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  • Palttala, Iida (2010)
    Automated dose dispensing is an increasing field in which medicines are packaged mechanically into small one-dose pouches in portions of two weeks. Suitability of tablets for automated dose dispensing has not been researched systematically earlier. The study was made in collaboration with the dose dispensing unit of Espoonlahti pharmacy. The aim of the study was to define optimal characteristics for an automatically dispensed tablet from a viewpoint of the dose dispensing process to reduce breakings and transitions. Breaking means that tablet crumbles, splits up or breaks up otherwise during mechanical dose dispensing process. Transition means that tablet is dispensed in a wrong dose pouch. Percentually breakings and transitions occur very little, but quantitatively plenty and increasingly when automated dose dispensing is becoming more common. Breakings and transitions cause plenty extra work because of correcting pouches, so their amount should be aimed to reduce. In addition, the aim is to find out matters to enquire from the manufacturers of medicines that would help concluding whether a product is suitable for automated dose dispensing based on written information. Results of the study indicate that to reduce breakings and transitions, an optimal tablet product for dose dispensing is rather small or middle sized, coated, strong and without a breakline and the optimal relative humidity of air in the product room of dose dispensing unit would be around 30 - 40 %. Matters to enquire from the manufacturers of medicines besides size, coating, breaking strength and breakline are stability of the product outside of its original package and light, heat and moisture sensitiveness of the product. Besides breakings and transitions, also stability of a moisture sensitive acetylsalicylic acid product (Disperin 100 mg) was investigated in 25 °C/60 % RH because air humidity in the product room is not adjustable. Duration of the test was four weeks. It is enough since it is the maximum time that tablets are outside their original packages during drug dispensing process before use. Tablets were kept in opened original container (bottle), in closed original container, in cassette of dispensing machine and in two different dose pouches (new material and the one in use). According to the results, cassettes are protecting tablets from moisture as poorly as an opened bottle. Instead, new pouch material protects tablets better than the material in use. Results of Raman spectroscopy measurements indicate no change in acetylsalicylic acid to salicylic acid during four weeks test. Moisture affects to tablets by decreasing breaking strength, which may cause more breakings. Air humidity should be adjusted in product rooms or tablets should be unpacked into cassettes as near operating the machine as possible to prevent breakings. Especially when air humidity is high. Among others, a heat sensitive drug product was researched because of the seaming unit of dose dispensing machine which is radiating heat of about 75 °C to pouches if machine is pulled over in the middle of work. Study was performed with variable temperature XRPD. Results of the study of heat sensitiveness indicate that 75 °C for 60 minutes doesn't induce changes in carbamazepine tablet (Neurotol 200 mg). However, results of the study reveal that researched product did not contain the most heat sensitive form of carbamazepine, so other heat sensitive drug products should be examined to get more information about effects of heat.
  • Mertanen, Sini (2023)
    For pediatric patients, it is often necessary to resort to off-label use of available commercial products. This may require manual modification of the preparations, which may result in reduced dose accuracy. In the past, there have been discussions about the potential of 3D printing technologies for on-demand manufacturing of medicines in hospitals. Printing technologies can be used to tailor medicines to the individual needs of patients. This could be a possible solution to the lack of commercial products for pediatric patients, for example. Semi-solid extrusion is a printing technique that could potentially be used in the future in hospitals. This study aims to design the simplest possible excipient composition for a printing material for semi-solid extruded preparations for pediatric patients. The finished products will be examined to determine the type of products achieved with this printing method and excipient composition. In addition, the suitability of semi-solid extrusion in a hospital environment will be observed and evaluated. Printing was performed with a pneumatic bioprinter. The desired formulations could not be prepared with a printing material containing only a gel former (poloxamer 407) and a solvent (water). Therefore, a filler (microcrystalline cellulose) was added to the printing material to improve the mechanical strength of the preparations. The model drug used in the study was warfarin sodium and the target strengths of the preparations were 0.1 mg, 0.5 mg and 1.0 mg. The preparations were dried at room temperature for 22-23 hours. The tablets produced in the study were small (diameter less than 7 mm, height less than 2 mm) grid-structured preparations. The method was successful in producing tablets of uniform mass. For all strengths, the tablets passed the European Pharmacopoeia test for uniformity of content of single-dose preparations. Only the 0.5 mg strengths passed the test of uniformity of dosage units. The excipient composition should still be optimized to improve the mechanical strength of the products. The overall preparation time of the formulations should be reduced, for example by shortening the drying time, to make semi-solid extrusion suitable for extemporaneus preparations in hospitals.
  • Palomäki, Emmi (2012)
    3D-imaging is based on combining two or more pictures to form one three-dimensional picture. Most of the methods used provide only surface pictures, but tomography acquires also information about the inside-structure of investigated material. Young's modulus is a method, which has been used for long time to determine toughness hard materials, such as steal. In traditional method a beam-shaped piece is bent. When the size of piece, used force and amount of bending are known, Young's modulus of piece can be calculated. Although the method has traditionally been used to research very hard materials, it has been applied without changes with pharmaceutical materials. It is, however, open to the question whether or not the method is appropriate for those materials. There are also methods to determine Young's modulus based on compressing a tablet or using ultrasound. Determining tablet's toughness with ultimate strength test is complicated because it breaks tablet. For that reason it would be good to find compensatory methods to measure strength of tablet. The aim of the study was to validate Flash Sizer 3D appliance, which is used in 3D-imiging. Another goal was to investigate possible correlations between 3D-imiging, Young's modulus and traditional ultimate strength method. Lastly, the feasibility of Young's modulus as a substitute for traditional ultimate strength measurement in self life studies was investigated. Flash Sizer 3D was validated by measuring particle size distribution of pellets, which were made of microcrystalline cellulose (Cellets). Sizes of the investigated pellets were 100 µm, 200 µm and 500 µm. Also binary mixture of 100 µm and 200 µm was investigated. From microcrystalline cellulose was made tablets and 3D-pictures were taken. Ultimate strength test was made for half of the tablets. Young's modulus was measured from half of the tablets in tableting day, day after that and nine days after tableting. Results show that Flash Sizer 3D is suitable for investigating bigger Cellet. With smaller particles distinguishing of tablets wasn't probably good enough. Still it seems to be quite good method to determine surface roughness of tablet. Young's modulus seems to be very promising as compensating method for traditional ultimate strength measurement. In future in self life studies tablets hardness might be able to investigate by measuring Young's modulus and not measuring ultimate strength. If correlation between Young's modulus and solubility meets the case, Young's modulus might also replace also solubility measurements in self life studies.
