Registration and local production of essential medicines in Uganda

Universal access to high quality essential medicines is critical to sustainable development (SDG 3.8). However low- and middle-income countries struggle to ensure access to all medicines on their national essential medicines lists (EML). Market registration is the first step in determining both access and availability yet the extent to which essential medicines are registered for use at country level is not known. Companies apply for a marketing authorisation, however low price or lack of a market is a disincentive. Local production has been promoted to ensure availability of essential medicines but research in this area is also limited.; The study took place between 2011 and 2015. We systematically examined the registration status of medicines and vaccines listed in the Ugandan 2012 EML and conducted 20 interviews with regulators, ministry of health representatives, donors, and pharmaceutical producers and analysed quality assurance issues affecting registration, procurement, and local production of medicines in Uganda. In 2017 we conducted a further three interviews to clarify issues around non-registration of essential medicines highlighted by our analysis.; Of the 566 essential medicines and vaccines nearly half (49%; 275/566) had no registered product in 2012. Of the 3130 registered products, just over a quarter (28%; 880/3130) were listed on the EML. Six local producers had registered 138 products of which 40 corresponded to 32 unique essential medicines. Interviews highlighted alternative routes to availability other than registration. Local producers faced considerable barriers to achieving international quality standards required for international procurement of medicines for the domestic market.; Monitoring and audit of the registration of essential and non-essential medicines should be a priority nationally and, regionally through harmonisation of registration requirements in the East African Community. National and regional manufacturing plans should consider local production of unregistered essential medicines

Antimicrobial resistance and rational prescription practices: knowledge, perceptions and confidence of health profession interns in Uganda

Background Antimicrobial resistance (AMR) is significantly driven by misuse and overuse of antibiotics. Graduate health profession interns often prescribe antimicrobials under minimum supervision. Objectives This study explored the knowledge, perceptions and confidence of health profession interns in Uganda regarding AMR and rational prescription practices. Methods This was a cross-sectional survey employing quantitative techniques carried out between October and November 2022 at six tertiary hospitals in Uganda. Health profession interns including doctors, nurses, midwives and pharmacists were recruited as study participants. Data were collected using online Kobo toolbox software. Data analysis was performed using STATA (StataCorp) version 16. Bivariate analysis and multivariable logistic regression were performed. P \u3c 0.05 was considered statistically significant. Results We recruited 281 participants with a mean age of 27 ± 3.8 years, of which few (n = 53; 19%) had good knowledge about AMR and rational prescription. The use of professional organization guidelines as a source of information was significantly associated with good knowledge (adjusted OR = 1.9; 95% CI: 1.0–3.5; P = 0.046). Nurses had the least knowledge compared with doctors and pharmacists. Continuous medical education (99%) and availability of clinical guidelines (98%) were identified as the most helpful intervention to improve knowledge. Most participants were confident about accurately diagnosing infections and sepsis and selecting appropriate antimicrobials. Conclusions Continuous medical education and availability of clinical and professional organization guidelines should be leveraged to improve the knowledge of AMR and rational prescription among health profession interns. Their high confidence in rational prescription practices should be pivotal to the fight against AMR

A method for the analysis of sugars in biological systems using reductive amination in combination with hydrophilic interaction chromatography and high resolution mass spectrometry

Separation of sugar isomers extracted from biological samples is challenging because of their natural occurrence as alpha and beta anomers and, in the case of hexoses, in their pyranose and furanose forms. A reductive amination method was developed for the tagging of sugars with the aim of it becoming part of a metabolomics work flow. The best separation of the common hexoses (glucose, fructose, mannose and galactose) was achieved when 2H5-aniline was used as the tagging reagent in combination with separation on a ZICHILIC column. The method was used to tag a range of sugars including pentoses and uronic acids. The method was simple to perform and was able to improve both the separation of sugars and their response to electrospray ionisation. The method was applied to the profiling of sugars in urine where a number of hexose and pentose isomers could be observed. It was also applied to the quantification of sugars in post-mortem brain samples from three control samples and three samples from individuals who had suffered from bipolar disorder

Metabolomic profiling of the synergistic effects of melittin in combination with cisplatin on ovarian cancer cells

Melittin, the main peptide present in bee venom, has been proposed as having potential for anticancer therapy; the addition of melittin to cisplatin, a first line treatment for ovarian cancer, may increase the therapeutic response in cancer treatment via synergy, resulting in improved tolerability, reduced relapse, and decreased drug resistance. Thus, this study was designed to compare the metabolomic effects of melittin in combination with cisplatin in cisplatin-sensitive (A2780) and resistant (A2780CR) ovarian cancer cells. Liquid chromatography (LC) coupled with mass spectrometry (MS) was applied to identify metabolic changes in A2780 (combination treatment 5 μg/mL melittin + 2 μg mL cisplatin) and A2780CR (combination treatment 2 μg/mL melittin + 10 μg/mL cisplatin) cells. Principal components analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) multivariate data analysis models were produced using SIMCA-P software. All models displayed good separation between experimental groups and high-quality goodness of fit (R2) and goodness of prediction (Q2), respectively. The combination treatment induced significant changes in both cell lines involving reduction in the levels of metabolites in the tricarboxylic acid (TCA) cycle, oxidative phosphorylation, purine and pyrimidine metabolism, and the arginine/proline pathway. The combination of melittin with cisplatin that targets these pathways had a synergistic effect. The melittin-cisplatin combination had a stronger effect on the A2780 cell line in comparison with the A2780CR cell line. The metabolic effects of melittin and cisplatin in combination were very different from those of each agent alone

