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Naravne snovi so spojine ali ekstrakti, ki izvirajo iz živih organizmov ter navadno izkazujejo biološko delovanje. So osnovna surovina za zdravila naravnega izvora. V farmakognoziji se ukvarjamo z njihovim izborom, karakterizacijo, identifikacijo in uporabo. Pogost pristop k odkrivanju potencialno zdravilnih naravnih snovi je preizkušanje učinkov rastlinskih izvlečkov, ki nas v povezavi s kromatografskimi separacijami vodi do izolacije biološko aktivnih molekul. Njihove kemijske strukture ugotavljamo s sodobnimi spektroskopskimi in spektrometrijskimi metodami in nadaljujemo s preizkušanjem njihovega farmakološkega delovanja. Ajda predstavlja bogat vir hranil z ugodnim vplivom na zdravje, zato njena poraba v skladu s popularnostjo zdravega načina življenja narašča. Poleg zdravju koristnih snovi pa ajda vsebuje tudi manj znane, fototoksične fagopirine, ki pod vplivom svetlobe oksidirajo biološko snov in povzročijo njene poškodbe. Za zagotavljanje varnosti ajde in njenih izdelkov je treba fagopirine vrednotiti tako s prehranskega kot s farmacevtskega vidika. Ne nazadnje gre tudi za spojine s potencialno terapevtsko uporabnostjo, na primer v fotodinamični terapiji. Razvili smo učinkovit postopek ekstrakcije fagopirinov z uporabo raztopine acetona in vode. Ugotovili smo, da so najprimernejši material za pridobivanje fagopirinov ajdovi cvetovi. Razvili smo preprosto in občutljivo metodo tekočinske kromatografije visoke ločljivosti s fluorescentno detekcijo. Metoda je primerna za analizo vzorcev ajde in kot izhodišče za razvoj preparativne metode za izolacijo fagopirinov. Ugotovili smo, da je poleg osnovne molekule fagopirina F v zeli ajde še vsaj sedem njegovih derivatov. Te derivate smo kromatografsko ločili na posamezne frakcije. Vrednotili smo jih z UV-Vis absorpcijo, masno spektrometrijo in NMR spektroskopijo. Ugotovili smo strukturi dveh novih fagopirinov, ki se razlikujeta po skupinah, pripetih na naftodiantronski obroč. Fagopirin F je že predhodno znana spojina z dvema piperidinskima obročema, fagopirin E je spojina s piperidinskim in pirolidinskim obročem, fagopirin A pa je spojina z dvema pirolidinskima obročema. Dokazali smo, da v ajdi ni fagopirinov, temveč zgolj protofagopirini kot njihove predstopnje z necikliziranim naftodiantronskim obročem. Po raztopitvi v ekstrakcijskem topilu in pod vplivom svetlobe se protofagopirini pretvorijo v fagopirine. Raziskovali smo topnost, fluorescenco in UV-Vis spektre fagopirinov v različnih topilih iz različnih vzorcev ajde. Za kvantitativno vrednotenje vsebnosti fagopirinov je treba pridobiti čiste spojine kot referenčne spojine. Raziskovalci namesto fagopirina kot referenčne spojine uporabljajo hipericin iz šentjanževke, ki pa zaradi nekoliko drugačne strukture in s tem drugačne absorpcije pri spektrofotometrijski detekciji ni ustrezen za točno kvantitativno vrednotenje fagopirinov. Naše ugotovitve in novo razvite metode bodo pripomogle k nadaljnjemu raziskovanju fagopirinov in vrednotenju vsebnosti in varnosti ajdovih izdelkov. V drugem delu doktorskega dela smo se ukvarjali z izvlečki debla navadne jelke. Izvlečki debel različnih vrst borovcev so znani po vsebnosti polifenolnih spojin z zanimivimi farmakološkimi učinki, predvidoma povezanimi z antioksidativnim delovanjem. Najbolj znan, raziskan in uporabljan je izvleček lubja obmorskega bora (Pinus maritima), Pycnogenol®. Ugotovili smo, da so izvlečki navadne jelke učinkovitejši antioksidanti od Pycnogenola® tako v in vitro antioksidativnih testih, kot tudi na humanih celičnih linijah, izoliranih iz periferne krvi. Komponente izvlečka lubja smo kromatografsko ločili z normalnofazno kolonsko kromatografijo pri zvišanem tlaku in reverznofazno tekočinsko kromatografijo visoke ločljivosti. Vsebovane spojine smo identificirali z metodami masne spektrometrije, UV-Vis absorpcijske spektroskopije in s primerjavo z referenčnimi spojinami. Identificirali smo šest fenolkarboksilnih kislin (galno, homovanilno, protokatehujsko, p-hidroksibenzojsko, vanilno in p-kumarno), tri flavonoide (katehin, epikatehin in katehintetrametileter) in štiri lignane (taksiresinol, 7-(2-metil-3,4-dihidroksitetrahidropiran-5-iloksi)-taksiresinol, sekolariciresinol