  • Hemminki, Nelli (2024)
    Tablets are solid medicinal products that are produced by compressing tablet mass in a tablet press. The reproducible and reliable functionality of the manufacturing process, facilities and equipment used in the manufacturing process of medicinal products must be validated. There must be a marketing authorisation for the sale of medicinal products, and pharmaceutical companies must comply with current legislation, Good Manufacturing Practices, and other binding guidelines. After marketing authorisation, certain changes, such as significant changes related to the manufacturing process or raw materials, must be notified to the competent authority by means of a variation application. Depending on the extent of the change, a revalidation and/or a regulatory approval before implementing the change may be required, which makes manufacturing process modification slow, expensive, and laborious. However, the pressure to enhance pharmaceutical manufacturing processes and reduce costs is prevailing. In this study, Lean Six Sigma methods were applied to enhance the manufacturing process of a tablet product. The study was divided into two parts. In the first part of the study, the objective was to investigate the manufacturing process of a tablet product X and to identify improvement actions that would make the manufacturing process of the tablet product X more efficient by achieving material and time savings. In addition, improvement actions were prioritized based on the benefits achieved by the improvement actions and the difficulty of implementation. The second part of the study was based on the improvement action selected from the first part of the study. The objective of the second part of the study was to investigate the factors influencing the amount of mass loss during tableting and how the amount of mass loss can be influenced. In this study, mass loss during tableting referred to the dusty mass inside the tablet press during tableting, which is removed from the tablet press to the equipment dedusting system. The purpose of the practical experiment conducted in the tablet production, which was part of the second part of the study, was to investigate how the magnitude of the exhaust airflow and fan power of the equipment dedusting system affects the amount of mass loss generated during tableting. Based on the experiment, adjusting the power of the equipment dedusting system can affect the amount of mass loss generated during tableting. However, further studies are needed to ensure that no dusty mass remains in the piping of the dedusting system or in tablet press at low exhaust airflow and fan power values during tableting. If the results are applied to other tablet products, the effect of a different formulation on powder dusting should be considered. Adequate dedusting during tableting is important so that the dusty mass inside the tablet press is removed as intended, facilitating the cleaning of the tablet press after tableting and reducing the risk of cross-contamination and exposure of workers to dust. The conclusion of this study is that Lean Six Sigma methods can be used more extensively to enhance the manufacturing processes of both tablet products and other pharmaceutical dosage forms and to reduce loss generated during the manufacturing process without revalidation or changes to a valid marketing authorisation.
  • Salonen, Iiro (2010)
    It was given a task to the Finnish Medicines Agency (Fimea) to prepare a national Medicines Information Strategy. The strategy process can be divided into four stages: 1) the collection and analysis of the information, 2) the determination of the strategy and the vision, 3) the realization and 4) the follow-up stage of the strategy. In the European Union (EU) the High Level Pharmaceutical Forum has drawn up the criteria for the high quality medicines information (MI) and the recommendations to improve the quality and availability of the MI directed to the consumers. The most significant medicines related political actions in Finland in the 21st century are the Medicine policy 2010 -document, the strategies of the National Agency for Medicines and the TIPPA-project. The objective of the Master's thesis was to produce the information to Fimea's MI work. The electric questionnaire was drawn up in the Ministry of Social Affairs and Health. After a pilot test, the questionnaire was sent by email to all national Medicines Authorities in EU (n=27) in November 2009. The purpose of the questionnaire was to find out 1) the significance of the medicine information in the national legislation, 2) the possible MI strategies and 3) the control mechanisms of the medicine information directed to the consumers. The medicines information strategies were found in the United Kingdom (UK), Italy and Germany. Furthermore, the strategy process was unfinished in four countries. In the strategy of the UK 25 concrete actions were presented during a three-year strategy period to improve the quality and availability of the MI and to improve the cooperation between public and private actors. The information and communication technology (ICT) was in the centre of the medicines information offered to the consumers. ICT was utilised by publishing Patient Information Leaflets in Internet and by developing medicines information web pages, digitally patient counseling services and quality certificates. The results of the survey can be utilised as a part of the Fimea's Medicines Information Strategy process. Further studies, for example an analysis of the interest groups, are needed before the preparation of the national strategy. Furthermore, experiences of the implementation of the strategy and the results reached in the UK should be clarified.
  • Hakoinen, Suvi (2014)
    Various cardiovascular diseases cause a significant portion of the direct healthcare costs. For this reason it is important to develop effective treatment strategies. In the treatment of heart failure maintaining healthy lifestyle is an important issue in addition to medical treatment. The aim of this study is to find out whether multidisciplinary interventions in the treatment of heart failure have had better patient outcomes and/or cost savings compared to the standard care and has a certain intervention method proved itself to be more effective than another. The study focuses on the published material in the last ten years, which is compared with prior systematic review about the same topic. The study was conducted as a systematic review. The literature search was performed on Medline (Ovid), CINAHL and Scopus databases. MeSH-terms and other keywords were used in the search. Search was limited to English-language studies. The article selection was made after 06.01.2004 published studies based on the inclusion and exclusion criteria. A quality assessment was made to the selected articles. Only valid articles exceeding 60 % of the maximum points were selected to this review. Literature search found 848 references, of which quality assessment was carried out for 17 articles. 12 articles were selected to this review. Based on statistical significance mortality rate was reduced only rarely. There were lots of heterogeneity in the results for reducing hospitalizations or improving the quality of life. Hospital or clinic-based interventions seemed to be slightly more effective than other intervention methods. There have been no major changes in the intervention methods, only the methods using mailed materials have been left out. Also the outcomes used are similar to those in the past, although validated instruments in different areas of determining the effectiveness have not been developed. This study found no clear evidence about the effectiveness of the treatment programs. Hospital or clinic-based interventions might be more effective than other intervention methods. There is a lot of research about the issue, but high-quality cost-effectiveness studies are lagging. The results from different studies are hard to integrate because the ways to measure vary. The effectiveness of the intervention depends on many factors, and the sheer increase in the knowledge does not seem to be crucial. For assessing the profitability of multidisciplinary interventions there is a need for a high-quality research about the cost-effectiveness of the treatment programs.