Effect of bee venom and its fractions on the release of pro-inflammatory cytokines in PMA-differentiated U937 cells co-stimulated with LPS

The venom of Apis mellifera (honey bee) has been reported to play a role in immunotherapy, but existing evidence to support its immuno-modulatory claims is insufficient. Four fractions from whole bee venom (BV) were separated using medium pressure liquid chromatography. Their ability to induce the production of cytokines TNFα, IL-1β and IL-6 in phorbol-12-myristate-13-acetate (PMA)-treated U937 cells was assessed. The levels of the three cytokines produced by stimulation with the four fractions and crude BV without LPS were not significantly different from negative control values. However, co-stimulation of the cells with LPS and Fraction 4 (F-4) induced a 1.6-fold increase in TNF-α level (p < 0.05) compared to LPS alone. Likewise, LPS-induced IL-1β production was significantly synergised in the presence of F-1 (nine-fold), F-2 (six-fold), F-3 (four-fold) and F-4 (two-fold) fractions, but was only slightly enhanced with crude BV (1.5-fold) relative to LPS. Furthermore, the LPS-stimulated production of IL-6 was not significantly increased in cells co-treated with F-2 and F-3, but the organic fraction (F-4) showed an inhibitory effect (p < 0.05) on IL-6 production. The latter was elucidated by NMR spectroscopy and found to contain(Z)-9-eicosen-1-ol. The effects observed with the purified BV fractions were more marked than those obtained with the crude sample

A study of the biological activity of bee venom and its fractions with regard to cosmetic science and immunology

Apis mellifera venom has been used since antiquity to treat various ailments but scientific evidence to justify its therapeutic claims is incomplete. The venom has recently entered cosmetic usage as a mimetic ingredient to alleviate signs of facial aging. This study investigated the potential of bee venom (BV) as a source of cosmetic and immunomodulatory agents for use in skin anti-ageing applications and as a vaccine adjuvant, respectively. BV fractions were obtained by reversed phase preparative chromatography and characterised by LC-MS and NMR techniques. The fractions were assayed for antimicrobial and cytotoxic activities against a range of microbes (Mycobacterium marinum, Norcardia farcinia and Trypanosoma brucei brucei) and human cell lines [NCTC2544 (normal keratinocytes), PNT2A (normal epithelial cells), and HS27 (normal fibroblasts)] respectively. Immunomodulatory effects were investigated in PMA-differentiated U937 cells with and without LPS co-stimulation. The aqueous stability of the venom and its susceptibility to protease action were as-sessed by LC-MS assays. An LC-MS method was also developed and validated for the assay of BV in commercial creams using the melittin signal as an indicator of BV content. Of the 4 BV fractions (F-1 to F-4), the largest, containing melittin, showed the most activity against N. farcinia (25-50μg/mL) and T. b. brucei (0.78-1.56μg/mL), but was not very active against M. marinum (>100μg/mL). The melittin fraction (96% pure) was the most cytotoxic against keratinocytes, fibroblasts and epi-thelial cells, with IC50 ≥ 2.5-4.0μg/mL. All fractions significantly enhanced IL-1β release, while only F-4, a lipophilic fraction, enhanced TNF-α release. F-4, which was revealed through NMR elucidation to contain (Z)-9-eicosen-1-ol, produced sig-nificant inhibition of IL-6 release. Melittin in aqueous solutions of BV, but not of the melittin fraction alone, underwent a temperature-dependent spontaneous degradation within the 21Lys-22Arg-23Lys-24Arg sequence due to a serine carboxypeptidase-like activity attributed to the BV allergen, Api m9. Taken together, these results suggest that a formulation matching the 3.2-37.2ppm content of melittin assayed in commer-cial creams would be safe for skin application based on IC50 values in human cells. The immunomodulatory effects observed in U937 cells highlight the potential of BV as a possible source of vaccine adjuvants.Apis mellifera venom has been used since antiquity to treat various ailments but scientific evidence to justify its therapeutic claims is incomplete. The venom has recently entered cosmetic usage as a mimetic ingredient to alleviate signs of facial aging. This study investigated the potential of bee venom (BV) as a source of cosmetic and immunomodulatory agents for use in skin anti-ageing applications and as a vaccine adjuvant, respectively. BV fractions were obtained by reversed phase preparative chromatography and characterised by LC-MS and NMR techniques. The fractions were assayed for antimicrobial and cytotoxic activities against a range of microbes (Mycobacterium marinum, Norcardia farcinia and Trypanosoma brucei brucei) and human cell lines [NCTC2544 (normal keratinocytes), PNT2A (normal epithelial cells), and HS27 (normal fibroblasts)] respectively. Immunomodulatory effects were investigated in PMA-differentiated U937 cells with and without LPS co-stimulation. The aqueous stability of the venom and its susceptibility to protease action were as-sessed by LC-MS assays. An LC-MS method was also developed and validated for the assay of BV in commercial creams using the melittin signal as an indicator of BV content. Of the 4 BV fractions (F-1 to F-4), the largest, containing melittin, showed the most activity against N. farcinia (25-50μg/mL) and T. b. brucei (0.78-1.56μg/mL), but was not very active against M. marinum (>100μg/mL). The melittin fraction (96% pure) was the most cytotoxic against keratinocytes, fibroblasts and epi-thelial cells, with IC50 ≥ 2.5-4.0μg/mL. All fractions significantly enhanced IL-1β release, while only F-4, a lipophilic fraction, enhanced TNF-α release. F-4, which was revealed through NMR elucidation to contain (Z)-9-eicosen-1-ol, produced sig-nificant inhibition of IL-6 release. Melittin in aqueous solutions of BV, but not of the melittin fraction alone, underwent a temperature-dependent spontaneous degradation within the 21Lys-22Arg-23Lys-24Arg sequence due to a serine carboxypeptidase-like activity attributed to the BV allergen, Api m9. Taken together, these results suggest that a formulation matching the 3.2-37.2ppm content of melittin assayed in commer-cial creams would be safe for skin application based on IC50 values in human cells. The immunomodulatory effects observed in U937 cells highlight the potential of BV as a possible source of vaccine adjuvants

