Quality assessment of common anti-malarial medicines marketed in Gambella, National Regional State, South Western-Ethiopia.

BACKGROUND: Over the past years, there has been a growing concern that a considerable amount of anti-malarial supply in the underdeveloped world particularly in the private sector, is of poor quality. The World Health Organization (WHO) has received about 1500 reports that mentions instances of substandard and falsified products since 2013. The majority of the reports concerned antibiotics and anti-malarials. The majority of reports (42%) originate from the WHO African region. OBJECTIVE: This study intends to assess the quality of the most widely used anti-malarial medications [artemether-lumefantrine tablets, chloroquine phosphate tablets, primaquine phosphate tablets, artesunate, and artemether injections] in Gambella, South-West, Ethiopia. METHODS: A total of 52 samples were collected on June 2022 from Gambella National Regional State, Ethiopia. Half of the districts (six) located in the four zones of the region were chosen using simple random sampling technique. All drug retail outlets available in the selected districts (locally known as woredas) were included. The samples were subjected to visual inspection with a tool adopted from the joint WHO/FIP/ USP checklist. The pharmacopeial tests for identification, uniformity of dosage forms, assay, thickness, diameter, hardness, friability, disintegration test, dissolution, and sterility tests were carried out according to the USP 44-NF 39 and International Pharmacopoeia 11th edition, 2022 monographs. RESULTS AND DISCUSSION: Only 25% of the samples were registered on the Ethiopian Food and Drug Authority (EFDA's) electronic regulatory/ registration system (ERIS). Besides, 88.8% of artemether injection products were presented in clear glass ampoules. This might expose the products to photochemical degradation that leads to in loss of anti-plasmodial activity. In addition, 50% of the artemether products assessed were not bioequivalent with the comparator product in the in vitro dissolution comparison tests. Overall, the study findings reveal a high prevalence (58.3%) of substandard anti-malarial drugs in the region. The stated percent of the samples had failed in one or more of the quality test parameters assessed in this study. CONCLUSION: The study findings reveal a high prevalence (58.3%) of substandard anti-malarial drugs in the region. Only a quarter were registered and 38% of the unregistered products failed the quality tests. Hence, the national, regional medicine regulatory bodies and other stake holders should perform the required roles to circumvent presence of Substandard and Falsified (SF) anti-malarial drugs in the study sites.

Pharmacopeial quality of artemether-lumefantrine anti-malarial agents in Uganda.

BACKGROUND: Substandard anti-malarial agents pose a significant challenge to effective malaria control and elimination efforts especially in sub-Saharan Africa. The quality of anti-malarials in most low-and-middle income countries (LMICs) is affected by several factors including inadequate regulation and limited resources. In this study, the pharmacopeial quality of artemether-lumefantrine (AL) in low and high malaria transmission settings in Uganda was assessed. METHODS: This was a cross-sectional study conducted among randomly selected private drug outlets. The AL anti-malarials available in drug outlets were purchased using overt method. The samples were screened for quality using visual inspection, weight uniformity, content assay and dissolution tests. The assay test was done using liquid chromatography-mass spectrometry (LC-MS). The samples were considered substandard if the active pharmaceutical ingredient (API) content was outside 90-110% range of the label claim. Dissolution test was conducted following United States Pharmacopoeia (USP) method. Data was analysed using descriptive statistics and presented as means with standard deviations, frequencies, and proportions. Correlation between medicine quality and independent variables was determined using Fisher's exact test of independence at 95% level of significance. RESULTS: A total of 74 AL anti-malarial samples were purchased from high (49/74; 66.2%) and low (25/74; 33.8%) malaria transmission settings. The most common batch of AL was LONART, 32.4% (24/74), with 33.8% (25/74) being 'Green leaf'. Overall prevalence of substandard quality artemether-lumefantrine was 18.9% (14/74; 95% CI: 11.4-29.7). Substandard quality AL was significantly associated with setting (p = 0.002). A total of 10 samples (13.5%) failed artemether content assay test while, 4 samples (5.4%, 4/74) failed the lumefantrine assay test. One sample from a high malaria transmission setting failed both artemether and lumefantrine assay content test. Of the samples that failed artemether assay test, 90% had low (< 90%) artemether content. All the samples passed visual inspection and dissolution tests. CONCLUSION: Artemether-lumefantrine agents, the recommended first-line treatment for uncomplicated malaria with APIs outside the recommended pharmacopeial content assay limit is common especially in high malaria transmission settings. There is need for continuous surveillance and monitoring of the quality of artemisinin-based anti-malarials across the country by the drug regulatory agency.