in lariciresinol). Nadaljevali smo s farmakološkim testiranjem. Ugotovili smo, da izvleček zmanjša nastajanje ateroskleroze pri budrah na kronični aterogeni dieti. Ovrednotili smo vpliv izvlečka in dveh v njem prisotnih fenolnih kislin, p-kumarne in protokatehujske, na izolirana podganja srca, podvržena akutnim ishemijsko-reperfuzijskim poškodbam. Ugotovili smo, da je izvleček značilno zmanjšal poškodbo srčnega tkiva, merjeno preko sproščanja laktat-dehidrogenaze in zmanjšal pojavnost aritmij. p-kumarna in protokatehujska kislina nista imeli tako izrazitih učinkov, zato verjetno samo delno prispevata k sinergističnemu delovanju v izvlečku prisotnih učinkovin. Kot ustrezno surovino za raziskovanje in uporabo smo oblikovali izvleček, raztopljen v polietilenglikolu. Pojavila se je potreba po ugotavljanju vsebnosti vode v tovrstnih vzorcih. Uveljavljena in dostopna Karl-Fischer metoda z opazovanjem spremembe barve pri končni točki titracije ni bila ustrezna, saj je bil izvleček obarvan. Zato smo razvili preprosto in finančno dostopno titracijsko metodo za ugotavljanje vsebnosti vode v obarvanih vzorcih, pri kateri končno točko titracije zaznamo spektrofotometrijsko. Tako ovrednoten izdelek predstavlja dobro osnovo za nadaljnje raziskovanje in razvijanje novih prehranskih dopolnil, kozmetičnih izdelkov in zdravil. Natural products are compounds or extracts that originate from natural sources and usually exhibit biological activity. They are raw materials for natural medicines. Pharmacognosy encompasses their selection, characterization, identification and use. Bioassay-guided fractionation of plant extracts linked to chromatographic separation techniques leads to the isolation of biologically active compounds whose chemical structures can readily be determined by modern spectroscopic and spectrometric methods, followed by testing of their pharmacological activity.Buckwheat contains many healthy nutrients, and its consumption is therefore increasing. The plant contains phototoxic compounds called fagopyrins that can act as photosensitisers upon excitation with visible light causing fagopyrism, but these compounds have received little research attention. To assure adequate safety of buckwheat and its products, fagopyrins should be characterized from a nutritional and pharmaceutical point of view. Nevertheless, fagopyrins are not only health-threatening compounds but also potent photodynamic agents that deserve further research. In this work, fagopyrins extraction procedure was improved employing a mixture of acetone and water. Flowers were recognized as the best material for the extraction. A chromatographic analysis of fagopyrins was performed by developing a simple, sensitive HPLC analytical method of high-resolution employing fluorescence detection. The developed method is a good starting point for a preparative method that would enable the isolation of pure compounds. At least eight fagopyrin derivatives were observed upon isolation and were characterized via UV-Vis absorption, NMR spectroscopy and mass spectrometry. The structures of two new fagopyrins were elucidated. Fagopyrin F is a previously known compound that consists of naphthodianthrone scaffold and two piperidine moieties. Fagopyrin A differentiates by two pyrrolidine moieties instead of piperidine moieties, and fagopyrin E consists of one pyrrolidine and one piperidine moiety. The existence of protofagopyrins that originate in buckwheat and can transform into fagopyrins upon light exposure was confirmed. Information on solubility, fluorescence and UV-Vis absorbance in different solvents were obtained. The need for reference compounds of different fagopyrins that would allow proper quantitative evaluation of buckwheat samples still remains. Researchers use hypericin for that purpose, but we recognize its use as inadequate due to the different structure and consequently uncomparable absorbance in spectrophotometric detection. Our methods complement the existing knowledge regarding fagopyrins and will facilitate their further isolation and analysis of their biological activity. In the second part of our work, silver fir trunk extracts were investigated. Extracts