  • Huttunen, Sari (2014)
    Long-term use of benzodiazepines is not recommended in the aged. Elimination of these drugs is delayed in the aged, which can prolong drug action and expose users to adverse effects. Long-term benzodiazepine use is associated with many adverse effects, including cognitive impairment and falls. However, there are only few published studies dealing with associations between benzodiazepine concentration and clinical outcomes in the aged in long-term use. The aim of this study was to explore association of residual concentration of temazepam, oxazepam and zopiclone with age, gender, kidney function, drug dose and clinical outcomes, such as self-perceived health and functional abilities in aged patients. The data were collected in Pori City Hospital in July 2004. The patients were between 60-98 years of age (median 81) and the majority (79%) of them were women. Residual drug concentrations were analysed from serum from patients using temazepam (n=14), oxazepam (n=11) or zopiclone (n=28) regularly. Residual oxazepam concentration correlated positively with evening dose (p<0.001) and daily dose (p=0.003). Also oxazepam concentration was higher (p=0.017) in patients who took the last dose later (21:00-02:45) compared to patients with earlier dosing time (15:35-19:00). There was no such association between temazepam or zopiclone and dose or dosing time. This might be explained by the fact that there was more dispersion in the dose and dosing time of oxazepam compared to temazepam and zopiclone. There were no other associations between drug concentrations and demographic variables studied. Concerning associations with clinical outcomes, zopiclone concentration tended to be higher in patients who often felt themselves tired during daytime (p=0.087). Surprisingly, residual serum concentration of oxazepam seemed to be higher in patients who were able to walk and manage their shopping independently. Apart from these findings, residual concentration of temazepam, oxazepam and zopiclone associated poorly with clinical outcomes. These findings do not support routine monitoring of residual benzodiazepine concentrations in aged patients.
  • Parhiala, Minna (2020)
    Suomessa on otettu käyttöön nykyiset lääketurvatoiminnot asteittain vuodesta 2012 lähtien. Merkittävimmät muutokset lainsäädännössä olivat haittavaikutuksen määritelmän muutos sekä erityisen turvallisuuprofiilin lääkkeiden asettaminen lisäseurannan piiriin. Haittavaikutusten spontaani ilmoittaminen on edelleen ensisijainen menetelmä lääkkeen turvallisuustietojen keräämiseksi lääkkeen markkinoille saattamisen jälkeisessä vaiheessa. Vaikka spontaani haittavaikutusilmoitusjärjestelmä on ollut olemassa jo 1960-luvulta, on haittavaikutusten aliraportointi valitettavan yleistä. Tämän tutkielman tavoitteena oli arvioida tietoja, kokemuksia ja asenteita lääkkeiden haittavaikutusten ilmoittamisesta, nykyisestä lääketurvalainsäädännöstä sekä lääkkeiden lisäseurannasta ja mustasta kärkikolmiosta terveydenhuollon ammattilaisten keskuudessa Suomessa. Lääkärit (n = 38), sairaanhoitajat (n = 45), farmaseutit (n = 115) ja proviisorit (n = 36) vastasivat verkkokyselyyn. Aineisto analysoitiin osin aineistolähtöisen ja osin teoriaohjaavan sisällönanalyysin avulla. Tutkimuksen teoriasidonnaisena viitekehyksenä toimi Rogersin diffuusioteoria. Tämä pro gradu -tutkielma paljasti joitain eroja haittavaikutusilmoituksiin liittyvissä teidoissa ja asenteissa terveydenhuollon ammattilaisten välillä. Suurin osa vastaajista koki tietävänsä lääkkeiden lisäseurannasta. Lisäseurannasta ennestään tienneiden joukossa, musta kärkikolmio symboli oli paremmin farmasistien (> 84,5%) tiedossa verrattuna lääkäreihin (45,2%) ja sairaanhoitajiin (32,4%). Lääkäreillä oli enemmän kokemusta, mutta vähemmän tietoa haittavaikutusten ilmoittamisesta kuin muilla terveydenhuollon ammattilaisilla. Suurin osa terveydenhuollon ammattilaisista ei muuttanut työskentelytapojaan lisäseurannassa olevien lääkkeiden kohdalla. Tämä tutkielma tuo esiin sen, että terveydenhuollon ammattilaisilla on riittävät tiedot haittavaikutusilmoitusten tekoon, mutta raportoinnin monimutkaisuuden, huonon saatavuuden ja ilmoitusten tekoon rohkaisemattomuuden takia ilmoituksia tehdään vähän. Tulevaisuudessa tietoisuutta lisäseurannan alaisista lääkkeistä on parannettava sekä raportointiprosessia yksinkertaistettava ja tuotava helpommin saataville.