Development of single- and multiplex immunoassays for rapid detection and quantitation of amodiaquine in ACT drugs and rat serum.

Amodiaquine (AQ) is a commonly used antimalarial drug, and N-desethyl-AQ (N-DEAQ) is an active metabolite of AQ. Given the significance of drug quality in the management of malaria cases, this study aims to develop antibody-based assays for the detection and quantitation of AQ without the need for sophisticated equipment. Two monoclonal antibodies (mAbs) against AQ, designated as JUN7 and TE7, were selected, which showed 72.7% and 9.5% cross-reactivity to N-DEAQ, respectively. These mAbs showed <0.1% cross-reactivity to other commonly used antimalarial drugs. An indirect competitive enzyme-linked immunosorbent assay (icELISA) based on JUN7 showed a 50% inhibitory concentration (IC50) of 0.16 ng/mL and a working range of 0.06-0.46 ng/mL. A lateral flow immunoassay (LFIA) based on JUN7 was also developed with a working range of 2.58-30.86 ng/mL. The icELISA and LFIA were applied for the quantification of AQ in commercial drugs, and the results were comparable to those determined using high-performance liquid chromatography. In addition, a combination dipstick for simultaneous, qualitative analysis of AQ and artesunate was developed. All immunoassays based on JUN7 can be applied for quality control of AQ-containing artemisinin-based combination therapies. As TE7 showed low cross-reactivity to N-DEAQ, an icELISA based on TE7 was developed with an IC50 of 0.38 ng/mL and a working range of 0.14-1.67 ng/mL. The TE7 icELISA was applied for the study of pharmacokinetics of AQ in rat serum after intragastric administration, and the results were consistent with those of previous studies.

Quantification of Tafenoquine and 5,6-Orthoquinone Tafenoquine by UHPLC-MS/MS in Blood, Plasma, and Urine, and Application to a Pharmacokinetic Study.

Analytical methods for the quantification of the new 8-aminoquinoline antimalarial tafenoquine (TQ) in human blood, plasma and urine, and the 5,6-orthoquinone tafenoquine metabolite (5,6-OQTQ) in human plasma and urine have been validated. The procedure involved acetonitrile extraction of samples followed by ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). Chromatography was performed using a Waters Atlantis T3 column with a gradient of 0.1% formic acid and acetonitrile at a flow rate of 0.5 mL per minute for blood and plasma. Urine analysis was the same but with methanol containing 0.1% formic acid replacing acetonitrile mobile phase. The calibration range for TQ and 5,6-OQTQ in plasma was 1 to 1200 ng/mL, and in urine was 10 to 1000 ng/mL. Blood calibration range for TQ was 1 to 1200 ng/mL. Blood could not be validated for 5,6-OQTQ due to significant signal suppression. The inter-assay precision (coefficient of variation %) was 9.9% for TQ at 1 ng/mL in blood (n = 14) and 8.2% for TQ and 7.1% for 5,6-OQTQ at 1 ng/mL in plasma (n = 14). For urine, the inter-assay precision was 8.2% for TQ and 6.4% for 5,6-OQTQ at 10 ng/mL (n = 14). TQ and 5,6-OQTQ are stable in blood, plasma and urine for at least three months at both -80 °C and -20 °C. Once validated, the analytical methods were applied to samples collected from healthy volunteers who were experimentally infected with Plasmodium falciparum to evaluate the blood stage antimalarial activity of TQ and to determine the therapeutic dose estimates for TQ, the full details of which will be published elsewhere. In this study, the measurement of TQ and 5,6-OQTQ concentrations in samples from one of the four cohorts of participants is reported. Interestingly, TQ urine concentrations were proportional to parasite recrudescence times post dosing To our knowledge, this is the first description of a fully validated method for the measurement of TQ and 5,6-OQTQ quantification in urine.

Development of a dissolution method for lumefantrine and artemether in immediate release fixed dose artemether/lumefantrine tablets.

