is lacking. The assembled database, comprising surveys conducted between 1970 and 2012 in 99 different locations, represents one of the most comprehensive compilations of STH infections in the country. It was found that the geographical distribution of STH varies considerably with no clear pattern across the surveyed locations. Our attempt to generate predictive risk maps of STH infections on the basis of ecological limits such as climate and other environmental factors shows that the prevalence of Ascaris lumbricoides is low along the western coast and the southern part of the country, whilst the prevalence is high in the central plains and in the North. In the present study, we demonstrate that GIS can play an important role in providing data for the implementation of sustainable and effective STH control programmes to policy-makers and authorities in charge.
Fulltext A randomised controlled trial on hypolipidemic effects of Nigella Sativa seeds powder in menopausal women

Ibrahim RM, Hamdan NS, Mahmud R, Imam MU, Saini SM, Rashid SN, et al.

J Transl Med, 2014;12:82.
PMID: 24685020 DOI: 10.1186/1479-5876-12-82

The risk of cardiovascular diseases (CVD) is increased tremendously among menopausal women, and there is an increasing demand for alternative therapies for managing factors like dyslipidemia that contribute to CVD development.
Fulltext Malaysian child infected with Plasmodium vivax via blood transfusion: a case report

Anthony CN, Lau YL, Sum JS, Fong MY, Ariffin H, Zaw WL, et al.

Malar J, 2013;12:308.
PMID: 24007496 DOI: 10.1186/1475-2875-12-308

Malaria may be a serious complication of blood transfusion due to the asymptomatic persistence of parasites in some donors. This case report highlights the transfusion-transmitted malaria of Plasmodium vivax in a child diagnosed with germ cell tumour. This child had received blood transfusion from three donors and a week later started developing malaria like symptoms. Nested PCR and sequencing confirmed that one of the three donors was infected with P. vivax and this was transmitted to the 12-year-old child. To the best of the authors' knowledge, this is the first reported transfusion-transmitted malaria case in Malaysia.
Multilocus genotyping of Giardia duodenalis in Malaysia

Huey CS, Mahdy MA, Al-Mekhlafi HM, Nasr NA, Lim YA, Mahmud R, et al.

Infect Genet Evol, 2013 Jul;17:269-76.
PMID: 23624189 DOI: 10.1016/j.meegid.2013.04.013

Giardia duodenalis is considered the most common intestinal parasite in humans worldwide. In Malaysia, many studies have been conducted on the epidemiology of giardiasis. However, there is a scarcity of information on the genetic diversity and the dynamics of transmission of G. duodenalis. The present study was conducted to identify G. duodenalis assemblages and sub-assemblages based on multilocus analysis of the glutamate dehydrogenase (gdh), beta-giardin (bg) and triose phosphate isomerase (tpi) genes. Faecal specimens were collected from 484 Orang Asli children with a mean age of 7 years and examined using light microscopy. Specimens positive for Giardia were subjected to PCR analysis of the three genes and subsequent sequencing in both directions. Sequences were edited and analysed by phylogenetic analysis. G. duodenalis was detected in 17% (84 of 484) of the examined specimens. Among them, 71 were successfully sequenced using at least one locus. Genotyping results showed that 30 (42%) of the isolates belonged to assemblage A, 32 (45%) belonged to assemblage B, while discordant genotype results were observed in 9 specimens. Mixed infections were detected in 43 specimens using a tpi-based assemblage specific protocol. At the sub-assemblages level, isolates belonged to assemblage A were AII. High nucleotide variation found in isolates of assemblage B made subtyping difficult to achieve. The finding of assemblage B and the anthroponotic genotype AII implicates human-to-human transmission as the most possible mode of transmission among Malaysian aborigines. The high polymorphism found in isolates of assemblage B warrants a more defining tool to discriminate assemblage B at the sub-assemblage level.
Fulltext Hyperparasitaemic human Plasmodium knowlesi infection with atypical morphology in peninsular Malaysia

Lee WC, Chin PW, Lau YL, Chin LC, Fong MY, Yap CJ, et al.

Malar J, 2013;12:88.
PMID: 23496970 DOI: 10.1186/1475-2875-12-88

Plasmodium knowlesi is a potentially life-threatening zoonotic malaria parasite due to its relatively short erythrocytic cycle. Microscopic identification of P. knowlesi is difficult, with "compacted parasite cytoplasm" being one of the important identifying keys. This report is about a case of hyperparasitaemic human P. knowlesi infection (27% parasitaemia) with atypical amoeboid morphology. A peninsular Malaysian was admitted to the hospital with malaria. He suffered anaemia and acute kidney function impairment. Microscopic examination, assisted by nested PCR and sequencing confirmed as P. knowlesi infection. With anti-malarial treatment and several medical interventions, patient survived and recovered. One-month medical follow-up was performed after recovery and no recrudescence was noted. This case report highlights the extreme hyperparasitaemic setting, the atypical morphology of P. knowlesi in the patient's erythrocytes, as well as the medical interventions involved in this successfully treated case.
Fulltext A simple isocratic HPLC method for the simultaneous determination of sinensetin, eupatorin, and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone in Orthosiphon stamineus extracts

Yam MF, Mohamed EA, Ang LF, Pei L, Darwis Y, Mahmud R, et al.

J Acupunct Meridian Stud, 2012 Aug;5(4):176-82.
PMID: 22898066 DOI: 10.1016/j.jams.2012.05.005

Orthosiphon stamineus extracts contain three flavonoids (3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin) as bioactive substances. Previous reported high performance liquid chromatography- ultraviolet (HPLC-UV) methods for the determination of these flavonoids have several disadvantages, including unsatisfactory separation times and not being well validated according to International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) standard guidelines. A rapid, specific reversed-phase HPLC method with isocratic elution of acetonitrile: isopropyl alcohol: 20mM phosphate buffer (NaH(2)PO(4)) (30:15:55, v/v) (pH 3.5) at a flow-rate of 1ml/minute, a column temperature of 25°C, and ultraviolet (UV) detection at 340 nm was developed. The method was validated and applied for quantification of different types of O stamineus extracts and fractions. The method allowed simultaneous determination of 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin in the concentration range of 0.03052-250 μg/ml. The limits of detection and quantification, respectively, were 0.0076 and 0.061 μg/ml for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, 0.0153 and 0.122 μg/ml for sinensetin and 0.0305 and 0.122 μg/ml for eupatorin. The percent relative standard deviation (% RSD) values for intraday were 0.048-0.368, 0.025-0.135, and 0.05-0.476 for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin, respectively, and those for intraday precision were 0.333-1.688, 0.722-1.055, and 0.548-1.819, respectively. The accuracy for intraday were 91.25%-103.38%, 94.32%-109.56%, and 92.85%-109.70% for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin, respectively, and those for interday accuracy were 97.49%-103.92%, 103.58%-104.57%, and 103.9%-105.33%, respectively. The method was found to be simple, accurate and precise and is recommended for routine quality control analysis of O stamineus extract containing the three flavonoids as the principle components in the extract.

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