Razvoj analizne metode za ugotavljanje vsebnosti terpenskih spojin v neobdelanih cvetovih navadne konoplje (Cannabis sativa, L.) z uporabo plinske kromatografije sklopljene z masno spektrometrijo

Tušar, Iza (author), Tavčar, Eva (mentor), Schoss, Katja (comentor)

Količina in vrsta terpenskih spojin med različnimi varietetami konoplje (Cannabis sativa L.) variirata, saj gre za genotipsko kot tudi fenotipsko pogojene lastnosti. Njihov profil postaja vedno zanimivejši v farmaciji, predvsem zaradi možne sinergije s kanabinoidi. Namen te naloge je bil razviti analizno metodo za ugotavljanje vsebnosti terpenskih spojin v neobdelanih cvetovih konoplje z uporabo plinske kromatografije sklopljene z masno spektrometrijo in pridobiti profil terpenskih spojin v različnih vzorcih cvetov konoplje. Tekom razvoja metode za snemanje rastlinske droge konoplje brez predpriprave vzorca smo običajen GC-MS sistem povezali z napravo OPTIC-4, ki kot sistem nove generacije vnosa vzorca na kolono omogoča prilagoditev različnih oblik (trdnih in tekočih) vzorcev za analizo. Z uporabo temperaturnega programa segrevanja vzorca v injektorju smo poskrbeli, da je nehlapen matriks ostal v mikroviali, hlapne spojine pa so potovale na kolono. Pod enakimi pogoji kot vzorce neobdelanih cvetov konoplje smo z OPTIC-4 ločeno snemali tudi referenčne raztopine terpenskih spojin. Pri tej metodi je bilo injiciranje na kolono direktno, kar pomeni večjo občutljivost metode. Referenčne raztopine terpenskih spojin smo poleg tega snemali tudi s klasično metodo GC-MS, kjer injiciranje na kolono ni bilo direktno, saj nismo operirali z OPTIC-4 napravo, temveč smo uporabili klasično injiciranje z razdelitvijo vzorca. Pogoje obeh metod smo zastavili tako, da smo na koncu lahko primerjali retencijske čase terpenskih spojin med metodama. Z razvito metodo smo v šestih preiskovanih vzorcih, varietetah konoplje s prevladujočim kanabidiolom, skupno zaznali 44 različnih terpenskih spojin. 16 spojin smo identificirali s pomočjo standardov. Retencijski časi, ki smo jih snemali s klasično metodo GC-MS, so variirali za RSD ⡤ 3,03 %. V preiskovanih vzorcih različnih varietet konoplje so se v največjem deležu nahajali β-kariofilen, α-pinen ali linalol. Delež monoterpenskih spojin v vzorcih je variiral od 24,6 % do 60,1 %, delež seskviterpenskih spojin pa od 39,6 % do 75,0 %. Prednost razvite metode je, da se izognemo ekstrakciji, kjer bi lahko prišlo do izgub analitov, poleg tega pa za analizo zadostuje že približno 1 mg rastlinske droge, kar je pomembno pri težje dostopnih rastlinskih drogah.

• Language: Slovenian
• Keywords: terpenske spojine, konoplja, analiza GC-MS
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2024
• PID: 20.500.12556/RUL-160096
• Publication date in RUL: 20.08.2024

Secondary language

• Language: English
• Title: Development of analytical method for content determination of terpenes and terpenoids in raw cannabis flowers (Cannabis sativa, L.) using gas chromatography coupled to mass spectrometry

Abstract

Terpene and terpenoid composition is a genotypic and also phenotypic trait that shows much variation across different cannabis varieties. Their profile in cannabis (Cannabis sativa L.) is becoming increasingly interesting in pharmacy, mainly due to the possible synergy with cannabinoids. The aim of this thesis was to develop an analytical method for content determination of terpenes and terpenoids in raw cannabis flowers using gas chromatography coupled to mass spectrometry and to obtain their profile in different samples of cannabis flowers. Attaching a next-generation multimode inlet, OPTIC-4, to a standard GC-MS system made it possible to analyze both solid and liquid samples. During the development of the method for analyzing raw cannabis material, a microvial containing the sample was heated at the injection port, leaving non-volatile impurities behind in the microvial, while volatile compounds traveled to the column. Under the same conditions as the raw cannabis flowers samples, we also separately recorded standard solutions of terpenes and terpenoids using OPTIC-4. In this method, we used direct column injection, which means greater method sensitivity. Additionally, we recorded standard solutions of terpenes and terpenoids using the classic GC-MS method, where split injection was used without the OPTIC-4 inlet. The conditions of both methods were set so that we were able to compare the retention times of terpenes and terpenoids between the methods. With the developed method, we detected a total of 44 different terpenes and terpenoids in six analyzed samples of cannabidiol-dominant cannabis varieties. 16 compounds were identified using standards. Retention times recorded using the classic GC-MS method varied with RSD values of ⡤ 3,03 %. In the analyzed samples of different cannabis varieties, β- caryophyllene, α-pinene, or linalool were found in the largest percentages. The monoterpene and monoterpenoid content in the samples varied from 24,6 % to 60,1 %, and the sesquiterpene and sesquiterpenoid content varied from 39,6 % to 75,0 %. The advantage of the developed method is that it avoids extraction, which could lead to losses of analytes. Additionally, the method only requires approximately 1 mg of plant material for analysis, which is important when dealing with herbal drugs that are harder to obtain.

• Keywords: terpenes and terpenoids, cannabis, GC-MS analysis