BACKGROUND: Dissolution of artemether (ART) and lumefantrine (LUM) active pharmaceutical ingredients (APIs) in fixed dose combination (FDC) ART/LUM tablets is one of the critical quality attributes. Thus, the verification of the release profile of ART and LUM from FDC ART/LUM tablets using a robust and discriminatory dissolution method is crucial. Therefore, the aim of this study was to develop and validate an appropriate dissolution method for quality control of FDC ART/LUM tablets. METHODS: The dissolution medium was selected based on saturation solubility data and sink conditions. The effect of agitation speed, pH and surfactant concentration on the release of ART and LUM was evaluated by employing a two-level factorial experiment. The resulting final method was validated for linearity, precision, robustness and API stability. In addition, the discriminatory power of the method was evaluated using expired and unexpired FDC ART/LUM products. RESULTS: A suitable dissolution profile of FDC ART/LUM tablets was obtained in 900 ml HCl (0.025 N, pH 1.6) with 1%Myrj 52 using paddle method at 100 rpm and 37 °C. ART and LUM were analysed using a HPLC method with UV detection at wavelengths of 210 and 335 nm, respectively. The results from the stability study showed that ART and LUM were sufficiently stable in HCl (0.025 N, pH 1.6) with 1%Myrj 52 at 37 °C. The method was linear (r2 = 0.999) over the concentration range of 6.25-100 µg/ml. The results for precision were within the acceptance limit (%RSD < 2). The percent relative standard deviation (< 2%) and statistically non-significant (p > 0.05) difference in release of ART and LUM observed between deliberately changed dissolution method settings (pH = 1.6 ± 0.2 or agitation speed = 100 ± 2) and optimized dissolution conditions revealed the robustness of the dissolution method. The method was capable to discriminate among different FDC ART/LUM products with different quality. CONCLUSIONS: The developed dissolution method is robust and discriminatory. It can be used in the quality evaluation of FDC ART/LUM tablets.

Green Analytical Methods of Antimalarial Artemether-Lumefantrine Analysis for Falsification Detection Using a Low-Cost Handled NIR Spectrometer with DD-SIMCA and Drug Quantification by HPLC.

Two green analytical approaches have been developed for the analysis of antimalarial fixed dose tablets of artemether and lumefantrine for quality control. The first approach consisted of investigating the qualitative performance of a low-cost handheld near-infrared spectrometer in combination with the principal component analysis as an exploratory tool to identify trends, similarities, and differences between pharmaceutical samples, before applying the data driven soft independent modeling of class analogy (DD-SIMCA) as a one-class classifier for proper drug falsification detection with 100% of both sensitivity and specificity in the studied cases. Despite its limited spectral range and low resolution, the handheld device allowed detecting falsified drugs with no active pharmaceutical ingredient and identifying specifically a pharmaceutical tablet brand name. The second approach was the quantitative analysis based on the green and fast RP-HPLC technique using ethanol as a green organic solvent and acetic acid as a green pH modifier. The optimal separation was achieved in 7 min using a mobile phase composed of ethanol 96% and 10 mM of acetic acid pH 3.35 (63:37, v/v). The developed method was validated according to the total error approach based on an accuracy profile, was applied to the analysis of tablets, and allowed confirming falsified drugs detected by spectroscopy.

Usefulness of combined screening methods for rapid detection of falsified and/or substandard medicines in the absence of a confirmatory method.

BACKGROUND: The influx of substandard and falsified medicines is a global public health challenge and its rapid detection is a key solution to the menace. This study used three screening methods and one confirmatory method for the quality assessment of 25 batches of artemether/lumefantrine dosage forms from the Ghanaian market to test that combined screening methods only can rapidly detect substandard and/or falsified medicines in areas where confirmatory methods may not be available. METHODS: The quality of artemether/lumefantrine tablet products obtained from pharmacies and licensed chemical seller shops within the Accra metropolis in Ghana were analysed using three screening methods (GPHF Minilab, Colorimetry and Counterfeit Drug Indicator) and one confirmatory method (high-performance liquid chromatography). RESULTS: The results showed that 18/25 batches of the artemether/lumefantrine samples passed using the combined screening and confirmatory methods and 5/25 batches of the artemether/lumefantrine samples failed using the combined screening and confirmatory methods. However, 1/25 batch of the artemether/lumefantrine samples failed using the combined screening methods but passed using the confirmatory method. Also, 1/25 batch of the artemether/lumefantrine samples passed using the combined screening methods but failed using the confirmatory method. This notwithstanding, the combined screening methods and the confirmatory method provided equivalent quality assessment profiles for 23/25 (92%) batches of the artemether/lumefantrine tablet products. Out of the 6 samples that failed the confirmatory test, 1/6, 2/6, and 3/6 failed on the high (> 110%), low (< 90%), and no active ingredient (0%), respectively. The sensitivity of Minilab, colorimetric, CoDI, and the combined screening methods at 95% confidence level were 0.5 ± 0.57, 0.83 ± 0.33, 0.75 ± 0.49, and 0.83 ± 0.33, respectively. Also, the specificity of Minilab, colorimetric, CoDI, and the combined screening methods at 95% confidence level were 1.00, 0.95 ± 0.10, 1.00, and 0.95 ± 0.10, respectively. CONCLUSION: The combined screening methods may be used for rapid detection of falsified and/or substandard medicines without using a confirmatory method. However, additional research on the best combinations of screening devices/methods to rapidly detect the quality of medicines is recommended.

