Optimization of HPLC-MS/MS method for determination of antimalarial adulterants in herbal products.

The use of herbal products is booming all over the world because of being believed as safer than conventional drugs and free of side effects. However, there are untrustworthy manufacturers who adulterate herbal products by adding conventional drugs which might eventually lead to microbial resistance and herb-to-drug interactions. There is a need to develop methods for detecting adulterants in herbal products. A high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for simultaneous identification and determination of conventional antimalarials (chloroquine, quinine, sulfadoxine, pyrimethamine, mefloquine, lumefantrine, amodiaquine, artemisinin, dihydroartemisinin, artesunate and artemether) in herbal products was developed. Stable isotopically labelled compounds (artemether-d3, quindine-d3, and sulfadoxine-d3) were used as internal standards (ISs) for quantitative analysis. Extraction of analytes was performed using methanol: water: formic acid (90:10:0.1, v/v) and chromatographic separation was done in a gradient mode using mobile phase A: Ultrapure water containing 0.1% formic acid and 1 mM ammonium formate and mobile phase B: Acetonitrile/methanol (50:50) containing 0.1% formic acid and 1 mM ammonium formate. The calibration curves were linear (r2 ≥ 0.991) over the range of 0.001-0.3 µg mL-1 for all compounds. The limit of detection (LOD) ranged from 0.002 to 0.02 µg mL-1 while the limit of quantification (LOQ) ranged from 0.006 to 0.08 µg mL-1. Accuracy, expressed as recovery of spiked herbal products ranged from 52 to 128%. The precision, expressed as percent relative standard deviation (%RSD) at two concentration levels, ranged from 1.0 to 13.8%. The matrix effect expressed as the matrix factor (MF) ranged from 0.77 to 0.97. The developed method was used to identify and quantify conventional antimalarials in herbal product samples from Tanzania. Ten out of 50 herbal products were found to contain amodiaquine, sulfadoxine, pyrimethamine, mefloquine, dihydroartemisinin, artemether and lumefantrine. The developed method is considered a valuable tool for getting a better understanding of the adulteration of conventional antimalarials in herbal products.

Effect of changes in business environments on traded medicinal plants products in Tanzania: An explorative study.

In Tanzania, a complex rural to urban supply network for the medicinal plants' products trade has developed over time driven by changes in business environments at the macro level notably in the policy and regulatory frameworks and the micro level resulting from traders' reactions and responses to those changes and pandemics. These changes in business environments and responses of traders have shaped the current trade practices. However, the response of traders to changes in business environments and the evolutionary paths over time are not well documented. Therefore, this paper aimed to synchronize changes in business environment over time and empirically determine how the trade has evolved concerning the responses of the medicinal plants' traders in Tanzania. The study applied economic evolution theory to describe the interactions of changes in business environments and responses of traders to demarcate the evolutionary stages. Primary data were collected from traders, regulators, and researchers through ten focus group discussions and sixteen in-depth interviews from five regions of Tanzania. The results indicated that the traders of medicinal plant products responded by improving product appearance and modernizing both practices and business premises. The study findings have identified four evolutionary stages of trade in medicinal plant products in Tanzania: The first stage was the colonial era (1882-1961), the second is the government supremacy era (1961-1984), the third is the emergence of the private sector era (1985-2004), and the fourth is the market and regulation integration era (started in 2005). Moreover, because of the partial implementation of the regulatory framework, the fifth stage of trade evolution is also expected. This stage is expected to be demarcated when the regulatory framework and market forces work together. Therefore, the study recommends that proper enforcement measures be put in place to ensure desired results whenever there are changes in business environments in traditional medicines practice. This is due to the expectation of the fifth stage whereby one of its characteristics is stiff competition among traders and which will require robust business models to survive in business.

A HPLC-MS/MS method for screening of selected antibiotic adulterants in herbal drugs.