from the bark of different conifer species are known to contain various polyphenols that possess interesting pharmacological activities. So far the most extensive research was done on the antioxidative extract of the maritime pine (Pinus maritima) bark Pycnogenol®, which is widely used in food supplements and cosmetic products. We have shown that antioxidant activity of silver fir (Abies alba) extract is higher comparing to the maritime pine bark extract in vitro and in cultured cells. Components of the extract were separated with normal phase flash chromatography and reversed phase HPLC. Structures of individual compounds were elucidated by mass spectrometry, UV-Vis absorption spectroscopy and comparison to reference compounds. Six phenolic acids were identified (gallic, homovanillic, protocatehuic, p-hydroxybenzoic, vanillic and p-coumaric), three flavonoids (catechin, epicatechin and catechin tetramethyl eter) and four lignans (taxiresinol, 7-(2-methyl-3,4-dihydroxytetrahydropyran-5-yloxy)-taxiresinol, secoisolariciresinol and laricinresinol).The research was continued with pharmacological testing. It was found that the extract exerts strong antioxidative and protective effects against atherogenic diet-induced arterial wall damage. Protective effects of the silver fir extract and its compounds as well as of two phenolic acids (p-coumaric and protocatechuic acids) were also evauated against ischemia-reperfusion-induced damage in isolated rat hearts. The silver fir extract significantly reduced tissue damage (measured as the lactate dehydrogenase release rate) and shortened the duration of arrhythmias, which was not observed in the case of phenolic acids. It was concluded that the two acids are only partially responsible for the observed effects and synergism of more compounds could be present. Silver fir extract, solubilized in polyethylene glycol was developed as an appropriate raw material for research and use. A need for water determination in such samples emerged. The most accessible technique for water content determination is Karl-Fischer titration with visual detection of a titration endpoint, which was unfortunately not suitable for the analysis of the silver fir extract due to its strong colour. Therefore an appropriate method for moisture determination in coloured samples was developed employing simple equipment and spectrophotometric detection of a titration endpoint. Silver fir extract, characterized in such manner is a suitable ingredient for development of food supplements, cosmetic products and medicines.

Konoplja je rastlina, ki se že vrsto let uporablja v medicinske, kozmetične, prehranske in industrijske namene. Njena uporaba je bila v preteklosti marsikje prepovedana, danes pa je v večini evropskih držav dovoljena v medicinske namene. Poznana je po svojih učinkih na živčevje in imunski sistem, na primer pri bolečinah in vnetjih. Te farmakološke lastnosti so značilne predvsem za kanabinoide, to so molekule, ki jih najdemo v vseh delih rastline. Z ustreznimi topili jih lahko iz rastline ekstrahiramo in uporabimo. V nalogi smo se osredotočili na optimizacijo etanolne ekstrakcije kanabinoidov iz konoplje. Najprej smo izvedli etanolno ekstrakcijo kanabinoidov iz posušenih cvetov industrijske konoplje in določali vsebnost kanabinoidov v različnih frakcijah ekstrakta. Izkazalo se je, da je njihova vsebnost približno enaka v vseh plasteh v bučki. Nadaljevali smo s profilom ekstrakcije, pri čemer smo sprva želeli ugotoviti, če se količina ekstrahiranih kanabinoidov razlikuje glede na temperaturo ekstrakcijskega topila in v kateri časovni točki je vsebnost kanabinoidov največja. Najprej smo uporabili nemleto drogo, pri kateri so bili rezultati ekstrakcij med seboj precej neprimerljivi. Zato smo v nadaljevanju drogo homogenizirali. Ugotavljali smo vpliv hlapnosti etanola na vsebnost kanabinoidov v supernatantu, ter vpliv stresanja in ultrazvočne homogenizacije na učinkovitost ekstrakcije. Rezultati so pokazali, da je za večjo učinkovitost ekstrakcije bolje uporabiti homogenizirano drogo, zatesnjen vsebnik in vključiti stresanje. Ker je bila učinkovitost ekstrakcije po dotedanjih poskusih še vedno le nekaj več kot 75 %, smo