Quality of fixed dose artemether/lumefantrine products in Jimma Zone, Ethiopia.

BACKGROUND: Malaria caused by Plasmodium vivax and Plasmodium falciparum is among the major public health problems in most endemic areas of the world. Artemisinin-based combination therapy (ACT) has been recommended as a first-line treatment for uncomplicated Plasmodium falciparum malaria almost in all endemic regions. Since ineffectively regulated medicines in resource limited settings could favour infiltration of poor quality anti-malarial medicines into pharmaceutical supply chain and jeopardize a positive treatment outcome, regular monitoring of the quality of anti-malarial medicines is critical. Thus, the aim of this study was to assess the quality of fixed dose combination (FDC) artemether (ART)/lumefantrine (LUM) tablets available in Jimma zone, Ethiopia. METHODS: This study was conducted in Jimma zone, Ethiopia. A total of 74 samples of FDC ART/LUM (20 mg ART/120 mg LUM) tablets were collected from 27 public facilities. All samples were subjected to visual inspection and the relevant information was recorded. The samples were transported to Jimma University Laboratory of Drug Quality (JuLaDQ) and stored at ambient temperature (20 °C to 25 °C) until analysis. The Pharmacopoeial conform/non-conform methods and the risk-based Derringer's desirability function approach were employed to assess the pharmaceutical quality of the investigated products. RESULTS: The visual inspection results revealed that there were no signs of falsified in the investigated products. Identification test results of samples indicated that all samples contained the stated active pharmaceutical ingredients (APIs). The results of uniformity of mass indicated that all samples complied with International Pharmacopoeial specification limits. The assay results, expressed as percent label claim (%lc) of ART (89.8 to 108.8%, mean ± SD = 99.1 ± 3.9%) and LUM (90.0 to 111.9%, mean ± SD = 98.2 ± 3.8%) revealed that, all samples complied with International Pharmacopoeia acceptance specification limits (i.e. 90-110%lc), except one generic product (IPCA Laboratories Ltd., India) which contains excessive LUM (111.9 ± 1.7%lc). The risk priority number (RPN) results revealed that assay (RPN = 392) is relatively the most critical quality attribute followed by identity (RPN = 280) and mass uniformity (40). Quality evaluation based on psycho-physical Harrington's scale revealed that more than 96% of samples were within the acceptable ranges (D ≥ 0.7-1.0). CONCLUSIONS: Both Pharmacopoeial and risk-based desirability function approaches to quality evaluation applied to the investigated products revealed that above 96% FDC ART/LUM tablets circulating in public settings of Jimma zone are of good quality.

Development of an agent-based model to assess the impact of substandard and falsified anti-malarials: Uganda case study.