The use of herbal products adulterated with conventional drugs increases the risk of developing microbial resistance and causes herb-to-drug interaction, leading to severe clinical consequences. The complex nature of herbal products has been a challenge for the unambiguous identification of adulterants. The improved analytical selectivity and sensitivity of hyphenated techniques such as high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) enable the confirmatory screening of adulterants in herbal products. Simultaneous screening of adulterants is necessary and efficient because it has been established that more than one chemical adulterant may be present in one herbal product. An HPLC-MS/MS method for the simultaneous detection and quantification of amoxicillin, ampicillin, metronidazole, ciprofloxacin, sulfamethoxazole, and trimethoprim in powdered herbal drugs was developed. Deuterated metronidazole-d3, trimethoprim-d3, ciprofloxacin-d8, and sulfamethoxazole-d4 were used as internal standards (ISs). For each analyte, two transitions were monitored using protonated molecules as precursor ions. The extraction of analytes from herbal products was performed using a simple methanol : water : formic acid (90 : 10 : 0.05, v/v) extraction solvent. Chromatographic separation was done in a gradient of 0.01% formic acid in methanol and 0.01% formic acid in MilliQ water. The calibration curves were linear (r2 ≥ 0.996) over the range of 0.005-2.5 µg mL-1 for all compounds except metronidazole, whose range was 0.005-1 µg mL-1. The limit of detection (LOD) ranged from 0.012 to 0.046 µg mL-1, while the limit of quantification (LOQ) ranged from 0.066 to 0.153 µg mL-1. The accuracy, expressed as the recovery of spiked herbal products, ranged from 45% to 114%. The precision, expressed as relative standard deviation (RSD) at two concentration levels, ranged from 1.6% to 15.9%. The matrix effect expressed as the matrix factor (MF) ranged from 0.79 to 0.92. The developed method was applied to powder herbal products purchased in Tanzania. Amoxicillin, ampicillin, trimethoprim, sulfamethoxazole, and ciprofloxacin were not detected in all samples. Metronidazole was detected in eight samples with the highest concentration of 1.38 µg g-1. The developed method is suitable for the detection of all the studied antibiotic adulterants in herbal products. Quantification can be performed for all the compounds except ciprofloxacin due to its lower recovery.

Ethnobotanical survey and toxicity evaluation of medicinal plants used for fungal remedy in the Southern Highlands of Tanzania.

BACKGROUND/AIM: Some of the antifungal drugs used in the current treatments regime are responding to antimicrobial resistance. In rural areas of Southern Tanzania, indigenous people use antifungal drugs alone or together with medicinal plants to curb the effects of antibiotic resistance. This study documented ethnobotanical information of medicinal plants used for managing fungal infections in the Southern Highlands of Tanzania and further assess their safety. MATERIALS AND METHODS: Ethnobotanical survey was conducted in Makete and Mufindi districts between July 2014 and December 2015 using semi-structured questionnaires followed by two focus group discussions to verify respondents' information. Cytotoxicity study was conducted on extracts of collected plants using brine shrimp lethality test and analyzed by MS Excel 2013 program. RESULTS: During this survey about 46 plant species belonging to 28 families of angiosperms were reported to be traditionally useful in managing fungal and other health conditions. Among these, Terminalia sericea, Aloe nutii, Aloe lateritia, Zanthoxylum chalybeum, Zanthoxylum deremense, and Kigelia africana were frequently mentioned to be used for managing fungal infections. The preparation of these herbals was mostly by boiling plant parts especially the leaves and roots. Cytotoxicity study revealed that most of the plants tested were nontoxic with LC50 > 100 which implies that most compounds from these plants are safe for therapeutic use. The dichloromethane extract of Croton macrostachyus recorded the highest with LC50 value 12.94 µg/ml. The ethnobotanical survey correlated well with documented literature from elsewhere about the bioactivity of most plants. CONCLUSIONS: The ethnobotanical survey has revealed that traditional healers are rich of knowledge to build on for therapeutic studies. Most of the plants are safe for use; and thus can be considered for further studies on drug discovery.

Determination of oxytetracycline residues in cattle meat marketed in the Kilosa district, Tanzania.

Oxytetracycline is used to treat various diseases in cattle. However, its use may be associated with unacceptable residue levels in food. Oxytetracycline residues in tissues from indigenous cattle were determined in a cross-sectional study conducted in the Kilosa district, Tanzania, between November 2012 and April 2013. A total of 60 tissue samples, including muscle, liver and kidney, were collected from slaughterhouses and butchers and analysed for oxytetracycline using high-performance liquid chromatography. Oxytetracycline residues were found in 71.1% of the samples, of which 68.3% were above acceptable regulatory levels. The mean concentration of oxytetracycline across tissues was 3401.1 µg/kg ± 879.3 µg/kg; concentrations in muscle, liver and kidney were 2604.1 µg/kg ± 703.7 µg/kg, 3434.4 µg/kg ± 606.4 µg/kg and 3533.1 µg/kg ± 803.6 µg/kg, respectively. High levels of oxytetracycline residue in meat from indigenous cattle may pose a health threat to consumers in Kilosa. The findings possibly reflect a general lack of implementation of recommended withdrawal periods, ignorance about drug use and lack of extension services. Strict regulation of the use of antimicrobial drugs in the livestock industry and associated testing of animal-derived food sources prior to marketing are required.