poskusili z večkratnim spiranjem droge z etanolom in z ožemanjem droge. Z ožemanjem droge smo izboljšali metodo ekstrahiranja za približno 15 %, medtem ko večkratno spiranje droge ni pripomoglo k večji učinkovitosti ekstrakcije. Nazadnje smo s pomočjo pristopa Design of experiments in programa Modde postavili model ekstrakcije. Spremljali smo vpliv časa ekstrakcije, temperature topila pri ekstrakciji in ultrazvočne homogenizacije na učinkovitost ekstrakcije, torej deleža kanabidiola, kanabidiolne kisline in nečistot v ekstraktu. Pri tem smo ugotovili, da daljši čas, višja temperatura in daljša homogenizacija pri ekstrakciji povečujejo ekstrakcijo drugih snovi in tudi kanabinoidov, medtem ko poteka optimalna ekstrakcija kanabinoidov pri nizkih temperaturah (-24 °C), srednjem času (1-4 min) in brez ultrazvočne homogenizacije. Hemp is a plant that has been used for many years for medicinal, cosmetic, nutritional and industrial purposes. Its use was banned in many places in the past, but today it is allowed for medical purposes in most European countries. It is known for its effects on the nervous system and the immune system, for example in pain and inflammation. These pharmacological properties are mainly characteristic of cannabinoids, which are molecules found in all parts of the plant. With appropriate solvents, they can be extracted from the plant and used. Within master thesis, we focused on the optimization of cannabinoids ethanol extraction. First, we performed ethanol extraction of cannabinoids from dried flowers of industrial hemp and determined the content of cannabinoids in different fractions of the extract. It turned out that their content is approximately the same in all layers in the flask. We continued with the extraction profile, initially looking to determine if the amount of cannabinoids extracted varies with the temperature of the extraction solvent and at what time point the cannabinoid content is highest. First, a non-ground drug was used, and the results of the extractions were quite incomparable. Therefore, we further homogenized the drug. We determined the influence of ethanol volatility on the content of cannabinoids in the supernatant, as well as the influence of shaking and ultrasonic homogenization on the extraction efficiency. The results showed that for higher extraction efficiency it is better to use a homogenized drug, a sealed container and include shaking. Since the extraction efficiency after the previous experiments was still only slightly more than 75%, we tried repeatedly rinsing the drug with ethanol and wringing the drug. The extraction method was improved by wringing the drug fo approximately 15%, while repeated washing of the drug did not contribute to greater extraction efficiency. Finally, with the help of the Design of experiments approach and the Modde program, we set up the extraction model. We monitored the influence of the extraction time, the temperature of the solvent during extraction and the use of an ultrasonic homogenization on the extraction efficiency, the proportion of cannabidiol, cannabidiolic acid and impurities in the extract. We found that longer time, higher temperature and longer ultrasonic homogenization during extraction increase the extraction of other substances and also of the cannabinoids, while optimal extraction of cannabinoids takes place at low temperatures (-24 °C), medium time (1-4 min) and without ultrasonic homogenization.

Navadna konoplja (Cannabis sativa, L.) je rastlina, ki jo človeštvo s pridom izkorišča že tisočletja. V konoplji je prisotnih preko 100 kanabinoidov, med katerimi sta najpomembnejša kanabidiol (CBD) in tetrahidrokanabinol (delta-9-THC). Farmakološki učinek pri vretenčarjih dosežeta z delovanjem na kanabinoidne in nekatere druge receptorje. THC je psihotropna substanca in zaradi tega zakonodajno regulirana kot prepovedana droga v veliko državah. Drugače velja za CBD, ki ni psihoaktiven in zato bolj varen za uživanje. V zadnjih desetih letih se je njegova uporaba v farmaciji, hrani in kozmetiki močno povečala. Vedno bolj je tudi poznan javnosti. Na tujih trgih sta že 2 zdravili, ki vsebujeta CBD (Epidyolex in Sativex). Kljub temu je v literaturi malo