BACKGROUND: Global efforts to address the burden of malaria have stagnated in recent years with malaria cases beginning to rise. Substandard and falsified anti-malarial treatments contribute to this stagnation. Poor quality anti-malarials directly affect health outcomes by increasing malaria morbidity and mortality, as well as threaten the effectiveness of treatment by contributing to artemisinin resistance. Research to assess the scope and impact of poor quality anti-malarials is essential to raise awareness and allocate resources to improve the quality of treatment. A probabilistic agent-based model was developed to provide country-specific estimates of the health and economic impact of poor quality anti-malarials on paediatric malaria. This paper presents the methodology and case study of the Substandard and Falsified Antimalarial Research Impact (SAFARI) model developed and applied to Uganda. RESULTS: The total annual economic impact of malaria in Ugandan children under age five was estimated at US$614 million. Among children who sought medical care, the total economic impact was estimated at $403 million, including $57.7 million in direct costs. Substandard and falsified anti-malarials were a significant contributor to this annual burden, accounting for $31 million (8% of care-seeking children) in total economic impact involving $5.2 million in direct costs. Further, 9% of malaria deaths relating to cases seeking treatment were attributable to poor quality anti-malarials. In the event of widespread artemisinin resistance in Uganda, we simulated a 12% yearly increase in costs associated with paediatric malaria cases that sought care, inflicting $48.5 million in additional economic impact annually. CONCLUSIONS: Improving the quality of treatment is essential to combat the burden of malaria and prevent the development of drug resistance. The SAFARI model provides country-specific estimates of the health and economic impact of substandard and falsified anti-malarials to inform governments, policy makers, donors and the malaria community about the threat posed by poor quality medicines. The model findings are useful to illustrate the significance of the issue and inform policy and interventions to improve medicinal quality.

Counterfeit and Substandard Test of the Antimalarial Tablet Riamet® by Means of Raman Hyperspectral Multicomponent Analysis.

The fight against counterfeit pharmaceuticals is a global issue of utmost importance, as failed medication results in millions of deaths every year. Particularly affected are antimalarial tablets. A very important issue is the identification of substandard tablets that do not contain the nominal amounts of the active pharmaceutical ingredient (API), and the differentiation between genuine products and products without any active ingredient or with a false active ingredient. This work presents a novel approach based on fiber-array based Raman hyperspectral imaging to qualify and quantify the antimalarial APIs lumefantrine and artemether directly and non-invasively in a tablet in a time-efficient way. The investigations were carried out with the antimalarial tablet Riamet® and self-made model tablets, which were used as examples of counterfeits and substandard. Partial least-squares regression modeling and density functional theory calculations were carried out for quantification of lumefantrine and artemether and for spectral band assignment. The most prominent differentiating vibrational signatures of the APIs were presented.

The quality of anti-malarial medicines in Embu County, Kenya.

BACKGROUND: Malaria is a major health problem in sub-Saharan Africa where over 90% of the world's malaria cases occur. Artemisinin-based combination therapy (ACT) is recommended by the World Health Organization as first-line and second-line treatments for uncomplicated falciparum malaria. However, there are a growing number of reports of sub-standard and falsified anti-malarial medicines in sub-Saharan Africa. METHODS: A cross-sectional study was conducted in Embu County, Kenya on the quality of anti-malarial medicines available in public and private facilities. Sampling of anti-malarial medicines from public and private hospitals, health centers and pharmacies was conducted between May and June 2014. Quality control tests were performed at the Drug Analysis and Research Unit, University of Nairobi, using ultraviolet spectrophotometry and high-performance liquid chromatography. A test for microbial load was also conducted for suspension formulations. RESULTS: A total of 39 samples were collected from public and private facilities across the Embu County. A visual inspection of the medicines showed no signs of sub-standard or falsification. All ACT passed identification, assay and dissolution tests. Of 11 suspension samples collected, none failed the microbial load test although one sample had 50 colony forming units (cfu). No oral artemisinin monotherapy medicines were encountered during the survey. Amodiaquine and chloroquine monotherapy products accounted for 5% of the collected samples, despite their ban in Kenya. Two herbal anti-malarial formulations were collected during the survey. Sulfadoxine/pyrimethamine (SP) was also found to be available use for malaria treatment, not in accordance with malaria treatment guidelines. CONCLUSION: All the anti-malarial drugs analysed in this study passed the quality control tests. This is encouraging given the high malaria burden in Kenya. Regulatory actions are required to counter SP and herbal products for malaria treatment.

A qualitative assessment of the challenges of WHO prequalification for anti-malarial drugs in China.