  • Hiltunen, Anna (2015)
    Periodontitis is a globally significant disease which destroys the attachment tissues and alveolar bone of teeth, eventually leading to tooth loss. Biofilms, the most intrinsic lifestyle of bacteria, play a pivotal role in the occurrence of this disease. Periodontal biofilms can be treated with topically administered chlorhexidine and strain-specific antibiotics. However, these antimicrobials do not offer solutions for periodontal attachment tissue and alveolar bone loss. Some therapeutical alternatives for these conventional treatments have been investigated. In numerous studies, periodontitis is treated successfully (increased attachment and/or alveolar bone levels) with topically and systemically administered bisphosphonates. Furthermore, a topically administered bone graft substitute (bioactive glass) has shown to improve periodontal parameters. In addition, bioactive glass has known antimicrobial and anti-biofilm effects. Moreover, a few bisphosphonates have shown antimicrobial activity against some bacterial strains. Hence, both bisphosphonates and bioactive glass are promising materials for dental applications, also raising interest in their combination. Indeed, it could be hypothesized that this combination product could simultaneously treat both the underlying cause (biofilms) and consequences (alveolar bone and attachment tissue loss) of periodontitis. Open research questions remain for the combination product. Is the anti-biofilm effect enhanced when bioactive glass is combined with bisphosphonates? Moreover, do bisphosphonates have intrinsic anti-biofilm properties? These questions are investigated in this thesis, which is a continuation of a recent doctoral dissertation. In this dissertation, a clodronate-bioactive glass combination product was studied by applying it into periodontal pockets. However, anti-biofilm effects were not assessed. In this thesis, a close examination is carried out on these effects, utilizing relevant biofilm models. The aims of this work were to investigate anti-biofilm effects of bisphosphonates (alendronate, clodronate, etidronate, risedronate and zoledronate) (i) alone, administered as solutions and (ii) combined with bioactive glass S53P4. Optimization of the used assay methods (96-well plate assay, Static Biofilm method) was performed. The anti-biofilm effects of bisphosphonate solutions were screened in the 96-well plate assay using a model organism Staphylococcus aureus Newman and a periodontopathogen Aggregatibacter actinomycetemcomitans ATCC 33384. After this, experiments were conducted with bisphosphonate-bioactive glass combinations. The experiments were performed with a single-specie (A. actinomycetemcomitans ATCC 33384) dental biofilm model based on the Static Biofilm method. The model mimics conditions encountered by periodontal bacteria in the oral cavity. In this part, bisphosphonate particle sizes were measured to determine a suitable control material. In addition to bacterial experiments, pH measurements were carried out to gain an insight to a possible anti-biofilm mechanism. Bisphosphonates administered as stand-alone compounds did not have an effect on either the Gram-positive model organism (S. aureus Newman) or the Gram-negative periodontopathogen (A. actinomycetemcomitans ATCC 33384). In contrast, most combinations of bisphosphonate-bioactive glass revealed a statistically significant increase in anti-biofilm effect on A. actinomycetemcomitans ATCC 33384. The combinations were compared to a control composed of inert glass and bioactive glass. In these assay conditions, the risedronate-bioactive glass-combination was the most effective (significant statistical difference, p < 0.05). Other combinations also reduced biofilms (significant statistical differences, p < 0.05), with the exception of clodronate-bioactive glass, where the change was not statistically significant. The most effective combinations (containing risedronate and etidronate) subjected the biofilms to a period of low pH. Conversely, the least effective combination (clodronate-bioactive glass) rapidly became alkaline, similarly to the control compounds (inert glass and bioactive glass). Thus, anti-biofilm efficacy could be connected to lowered pH. This observation is supported by recent literature where A. actinomycetemcomitans has been deemed highly sensitive to acidity. However, establishing the anti-biofilm rank order of bisphosphonate-bioactive glass combinations would benefit from experiments with equal bisphosphonate particle sizes.
  • Kulmala, Veera (2022)
    Parkinson’s disease (PD) is a progressive neurodegenerative disorder with the neuropathological hallmark of intraneuronal inclusions called Lewy bodies (LB). Accumulation of α-synuclein (α-syn) and cellular components into LBs coincides with degeneration of dopaminergic neurons in the midbrain, substantia nigra. Degeneration of dopaminergic neurons eventually leads to motor dysfunctions. Currently, the treatments for PD are symptomatic. For this reason, new disease-modifying treatments are needed to slow down or prevent the disease progression. Neurotrophic factors (NTFs) have been an interest of research for a couple of decades because of their neuroprotective properties. The main aim of this study was to investigate if brain-derived neurotrophic factor (BDNF) reduces pre-formed fibril (PFF) induced aggregation of α-syn in dopaminergic neurons. PFF-model was used to mimic the accumulation of LBs in neurons, as PFFs induce aggregation of endogenous α-syn in neurons. Additionally, the dose dependence of BDNF was tested. The secondary objective was to investigate the interaction of tropomyosin receptor kinase B (TrkB) signaling pathway and α-syn aggregation using TrkB agonists and antagonists. The cultured dopaminergic neurons isolated from the midbrain of mouse embryos were treated with PFFs on the day in vitro (DIV) 8. BDNF or control treatments were added either 1 hour after the PFF-treatment or on DIV 12. Neurons were fixed on DIV 15 and fluorescent immunohistochemistry was performed. After the detection of fluorescence with automated, high-content imaging, image analysis was done for quantifying dopaminergic neurons, and dopaminergic neurons positive for LB-like aggregates by using unbiased image analysis CellProfilerTM software. Both BDNF and positive control glial cell line-derived neurotrophic factor (GDNF) significantly reduced LB-like aggregates in dopaminergic neurons at both timepoints. GDNF was more effective at both timepoints than BDNF. Both tested doses of BDNF lowered the number of LB-like aggregates, but a more robust effect was seen with the higher dose. The highest tested dose for the TrkB agonists was toxic to the cultured dopaminergic neurons, whereas the lower doses did not affect either the survival or the number of LB-like aggregates. BDNF promoted the survival of the dopaminergic neurons despite the survival-reducing adverse effect of TrkB antagonist K252a. This study provided new information on the effects of exogenously added BDNF on PFF-model with primary neuronal culture. Research on the underlying mechanisms of α-syn aggregation and the protective effects of NTFs can forward the development of new therapies against PD.