podatkov, ki opisujejo njegovo topnost v različnih topilih. Zato smo v magistrski nalogi določili topnost CBD v topilih, ki so za njegovo uporabo najbolj pomembna. Najprej smo raziskali spletni trg izdelkov s CBD, pri katerih so prevladovale kapljice in pršila, da smo ugotovili, katera topila se najpogosteje uporabljajo. Zadetke smo v spletnem brskalniku iskali v slovenščini in angleščini. Največkrat so pregledani izdelki kot nosilna topila vsebovali MCT olje, olje konopljinih semen, olivno olje in njihove zmesi. Zato smo topnost določili v vseh treh omenjenih oljih. Poleg tega smo topnost določili tudi v čijevem olju, propilenglikolu (in njegovih vodnih raztopinah), trietil citratu, PEG 400, triacetinu, 96% etanolu, metanolu, izooktanu, acetonu, izopropanolu, DMSO, 1-butanolu, DMF, cikloheksanu, diklorometanu in dietiletru. Za določanje topnosti smo uporabili »shake-flask« metodo, pri čemer smo pripravili suspenzije CBD (na sobni temperaturi), jih stresali 48 h na stresalniku, filtrirali, redčili in določili koncentracijo CBD s HPLC metodo. CBD je v vodi zelo slabo topen, vendar to ne velja za večino drugih topil, v katerih smo določali topnost. Najbolje so ga raztapljali aceton, metanol in dietileter ter le malenkost slabše ostala laboratorijska topila. Tudi v topilih, ki se uporabljajo za izdelavo izdelkov, je bil CBD zelo dobro topen. Med njimi je bila topnost najvišja v triacetinu, PEG 400 in trietil citratu. Nekoliko slabša je bila v propilenglikolu in še nižja v rastlinskih oljih. V vodnih raztopinah propilenglikola se je izkazalo, da že nizka vsebnost vode močno zniža topnost CBD. Med rastlinskimi olji je bil CBD najbolje topen v MCT olju, najslabše pa v olivnem olju. Cannabis (Cannabis sativa, L.) is a plant that has been used by mankind for thousands of years. It contains over 100 cannabinoid molecules, the most important of which are cannabidiol (CBD) and tetrahydrocannabinol (delta-9-THC). They achieve their pharmacological effects on vertebrates by acting on cannabinoid receptors and some other receptors. THC is a psychotropic substance and is therefore highly regulated as an illicit drug by most authorities around the world. CBD on the other hand is not psychotropic and has a more favorable safety profile. Over the last ten years, its use has increased significantly as has its recognition among the general public. There are already two CBD-containing medicines on the market (Epidyolex and Sativex). Nevertheless, there is little data in the literature describing its solubility in various solvents. Therefore, we determined the solubility of CBD in the solvents that are most important for its use. Firstly, we researched the online market of CBD products (drops and sprays) to find out which solvents are most commonly used. We searched for results in the web browser in Slovene and English language. The reviewed products most often contained following carrier solvents: MCT oil, hemp seed oil, olive oil, and their mixtures. Therefore, we determined the solubility in all 3 mentioned oils. We also measured the solubility of CBD in chia seed oil, propylene glycol, and its aqueous mixtures, triethyl citrate, PEG 400, triacetin, 96 % ethanol, methanol, isooctane, acetone, isopropanol, DMSO, 1-butanol, dimethylformamide, cyclohexane, dichloromethane, and diethyl ether. Solubility was determined by using shake-flask method. Firstly, CBD suspensions were prepared (at room temperature), then shaken for 48 h on a shaker, filtered, and diluted. Then the CBD concentration was determined by the HPLC method. CBD is very poorly soluble in water in contrast to most other solvents we tested. Measured solubility was highest in acetone, methanol, and diethyl ether, followed by other laboratory solvents. CBD was also very soluble in the solvents that are often used for production of CBD products. Among them, solubility was highest in triacetin, PEG 400, and triethyl citrate. It was slightly worse in propylene glycol and even lower in vegetable oils. By adding water to propylene glycol, we discovered that even a small addition of water greatly reduces the solubility of CBD. Among vegetable oils, CBD was most soluble in MCT oil and least soluble in olive oil.