BACKGROUND: While China is a major manufacturer of artemisinin and its derivatives, it lags as a global leader in terms of the total export value of anti-malarial drugs as finished pharmaceutical products ready for marketing and use by patients. This may be due to the limited number of World Health Organization (WHO) prequalified anti-malarial drugs from China. Understanding the reasons for the slow progress of WHO prequalification (PQ) in China can help improve the current situation and may lead to greater efforts in malaria eradication by Chinese manufacturers. METHODS: In-depth interviews were conducted in China between November 2014 and December 2016. A total of 26 key informants from central government agencies, pharmaceutical companies, universities, and research institutes were interviewed, all of which had current or previous experience overseeing or implementing anti-malarial research and development in China. RESULTS: Chinese anti-malarial drugs that lack WHO PQ are mainly exported for use in the African private market. High upfront costs with unpredictable benefits, as well as limited information and limited technical support on WHO PQ, were reported as the main barriers to obtain WHO PQ for anti-malarial drugs by respondents from Chinese pharmaceutical companies. Potential incentives identified by respondents included tax relief, human resource training and consultation, as well as other incentives related to drug approval, such as China's Fast Track Channel. CONCLUSIONS: Government support, as well as innovative incentives and collaboration mechanisms are needed for further adoption of WHO PQ for anti-malarial drugs in China.

Influence of abiotic elicitors on improvement production of artemisinin in cell culture of Artemisia annua L.

A significant sesquiterpene lactone used as a drug is artemisinin. It is definitely an anti-parasitic drug isolated from field-grown Artemisia annua L. a plant from Asteraceae family. It is the best treatment for Plasmodium falciparum malaria. Unfortunately, artemisinin content in A. annua is extremely low (0.01-0.8% dry weight). So, some researchers focused on enhancing artemisinin content either in tissue/cell culture or the whole plant of A. annua sp. The aims of the current study were the effect of plant growth regulators on callus production and improvement of artemisinin content in cell suspension culture of A. annua, an alternative to the whole plant using abiotic elicitors. For callus induction, an experiment was laid out as a factorial experiment with three factors (explant type, different concentrations of BAP and 2,4-D) based on completely randomized design with three replications. The maximum frequency of callus induction (100%) was found in leaf explant on MS medium with a combination of 2, 4-D (3 mg/l) and BAP (1.5 mg/l). Therefore, the best calli were used for cell suspension culture and the effects of GA3 and ABA as abiotic elicitors were evaluated on the improvement of artemisinin production. The results indicated that both ABA and GA3 increased artemisinin content (2.02 fold and 1.67 fold in comparison to control respectively) in cell suspension culture.

Metabolite identification of the antimalarial naphthoquine using liquid chromatography-tandem high-resolution mass spectrometry in combination with multiple data-mining tools.

Naphthoquine (NQ) is one of important partner drugs of artemisinin-based combination therapy (ACT), which is recommended for the treatment of uncomplicated Plasmodium falciparum. NQ shows a high cure rate after a single oral administration. It is absorbed quickly (time to peak concentration 2-4 h) and has a long elimination half-life (255 h). However, the metabolism of NQ has not been clarified. In this work, the metabolite profiling of NQ was studied in six liver microsomal incubates (human, cynomolgus monkey, beagle dog, mini pig, rat and CD1 mouse), seven recombinant CYP enzymes (1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4) and rat (plasma, urine, bile and feces) using liquid chromatography tandem high-resolution LTQ-Orbitrap mass spectrometry (HRMSn ) in conjunction with online hydrogen/deuterium exchange. The biological samples were pretreated by protein precipitation and solid-phase extraction. For data processing, multiple data-mining tools were applied in tandem, i.e. background subtraction and followed by mass defect filter. NQ metabolites were characterized by accurate MS/MS fragmentation characteristics, the hydrogen/deuterium exchange data and cLogP simulation. As a result, five phase I metabolites (M1-M5) of NQ were characterized for the first time. Two metabolic pathways were involved: hydroxylation and N-oxidation. This study demonstrates that LC-HRMSn in combination with multiple data-mining tools in tandem can be a valuable analytical strategy for rapid metabolite profiling of drugs.

Plasmodium falciparum parasites overexpressing farnesyl diphosphate synthase/geranylgeranyl diphosphate synthase are more resistant to risedronate.

Farnesyl diphosphate synthase/geranylgeranyl diphosphate synthase (FPPS/GGPPS) is a key enzyme in the synthesis of isoprenic chains. Risedronate, a bisphosphonate containing nitrogen (N-BP), is a potent inhibitor of blood stage Plasmodium. Here, we show that P. falciparum parasites overexpressing FPPS/GGPPS are more resistant to risedronate, suggesting that this enzyme is an important target, and bisphosphonate analogues can be used as potential antimalarial drugs.