  • Åman, Olli (2014)
    Plant cells in plant cell cultures can be used for production of secondary metabolites and recombinant proteins. Producing the desired compounds can be problematic since cells grow slowly, yields can be low and sometimes plant cells do not produce the desired compounds. Yields can be increased by various methods, of which optimisation of growth conditions to favour growth and secondary metabolite biosynthesis is one of various strategies. Light quality is known to have an impact on growth of plants and on accumulation of secondary metabolites. Plants receive information of their environment with photoreceptors, which gives plants ability to alter their morphology and biochemistry to adapt to the prevailing conditions. One of the most important factors involved in controlling morphology and metabolism is activity of bZIP protein HY5, which levels are controlled by degradation by E3 ubiquitin ligase COP1. The photoreceptors are divided to three main groups. A group of Blue/UV-A photoreceptors consists of cryptochromes and phototropins. Phytochromes are photochrome photoreceptors of wavebands of red and far-red. UVR8 photoreceptors are specialized to sense UV-B wavebands. Activated photoreceptors reduce the activity of COP1 individually or inductively. Plant cells contain the same genetic information as intact plants. Object of this study is to investigate effects of different light spectra on plant cell mass pigment accumulation, lipid content and accumulation of secondary metabolites. Additionally, the obtained results can be utilized in designing new artificial light sources to enhance growth and nutritional value of horticultured plants grown under artificial light. VTT's callus cultures established from berries of Rubus (raspberry, cloudberry, arctic bramble) and Vaccinium (lingonberry, bilberry, cranberry) were used in this study. The cell cultures were grown in hormone balanced solid media. For this research Valoya provided four different LED light sources with different spectra, ranging between wavebands 400–800 nm. All berry callus cultures were grown for continuous period of 28–31 days under different light sources. Mass pigments, lipid composition, total phenolic concentration and anthocyanins were analysed from each cell cultures which received different light treatments. Samples were pooled and were by freeze dried and milled. Mass pigments were extracted with acetone and analysis was carried out with UPLC-DAD. Extraction of lipids was carried out with petroleum ether followed with transesterification of glycerolipids and silylation of free fatty acids. The lipid extracts were analysed with GC-MS. Phenolic compounds were extracted with methanol and the extracts were treated with Folin-Ciocalteu's reagent and then analysed with spectrophotometer. Anthocyanins were extracted with acidified methanol and a portion of the extracts were hydrolysed to qualify anthocyanidin moieties of anthocyanins. The extracts and the hydrolysed extract were analysed with UPLC-DAD. Analysis of volatile compounds from each light treated samples was carried out with SPME GC-MS. The obtained results were used to compare concentration differences of the analytes under different light treatments. Correlations between the concentrations of the analytes and different wavebands were possible to establish from the results. Activation of cryptochromes and phytochromes reduced certain lipids that are precursors in LOX-pathway which indicates to increased activity of the pathway. Same wavebands which activated the photoreceptors reduced accumulation of mass pigments, whereas, wavebands of far-red increased the concentrations of mass pigments. In some cases it was observed that small difference in light spectra reduced mass pigment accumulation significantly. The plant cell cultures produced mainly anthocyanins which anthocyanidin moieties were same as in intact plants. Cryptochrome and phytochrome activation increased accumulation of anthocyanins. Yields of anthocyanins can be increased significantly with certain spectra significantly. The effect of light spectra did not have as straightforward effect on total phenolic content. Specie- and linewise differences were observed in light conditions where the highest concentrations of total phenolics were obtained.
  • Jormanainen, Miika (2021)
    Pharmaceutical industry is in a process of adopting new technologies due to the growing interest towards the continuous manufacturing approach. However, while the continuous wet granulation process with twin-screw granulator (TSG) has been studied widely, there has been less focus on subsequent continuous granule drying process. As a result, truly continuous granule drying device has not been available for a long time. However, L.B. Bohle has introduced a horizontal truly continuous fluid bed dryer (CFBD) in form of a perforated belt. In such system the wet granules are transported via vibration along the bed without actual fluidization while the hot air dries the granules. Accordingly, Bohle QbCon-25 fully integrated powder-to-tablet system facilitates the combination of a TSG and CFBD. However, the combination is relatively new and only few studies using these methods are available. Aim of this study was to experimentally evaluate the novel Bohle’s CFBD and elucidate the effect of formulation and process parameters of TSG and CFBD on granule residual water content. Additionally, the granules were characterized by their size distribution and bulk and tapped densities. Granule temperature and residence time in the dryer was determined with CubiSens mini sensors. In total 84 wet granulation trials in design of experiments setup were performed and multiple linear regression (MLR) model was used to investigate the effects of different process parameters on granule loss-on-drying (LoD) response. As a result, formulation microcrystalline cellulose (MCC) amount and all studied process parameters of TSG and CFBD showed significant effect on drying result indicating a robust manufacturing process. The increase in the amount of MCC in the formulation as well as the increase in L/S ratio and line rate in the wet granulation was reflected in a higher residual water content in dried granules. However, with increased drying time, airflow rate and inlet air temperature the granule residual water content decreased. The most influential process parameter affecting the granule residual water content was the used L/S ratio. In contrast, the material acceleration which corresponds to the granule drying time was the most significant process parameter of the CFBD affecting the granule residual water. However, the material acceleration did not only correspond to the material residence time in the dryer, but also to the thickness of the granule bed in the dryer. This indicated that the material acceleration is a critical process parameter of the CFBD. However, the thickness of the granule bed could not be measured in real time in this study. Additionally, at intense drying conditions granules showed a very dry outcome and further studies are required to elucidate the operational limit of the CFBD device. Furthermore, the vibration during the drying phase did not have an effect on granule size or the bulk and tapped densities. Material behavior in system was plug-flow like and mean material residence time in the CFBD was only 40 seconds with the middle material acceleration setting. Moreover, the temperature of the granules rose close to the process air temperature, but only for a very short time. Additionally, the used process settings in wet granulation affected the granule temperature, however, the most influential factor on the granule temperature was the used inlet air temperature. Overall, Bohle QbCon25 manufacturing system with TSG and CFBD showed high suitability to wet granulate and dry the produced granules with a uniform residual water content up to 20 kg/h throughput rate without process instability issues.
  • Hämäläinen, Noora (2021)
    Mini-tablets are 1-3 mm in diameter and administered as a single tablet or as a multi-particulate formulation. Mini-tablets are an attractive alternative for conventional solid dosage forms due to the ease of administration and the possibility for combination and individualised drug therapy. In mini-tablet production, good flowability of the formulation is critical as minor variations in die filling can lead to significant changes in mini-tablet weights. In addition, to reduce weight variation, the particle size should not exceed 1/3rd of the die diameter. This study aimed to determine the influence of the granule size on mini-tablet weight variability and content uniformity. The feasibility of direct compression, as well as high-shear wet granulated and roller-compacted formulations, were evaluated. From the nine final formulations manufactured, particle size distribution, Hausner ratio, Carr’s index, angle of repose and flowability were determined. The mini-tablets were made on a rotary tablet press using single punches of 3 mm in diameter. Content uniformity and weight variation of the mini-tablets were determined. The direct compression formulation had the smallest particle size, and the roller-compacted formulation milled through a 1.0 mm and 1.25 mm square screen had the largest particle size. Surprisingly, the RC 0.8 mm grater screen formulation had a very wide particle size distribution and is classified as a very fine blend. The wide particle size distribution might result from a high fill ratio during the milling of the roller-compacted ribbons. The four different high-shear wet granule formulations had a very similar particle size distribution. According to the Hausner ratio and Carr’s index values, the flow properties of the formulations varied between fair and very poor, while according to the angle of repose, the flow properties were between excellent and poor. However, all nine formulations were used to make mini-tablets with acceptable uniformity of mass, mini-tablets were within ± 8 % of the target weight, and none exceeded the 10 % limit set by Ph. Eur. The weight variation is small, as indicated by the low RSD of 1.0-2.9 %. The differences in the weight variation may be attributed to segregation due to particle or granule size and density. This is further supported by the fact that no force feeder or vacuum was utilised in the rotary tablet press, possibly causing re-circulation of the formulation and shearing forces. In addition, the fill level of the feeder might have varied between the nine formulations and affect the weight variation in a way that is not recognised in this study. Only direct compression formulation was within the limits of uniformity of content of single-dose preparations set by Ph. Eur. In the final formulations, the amount of paracetamol was in the HSWG 0.8 mm round screen 98.5 % and in the RC 1.0 mm square screen 97.7 %. These results suggest that the formulations contained an adequate amount of paracetamol, which does not explain why the mini-tablets made from high-shear wet granules did not meet the content uniformity criteria. Furthermore, the weight variation might not entirely explain why high-shear wet granulated formulations performed so poorly in the content uniformity analysis. In summary, that direct compression is a feasible manufacturing method for mini-tablets of 3 mm in diameter. However, further studies are needed on the content uniformity of mini-tablets made using high-shear wet granulated and roller-compacted formulations as these did not meet the content uniformity criterion. In particular, the content uniformity of the mini-tablets made from the high-shear wet granulated formulations was not acceptable, and the reason for this was not identified.