Dosedanje raziskave potrjujejo obstoj funkcionalnega endokanabinoidnega sistema v koži, ki med drugim vpliva na pravilno in uravnoteženo proliferacijo, diferenciacijo in toleranco kožnih celic ter regulacijo imunskega in vnetnega odziva. V zadnjem obdobju se zato izvaja vedno več raziskav topikalne uporabe kanabidiola, ki je eden izmed kanabinoidov, prisotnih v konoplji. Poleg pozitivnega vpliva kanabidiola na kožo, ugotavljajo tudi pozitivne učinke v ustni votlini, saj kanabidiol izkazuje obetavne baktericidne lastnosti proti aerobnim bakterijam v zobnih oblogah, ki so glavni vzrok za pojav različnih zobnih težav in bolezni. Izvedli smo dvojno slepo randomizirano in s placebom nadzorovano raziskavo kozmetičnih izdelkov s kanabidiolom. Preizkušali smo vlažilni in oljni serum za obraz, ustno vodo in peno za intimno nego. V raziskavi je sodelovalo dvajset prostovoljcev, ki so štirinajst dni preizkušali kozmetične izdelke s kanabidiolom in štirinajst dni placebo kozmetične izdelke. Namen raziskave je bil ugotoviti, kako uporaba kozmetičnih izdelkov s kanabidiolom, po mnenju prostovoljcev, vpliva na kožo, sluznico intimnih predelov in stanje v ustni votlini v primerjavi s placebom. Po preizkušanju vseh naštetih kozmetičnih izdelkov smo s pomočjo vprašalnikov pridobili in primerjali subjektivne ocene prostovoljcev ter ugotavljali učinkovitost vlažilnega in oljnega seruma za obraz z uporabo fotografij. Prostovoljci so v povprečju ocenili, da sta imela seruma za obraz s kanabidiolom v primerjavi s placebom večji vpliv na izboljšanje elastičnosti, vlažnosti in gladkosti kože na obrazu ter da sta izboljšala suhost kože, zmanjšala trenutno prisotnost rdečice in malo zmanjšala vidnost gub ter por na obrazu. Pri primerjavi ustnih vod so ocenili, da med njima ni razlik. Pri intimnih penah so v povprečju ocenili, da ima placebo večji pozitiven vpliv na pekoč občutek in občutek suhosti na intimnih predelih. Učinkovitost serumov za obraz smo preverjali tudi s pomočjo fotografij obrazov prostovoljcev, katere smo obdelali v aplikaciji Vichy SKINCONSULT AL®, ki ovrednoti številne parametre na koži obraza. Po statistični obdelavi smo sklenili, da aplikacija ni bila primerna za ugotavljanje učinkovitosti, saj je preveč občutljiva na svetlobo in kot fotografiranja obraza, v naši raziskavi pa ta parametra nista bila konstantna, saj fotografiranj nismo izvajali v ponovljivih razmerah. Poleg tega so ocenjevalci poskušali oceniti, katere fotografije so bile pridobljene po uporabi serumov s kanabidiolom, in tako ugotoviti, če se po uporabi kažejo vidni učinki na izgled kože obraza. Izkazalo se je, da ta način ocenjevanja zaradi kakovosti fotografij ni bil primeren za pridobitev relevantnih rezultatov. Studies have confirmed that there is a functional endocannabinoid system in the skin which plays a role in healthy and balanced proliferation, differentiation, tolerance of skin cells, and in the regulation of immune and inflammatory response, among other functions. For this reason, there is more and more research into the topical use of cannabidiol, one of the cannabinoids in cannabis. Besides cannabidiol’s beneficial effects on the skin, positive effects have also been observed in the oral cavity due to its promising bactericidal properties against the aerobic bacteria in dental plaques, the main cause of various dental problems. We performed a double blind, randomised, placebo-controlled trial of cosmetic products that contain cannabidiol. A moisturising face serum, an oily face serum, a mouthwash, and an intimate foam were tested. The study included 20 volunteers, who were using cosmetic products with cannabidiol for 14 days and placebo products for 14 days. The purpose of the trial was to assess how cosmetic products with cannabidiol affect the skin, the intimal mucosa and the condition of the oral cavity in comparison with placebo, according to volunteer reports. After completed testing of all the products we used questionnaires to record and compare the subjective opinions of the volunteers. Furthermore, we evaluated the efficacy of the moisturising and oily face serums with photography. On average, the volunteers estimated that the two cannabidiol face serums outperformed placebo in the improvement of face skin elasticity, hydration and smoothness. The serums also improved skin dryness, decreased redness, and led to a small reduction of visible wrinkles and pores on the face. For the mouthwash, no difference with placebo was observed on average. For the intimate foam, however, the placebo led to a greater average reduction of burning and dryness sensations in the intimal mucosa, according to the volunteer reports. The efficacy of the face serums was also studied by processing the photos of the volunteers' faces with the Vichy SKINCONSULT AL® application, which takes into account a number of facial skin parameters. After statistical analysis, we realised that the application is not an appropriate tool for evaluating efficacy, since it is overly sensitive to the lighting and the camera angle. These two parameters were not constant in our trial, since the photo shoots were not conducted in a laboratory. Moreover, the photos were searched to try to identify those taken after the use of the cannabidiol serums, in order to see if any effects on the appearance of facial skin can be observed. We concluded that this method of evaluation, due to the quality of the photographs, was not suitable for obtaining relevant results.