Immuno-Kachiks formula immunomodulates and ameliorates hepatic damage induced by monosodium glutamate in rats

The immune system plays a vital role in controlling liver fibrosis and enhancing the pathogenesis of liver inflammation. Monosodium glutamate is a common flavor-enhancement food additive. This study evaluated the immunomodulatory and hepato-curative effects of the Immuno-Kachiks polyherbal formulation against monosodium glutamate-induced immune suppression and hepatic damage in rats. Monosodium glutamate was given orally at a 2000 mg/kg dose to male Wistar rats for three months to induce liver damage and immune suppression. After three months of successful induction, three groups were separately administered orally with Immuno-Kachiks formula at 400, 800, and 1500 mg/kg/day for 28 days. At the end of the treatment period, liver and blood samples were collected for histological and biochemical analysis. The lymphocyte count remained significantly low while the neutrophil count and the neutrophil-to-lymphocyte ratio increased significantly, despite the cessation of monosodium glutamate ingestion for 28 days. The Immuno-Kachiks formula (IKF) significantly increased the lymphocyte count, reduced the neutrophil-to-lymphocyte ratio, and normalized the neutrophil count. Neither monosodium glutamate nor the IKF significantly caused alpha-fetoprotein levels to rise or fall below normal. High doses (800 and 1500 mg/kg) of the Immuno-Kachiks formula significantly raised serum levels of aspartate aminotransferase, alkaline phosphatase, and total bilirubin. 1500 mg/kg of the IKF caused mild liver inflammation. The IKF restored the liver morphologic alterations observed in monosodium glutamate-induced liver damage in rats. The results suggest that the Immuno-Kachiks herbal formulation is a potential curative agent for early-stage liver damage and could restore suppressed adaptive immunity

Metabolomic Profiling of the Effects of Melittin on Cisplatin Resistant and Cisplatin Sensitive Ovarian Cancer Cells Using Mass Spectrometry and Biolog Microarray Technology

In the present study, liquid chromatography-mass spectrometry (LC-MS) was employed to characterise the metabolic profiles of two human ovarian cancer cell lines A2780 (cisplatin-sensitive) and A2780CR (cisplatin-resistant) in response to their exposure to melittin, a cytotoxic peptide from bee venom. In addition, the metabolomics data were supported by application of Biolog microarray technology to examine the utilisation of carbon sources by the two cell lines. Data extraction with MZmine 2.14 and database searching were applied to provide metabolite lists. Principal component analysis (PCA) gave clear separation between the cisplatin-sensitive and resistant strains and their respective controls. The cisplatin-resistant cells were slightly more sensitive to melittin than the sensitive cells with IC50 values of 4.5 and 6.8 μg/mL respectively, although the latter cell line exhibited the greatest metabolic perturbation upon treatment. The changes induced by melittin in the cisplatin-sensitive cells led mostly to reduced levels of amino acids in the proline/glutamine/arginine pathway, as well as to decreased levels of carnitines, polyamines, adenosine triphosphate (ATP) and nicotinamide adenine dinucleotide (NAD+). The effects on energy metabolism were supported by the data from the Biolog assays. The lipid compositions of the two cell lines were quite different with the A2780 cells having higher levels of several ether lipids than the A2780CR cells. Melittin also had some effect on the lipid composition of the cells. Overall, this study suggests that melittin might have some potential as an adjuvant therapy in cancer treatment