  • Sundberg, Enikö (2024)
    Viimeaikaisten tutkimusten perusteella suun sairauksilla on havaittu olevan vakavia systeemisiä vaikutuksia sekä vaikutuksia yleisterveyteen ja hyvinvointiin. Antiseptisia valmisteita käytetään tulehduksellisten suun sairauksien hoidossa, mutta niillä on todettu riittämätöntä tehoa ja paikallisia sivuvaikutuksia. Antimikrobisia aineita käytetään vakavampien suun tulehdusten hoitoon, mutta ne lisäävät antibioottiresistenssin riskiä ennaltaehkäisevässä ja toistuvassa käytössä. Kaksoisvalohoito vähentää mikrobilääkkeiden ja antiseptisten aineiden tarvetta ja siten haitallisia sivuvaikutuksia. Varsinkin ennaltaehkäisevänä hoitona kaksoisvalon uskotaan aiheuttavan vähemmän haittaa, mutta olevan silti riittävän tehokas estämään tulehduksia. Fotodynaamista hoitoa (PDT) on tutkittu mm. parodontiitin, peri-implantiitin ja suun punajäkälän hoidossa. Satunnaistettu, yhdessä keskuksessa toteutettava kliininen lääketutkimus suunniteltiin ja toteutettiin määrittämään säännöllisesti käytetyn antibakteerisen lääkinnällisen laitteen, Lumoral®-kaksoisvalohoidon, tehoa ja turvallisuutta plakin hallinnassa ja ienterveydessä julkisessa terveydenhuollossa. 40 molempaa sukupuolta edustavaa, 37–77-vuotiasta koehenkilöä, joilla oli diagnosoitu parodontiitin vaihe II tai korkeampi, jotka harjaavat hampaitaan päivittäin ja käyttävät hammaslankaa tai hammasharjaa, satunnaistettiin 1:1 kontrolli- ja tutkimusryhmään. Kaikilta koehenkilöiltä kerättiin aMMP-8 ja mikrobiologiset näytteet ja kliiniset parametrit, kuten PPD, BOP, VPI, sekä lähtötilanteessa että tutkimuksen lopetuskäynnillä. Tutkimusryhmä sai aPD-hoitoa päivittäin noin 5 viikon ajan. aPD-hoito suoritettiin CE-merkityllä Lumorinse®-suuvedellä yhdessä CE-merkityn Lumoral®-valoaplikaattorin kanssa. Suuvettä purskuteltiin minuutin ajan, jonka jälkeen annosteltiin 10 minuutin ajan samanaikaisesti 405 nm aBL ja 810 nm lähi-infrapuna (NIR) LED-valoa, kokonaissäteilyaltistuksen ollessa 40 J/cm2. Seurantakäynnit suoritettiin keskimäärin 7 viikon kuluttua lähtötilanteen arvioinnista, jolloin tutkimusryhmässä oli keskimäärin 8 päivän hoitotauko. Tutkimustulokset viittaavat Lumoral® aPDT -hoidon käytön vähentävän ienverenvuotoa, auttaen hoitamaan tulehduksellisia suusairauksia, kuten parodontiittia, peri-implantiittia ja ientulehdusta. Suurempi hyöty on osoitettu tupakoimattomille henkilöille. Tutkimustulokset viittaavat tupakoimattomien henkilöiden, joilla on diagnosoitu vaiheen II tai III parodontiitti, voivan hyötyä Lumoral® hoidosta arvioitaessa syviä ientaskuja. Tutkimus osoittaa potilaiden, joilla on vaiheen II tai sitä korkeampi parodontiitti ja joilla on jo kohtalaisen hyvät kotihammashoitorutiinit, olevan motivoituneita näkemään ylimääräistä vaivaa hoitonsa parantamiseksi. Lumoral®-hoidon tehokkuuden arviointia syvien ientaskujen paranemisessa varten tarvitaan lisätutkimuksia. Lumoral®-hoidon vaikutusta hammasplakkiin, parodontiitin vaiheeseen ja mikrobiologiseen taakkaan tulee varmistaa lisätutkimuksin. Tupakkatuotteiden vaikutus Lumoral®-hoidon tehoon tulee vahvistaa. Optimaalista käyttötiheyttä ja Lumoral®-hoidon kestoa tulee tutkia lisää. Lumoral®-hoidon hyöty potilailla, joilla on vaikea vaiheen IV ja asteen C parodontiitti, tulee varmistaa kohdennetuin tutkimuksin.