Anti-malarial medicine quality field studies and surveys: a systematic review of screening technologies used and reporting of findings.

BACKGROUND: Assessing the quality of medicines in low-middle income countries (LMICs) relies primarily on human inspection and screening technologies, where available. Field studies and surveys have frequently utilized screening tests to analyse medicines sampled at the point of care, such as health care facilities and medicine outlets, to provide a snap shot of medicine quality in a specific geographical area. This review presents an overview of the screening tests typically employed in surveys to assess anti-malarial medicine quality, summarizes the analytical methods used, how findings have been reported and proposes a reporting template for future studies. METHODS: A systematic search of the peer-reviewed and grey literature available in the public domain (including national and multi-national medicine quality surveys) covering the period 1990-2016 was undertaken. Studies were included if they had used screening techniques to assess the quality of anti-malarial medicines. As no standardized set of guidelines for the methodology and reporting of medicine quality surveys exist, the included studies were assessed for their standard against a newly proposed list of criteria. RESULTS: The titles and abstracts of 4621 records were screened and only 39 were found to meet the eligibility criteria. These 39 studies utilized visual inspection, disintegration, colorimetry and Thin Layer Chromatography (TLC) either as components of the Global Pharma Health Fund (GPHF) MiniLab® or as individual tests. Overall, 30/39 studies reported employing confirmatory testing described in international pharmacopeia to verify the quality of anti-malarials post assessment by a screening test. The authors assigned scores for the 23 criteria for the standard of reporting of each study. CONCLUSIONS: There is considerable heterogeneity in study design and inconsistency in reporting of field surveys of medicine quality. A lack of standardization in the design and reporting of studies of medicine quality increases the risk of bias and error, impacting on the generalizability and reliability of study results. The criteria proposed for reporting on the standard of studies in this review can be used in conjunction with existing medicine quality survey guidelines as a checklist for designing and reporting findings of studies. The review protocol has been registered with PROSPERO (CRD42015026782).

In vivo efficacy of top five surveyed Ghanaian herbal anti-malarial products.

BACKGROUND: Anti-malarial herbal preparations (HPs) continue to enjoy high patronage in Ghana despite reports that the artemisinin-based combination therapy (ACT), the recommended first choice for treatment of uncomplicated malaria in the country, remains efficacious. A major issue with the use of these preparations is inadequate or unreliable data on their efficacy and quality. An assessment of the potency and quality of the most popular commercial anti-malarial HPs in Ghana was, therefore, carried out. The outcome of this investigation is herein discussed preceded by a short literature review of herbal medicines in Ghana. METHODS: Using a questionnaire survey of 344 individuals in parts of Ghana, five of the most frequently used HPs were identified and selected for test of their efficacy and quality. The effect of the selected compounds on Plasmodium berghei in vivo was assessed using standard methods. RESULTS: All five tested HPs (HP-A, HP-B, HP-C, HP-D and HP-E) showed chemo-suppressive activity against P. berghei in vivo. However the degree of parasites inhibition is significantly lower compared to the WHO-recommended artemether-lumefantrine combination (p < 0.05, 99.9% chemosuppression/activity, 28 days survival). Using the Solomon Saker's Test, two of the preparations were found to contain chloroquine or compounds with chemical properties like that of chloroquine. CONCLUSION: Popular anti-malarial HPs used in southern Ghana were found to have chemo-suppressive properties. Intentional addition of chloroquine or SCs to these preparations in order to enhance their effectiveness has serious public health concerns as it may induce cross resistance to amodiaquine, one of the partner drugs in the recommended ACT for use in Ghana.

Antiplasmodial potential and quantification of aloin and aloe-emodin in Aloe vera collected from different climatic regions of India.