  • Ignatius, Adele (2021)
    Misfolding and aggregation of alpha-synuclein (α-syn) protein, leading to dysfunctional proteins and toxic protein aggregates, are seen as major factors in the pathogenesis of Parkinson’s disease (PD). Direct protein-protein interactions (PPI) between α-syn and a serine endopeptidase, prolyl oligopeptidase (PREP), have been shown to increase α-syn aggregation. Small molecular PREP inhibitors, in turn, have been shown to reduce the ɑ-syn aggregation process both in vitro and in vivo. Inhibition of PREP has been shown to have dual effects on ɑ-syn aggregation: first of all, blocking PREP mediated seeding and secondly, inducing the clearance of ɑ-syn aggregates via increased autophagy. Thus, PREP inhibitors should be further studied as a potential treatment for PD and other synucleinopathies. In this study, we evaluated the effect of two different PREP inhibitors, 4-phenylbutanoyl-L-prolyl-2(S)-cyanopyrrolidine (KYP-2047) and HUP-115 in a virus vector-based unilateral A53T-ɑ-syn overexpression mouse model. AAV-A53T-ɑ-syn injections used in this study caused a significant increase in oligomer-specific alpha-synuclein (ɑ-synO5) immunoreactivity and a mild dopaminergic neuron loss, together with mild motor deficits. Neither 2-week PREP inhibition with KYP-2047 or 4-week PREP inhibition with HUP-115 reduced ɑ-synO5 immunoreactivity or protected dopaminergic neurons in the substantia nigra (SN). Concordant to this, the treatments did not restore the slight behavioral deficit AAV-A53T-ɑ-syn injections caused in the cylinder test. In previous studies, PREP inhibition with KYP-2047 decreased ɑ-synO5 immunoreactivity, attenuated dopaminergic neuron loss and restored behavioral deficits in other α-syn overexpression mouse models. It is suggested that PREP inhibitors mainly have an effect on soluble ɑ-syn oligomers, rather than insoluble fibrils. In case A53T-ɑ-syn forms insoluble fibrils too rapidly in mice, overexpression of A53T-ɑ-syn might not be a suitable option when studying the effects of PREP inhibitors. Our results suggest that further characterization of this model in mice is much needed before drawing any conclusions about the effect of these PREP inhibitors.
  • Jaskari, Iida (2022)
    Multiple sclerosis is a progressive inflammatory disease of the central nervous system that affects young adults. The pathological hallmark of MS is the degradation and loss of oligodendrocytes resulting in demyelination. Damage to axons caused by demyelination severely impairs physical function. Currently there is no cure for MS, but current drugs aim to modify the course of the disease and relieve symptoms. However, they are unable to promote the repair of damaged myelin sheaths, and thus new therapies are needed. In this study, the effect of V-MANF on remyelination was investigated in two commonly used experimental toxin models. V-MANF is a modification of the endoplasmic reticulum located protein MANF, which has been found to have neuroprotective and regenerative properties. Additionally, MANF can regulate ER stress, which contributes to demyelination in MS. The effect of V-MANF on lysolecithin-induced demyelination was examined in organotypic cerebellar brain sections from C57B/6 mice. The study was conducted exceptionally using the brains of adult mice because they are a better model for neurodegenerative diseases. However, when analyzing the results, it was found that there was no demyelination in the tissue cultures, so the effect of V-MANF could not be analyzed. In the other study, C57B/6 mice were given dietary cuprizone for six weeks, followed by daily intranasal administration of either V-MANF or vehicle for seven days. Mice were subjected to behavioral experiments, in which a light/dark box test showed that V-MANFs had a potential anxiolytic effect in mice receiving cuprizone. No significant demyelination was observed by immunohistochemical analysis and therefore the effect of V-MANF on remyelination could not be assessed. However, the results of the study can be utilized in the design of further studies.
  • Turunen, Iida (2023)
    Metastatic prostate cancer is often fatal disease stage. Mechanism causing prostate cancer remains unknown, but possible mechanism relies on hormones. Testosterone may activate spontaneous cell division of oncogenes. Prostate cancer cells require androgen cell stimulation of AR to grow in early stages of prostate cancer, approximately 80-90% of prostate cancer cases are androgen dependent. 3bHSD1, encoded by HSD3B1, catalyzes the conversion of dehydroepiandrosterone to androstenedione and further to T and DHT. SNP (1245A to C) in HSD3B1 changes asparagine to threonine in position 367 resulting the enzyme accumulation and increased function. With androgen deprivation therapy castrate levels of testosterone are often achieved and it induces positive response in most PCa patients, but the polymorphism of 1245C is related with lower survival rate and higher probability for PCa to develop into CRPC. The aim of this study was to find out the effect of SNP in 3bHSD1 to androgen levels in patients treated with ADT. 32 patients were first genotyped based on SNP in the HSD3B1 gene (rs1047303) with 96.9 % success rate. 21 patients represented genotype AA, 9 AC and 1 patient CC. Other mutation in rs6203 was also detected. Genotyping was done by isolating DNA from blood samples and preparing it further for Sanger sequencing. Steroid analysis was performed by using LC/MS, using liquid-liquid extraction as sample preparation method. Altogether 21 steroids were analyzed from serum samples. Samples were taken every 3 months, during 33 months period for longest. The concentrations of T and DHT were reduced in AA genotype group after ADT as was expected to happen in all of the groups. In fact, the only significant changes were seen in AA genotype with for example the concentrations of previously mentioned T, DHEA and also A4. The changes in measured androgen levels cannot be generalized to concern especially the CC genotype, as there was only one patient homozygote with the mutation. Even though these results gave promising data of possible androgen synthesis pathways, a similar study must be rerun with larger patient data to be sure of the characteristics of different genotypes. Also, the effect of SNP in rs6203 remains still unknown.
  • Niittymäki, Erika (2021)
    Since the discovery of ketamine’s antidepressant response, numerous of studies have been observed it to alleviate depressive symptoms rapidly and effectively within hours. This is a significant advantage compared to traditional antidepressants, which take weeks to show treatment efficacy. Ketamine is a N- methyl-D-aspartate receptor (NMDA) antagonist and its underlying mechanism of is proposed to be in its ability to increase synaptic plasticity and this is ultimately believed to improve mood. On a molecular level, the antidepressant effects have been observed to be dependent on the activation of tropomyosin receptor kinase B (TrkB) signalling pathway. However, the antidepressant mechanism of ketamine remains still poorly understood as no new NMDA-antagonist or other rapid-acting antidepressants have been successfully developed for clinical use despite many years of effort. Therefore, some have proposed that the missing pieces of understanding its antidepressant effects might be linked to ketamine’s ability to modify sleep patterns and circadian-related molecules. Ketamine has especially been demonstrated to increase slow-wave activity during the following night of treatment and these changes have been shown to predict the clinical outcome in patients with major depressive disorder (MDD). Slow-wave activity is a low-frequency and high-amplitude wave seen in electroencephalography, which is highly expressed during the deepest stage of sleep, and this has been prominently found to be reduced in MDD patients. Even more intriguing, there are indications that ketamine might increase slow-wave activity also immediately after its administration. During this time, TrkB signalling is observed to became active. Following these molecular findings, we sought to investigate the link between the TrkB signalling pathway and two prominent processes occurring during slow-wave sleep. During slow-wave sleep processes such as (1) reduction of brain’s energy expenditure and (2) the activation of glymphatic system is known to occur. The glymphatic system is as lymphatic-perivascular network, which is responsible for clearing the brain from the metabolic waste. Thus, in this study, our objective was to investigate whether by causing an acute decline in adenosine-triphosphate (ATP) production or by stimulating the glymphatic network, we could activate the same plasticity-related pathways as ketamine is capable of activating in mice prefrontal cortex. The results of this study suggest that acute metabolic reduction can trigger pathways regarding synaptic plasticity. The metabolic inhibitor, 2-deoxy-D-glucose and mercaptoacetate (2DG+MA), was found to phosphorylate the TrkB receptor and its downstream signalling molecules GSK3β and p70S6K, while MAPK was dephosphorylated. These results correlate with the previous findings of ketamine’s effect after its administration. We also found a plasticity-related marker, MAP2, to be heavily phosphorylated by 2DG+MA, indicating 2DG+MA having a surprising role on neuroplasticity. These results are promising indication of understanding the rapid effects of ketamine and might even give important insight to developing novel antidepressants. However, these findings are only preliminary, and more research is needed to directly link antidepressant effects and energy metabolic inhibition together, as our study did not directly investigate antidepressants and depression-like behaviour in mice.
  • Snellman, Nana (2023)
    Chlamydia pneumoniae is an intracellular Gram-negative bacterium, that can cause respiratory infections. Infections are typically mild or asymptomatic, but it can also lead to more severe infections, for example, pneumonia. Severe infections might need antibiotic treatment. When the bacteria are exposed to stressful conditions, they can change to a chronic, persistent form. Amoxicillin and penicillin are known to transform bacterium into persistent forms. Antibiotics are not effective for persistent infection very often. Amoxicillin is the recommended treatment for pneumonia in Finland and worldwide, which is problematic from the perspective of C. pneumoniae. That is why there is a need for effective treatment for persistent C. pneumoniae infection. Probiotics and their by-products short chain fatty acids (SCFAs) are known to have beneficial effects on human health. Based on the current knowledge, SCFAs and other probiotic by-products are known to inhibit pathogen bacterial growth. Thus, SCFAs could have a potential effect on the treatment of C. pneumoniae infection. The aim of this work is to study the impact of SCFAs, acetate, propionate, and butyrate on C. pneumoniae infection and its antibiotic susceptibility. To study the impact of acetate, propionate and butyrate on C. pneumoniae infection and its antibiotic susceptibility, three different infection models were used: productive C. pneumoniae infection model with A549 cells, amoxicillin-induced persistent infection model with A549 cells, and persistent infection model with THP1 cells. Bacterial growth was followed with immunofluorescence and the number of the bacterial genome was studied with quantitative polymerase chain reaction (qPCR). The studied SCFAs did not have a significant impact on productive C. pneumoniae infection. With amoxicillin- induced persistent infection, the results were varying. For example, sodium acetate, and propionate showed some increase in bacterial growth on the first infection, but with sodium butyrate, there were not any impact. The only SCFA that decreased the bacterial growth in the persistent infection model with THP1 cells was sodium butyrate (200 μM). The same treatment also decreased the number of bacterial genomes with qPCR in the same infection model. In addition, the same condition increased the antibiotic susceptibility of persistent C. pneumoniae to azithromycin in THP1 cells. In conclusion, the studied SCFAs seemed to have more impact on C. pneumoniae infection with human immune cells compared to human lung epithelial cells. Based on this study, sodium butyrate could have positive impacts against persistent C. pneumoniae infection. Nevertheless, further studies of the impact of sodium butyrate on persistent C. pneumoniae infection are needed.
  • Savolainen, Heikki (2018)
    Tablet manufacturing requires both high-quality equipment and powder blend with high flowability and compactability and low segregation tendency. The process is complex and tablet formation process still remains not fully understood. Adequate powder flow is a necessity for the pharmaceutical manufacturing process, i.e., powder flowability and flow properties play a great role when designing manufacturing processes for solid dosage forms. As such, the powder characteristics need to be investigated. However, one property is seldom enough to predict the flowability of a powder in specific processes and different test methods need to be used to fully understand the tableting performance of a particular powder. It is crucial to know how the assessed properties reflect the manufacturing conditions. The need for test batches and the use of empirical testing still exists despite the numerous powder characterization tests available. The main aim of the study was to understand the influence of material properties, flow properties and segregation tendencies on both the processability of a formulation during tablet compression and the critical quality attributes, such as mass, tensile strength and dose uniformity of the final drug product. Additionally, testing of an in-line NIR method to observe the homogeneity of the powder inside the force feeder right before the compression step and transmission Raman as an at-line method for tablet content were also evaluated. A number of powder characterization tests were employed in order to fully understand the impact of the formulation on the process performance. Three formulations with different particle size of the active substance and mannitol were used throughout the study. Both the sifting segregation and fluidization segregation tests’ results predicted the formulations’ tabletability particularly well. Fluidization segregation test predicted the changing composition of the formulation throughout tableting whereas sifting segregation results showed the constantly fluctuating API concentration in the manufactured tablets. Moreover, the Raman results confirmed the tablets of variable content despite the offset caused by the different particle size of the raw materials used. The functionality of the NIR in the force feeder was tested successfully. The residence time distribution could be determined at a sufficient level to point out tablets of a bad quality from the batch on grounds of the NIR data. Results from the powder flow property tests were rather conflicting. Angle of repose, Carr’s index and volume flow rate gave the best characterizing results, whereas the mass flow rate, shear test with higher normal stress in pre-shear gave the worst results, considering the experienced flow character of the formulations. As stated above, different flow property tests may give conflicting result, and hence, it is crucial to know which results are the most relevant ones. Furthermore, the right settings for the test should be known to gain applicable results, best exemplified by the shear cell test.