BACKGROUND: In this study, Aloe vera samples were collected from different climatic regions of India. Quantitative HPTLC (high performance thin layer chromatography) analysis of important anthraquinones aloin and aloe-emodin and antiplasmodial activity of crude aqueous extracts was done to estimate the effects of these constituents on antiplasmodial potential of the plant. METHODS: HPTLC system equipped with a sample applicator Linomat V with CAMAG sample syringe, twin rough plate development chamber (20 x 10 cm), TLC Scanner 3 and integration software WINCATS 1.4.8 was used for analysis of aloin and aloe-emodin amount. The antiplasmodial activity of plant extracts was assessed against a chloroquine (CQ) sensitive strain of P. falciparum (MRC-2). Minimum Inhibitory Concentration (MIC) of aqueous extracts of selected samples was determined according to the World Health Organization (WHO) recommended method that was based on assessing the inhibition of schizont maturation in a 96-well microtitre plate. EC (effective concentration) values of different samples were observed to predict antiplasmodial potential of the plant in terms of their climatic zones. RESULTS: A maximum quantity of aloin and aloe-emodin i.e. 0.45 and 0.27 mg/g respectively was observed from the 12 samples of Aloe vera. The inhibited parasite growth with EC50 values ranging from 0.289 to 1056 µg/ml. The antiplasmodial EC50 value of positive control Chloroquine was observed 0.034 µg/ml and EC50 values showed by aloin and aloe-emodin was 67 µg/ml and 22 µg/ml respectively. A positive correlation was reported between aloin and aloe-emodin. Antiplasmodial activity was increased with increase in the concentration of aloin and aloe-emodin. The quantity of aloin and aloe-emodin was decreased with rise in temperature hence it was negatively correlated with temperature. CONCLUSIONS: The extracts of Aloe vera collected from colder climatic regions showed good antiplasmodial activity and also showed the presence of higher amount of aloin and aloe-emodin in comparison to collected from warmer climatic sites. Study showed significant correlation between quantities of both the anthraquinones used as marker compounds and EC50 values of the different Aloe vera extracts. Although, both the anthraquinones showed less antiplasmodial potential in comparison to crude extracts of different Aloe vera samples. Diverse climatic factors affect the quantity of tested compounds and antiplasmodial potential of the plant in different Aloe vera samples.

Mini Review - Analysis of Artemether and Dihydroartemisinin by high performance high liquid chromatography in biological fluids-issues and solutions.

Artemether-Lumefantrine is the most widely recommended antimalarial combination used to treat millions of patients suffering from malaria. Artemether undergoes rapid metabolism and gets converted to its active metabolite dihydroartemisisn. Drug analysis is a vital aspect to evaluate drugs in research. There are a number of methods available for the determination of artemether in biological fluids. These methods include HPLC based UV detection, GS-MS, HPLC-ECD and HPLC-MS/MS. This article reviews different methods for the determination of artemether in the biological fluids. Among the available methods HPLC-MS/MS proves to be the most accurate and reliable one for analysis. This has the advantage of improved sensitivity and selectivity with smaller sample volume.

Use of thin-layer chromatography to detect counterfeit sulfadoxine/pyrimethamine tablets with the wrong active ingredient in Malawi.

BACKGROUND: Substandard and falsified anti-malarial medicines pose a serious threat to public health, especially in low-income countries. Appropriate technologies for drug quality analysis in resource-limited settings are important for the surveillance of the formal and informal drug market. The feasibility of thin-layer chromatography (TLC) with different solvent systems was tested using the GPHF Minilab in a study of the quality of sulfadoxine/pyrimethamine tablets in Malawi. METHODS: Twenty eight samples of sulfadoxine/pyrimethamine tablets were collected from randomly selected health facilities of four districts of southern Malawi. A mystery shopper approach was used when collecting samples from illegal street vendors, and an overt approach for the other facilities. Samples were subjected to visual inspection, disintegration testing and TLC analysis. 10 samples were further investigated according to the methods of the US Pharmacopeia using high performance liquid chromatography (HPLC). RESULTS: One sample was found to be falsified, containing a mixture of paracetamol tablets and co-trimoxazole tablets. These had been repackaged into paper strip packs labelled as a brand of sulfadoxine/pyrimethamine. TLC with different solvent systems readily proved that these tablets did not comply with their declaration, and provided strong evidence for the active pharmaceutical ingredients which were actually contained. Full pharmacopeial analysis by HPLC confirmed the results suggested by TLC for this sample, and showed two further samples to be of substandard quality. CONCLUSIONS: Due to the absence of the declared anti-malarial ingredients and due to the presence of other pharmaceutical ingredients, the identified falsified medicine represents a serious health risk for the population. Thin-layer chromatography (TLC) using different solvent systems proved to be a powerful method for the identification of this type of counterfeiting, presenting a simple and affordable technology for use in resource-limited settings.