Natural products have been crucial globally in treating and preventing fungal infections and diseases with significant inflammatory components, with numerous studies highlighting the therapeutic potential of these metabolites.[1] Herbs and spices are vital resources, commonly used in everyday life as food additives, flavor enhancers, fragrances, pharmaceuticals, colorants, or directly in medicinal applications. The practice of utilizing plants in these ways has a long-standing history worldwide, and over the centuries, humans have improved techniques for extracting phytochemicals and essential oils from these natural sources. Essential oils consist of complex mixtures of volatile compounds, typically present in low concentrations. They are intricate mixtures of volatile compounds that are usually found in low concentrations. Before these compounds can be analyzed, they must be extracted from their source material. [2] Several methods are available for extraction, including hydro-distillation (HD), steam distillation, Soxhlet extraction, and simultaneous distillation–extraction. However, these molecules are known to be sensitive to heat and prone to chemical alterations, making careful extraction crucial. [3] 

Phytochemicals are essential for creating therapeutic agents with natural alternatives. A novel method for identifying medicinally active substances in different plant species is the screening of plant extracts. Flavonoids, tannins, saponins, alkaloids, and terpenoids are examples of phytochemicals with a variety of biological characteristics, such as antioxidant, anti-inflammatory, anti-diarrhea, anti-ulcer, and anticancer effects. [4, 5].

MATERIALS AND METHODS

Materials

The spices such as Anise seeds, Ajwain seeds, Bay leaves, Dill seeds, Coriander seeds, Kalonji seeds and herbs such as Dill leaves, Scallion (Spring onion leaves) were identified and collected from the local market. The herbs and spices were air dried at room temperature and crushed into fine powder for further extractions. Analytical grade chemicals were used for the research work.

Methodology

I. Extraction of phytochemicals

Phytochemicals were extracted using Soxhlet’s distillation method. 100 g of dried powdered samples were extracted using 200 ml of ethanol in Soxhlet apparatus for 48 hours. The crude extracts were concentrated by rotary vacuum flash evaporation, weighed and  stored  at refrigerated condition [6, 7].

II. Extraction of Essential oils

500 grams of dried samples in powdered form were subjected to hydrodistillation in a Clevenger apparatus for 4-5 hours. The essential oil was collected, dried under anhydrous sodium sulphate and stored at 00C. [7]

III. Phytochemical Screening

Phytochemical screening of secondary metabolites such as alkaloids, flavonoids, terpenoids, saponins, phenolic compounds and tannins, phytosterols, carbohydrates, starch, fixed oils and fats were carried out using the standard  procedures as described by Harborne, [8]

Khandelwal, [9] and Daniel M. [10]

IV. GC-MS analysis

The phytochemicals and  essential oils were analysed by gas chromatography coupled to mass spectrometry (GC–MS) (Perkin Elmer Clarus 600 Gas chromatograph) using a fused-silica-capillary column with an apolar stationary phase GSBP-5MS (30M x 0.25mm x 0.2 micrometer).

GC–MS were obtained using the following conditions: carrier gas He (99.99%); flow rate 1.0 mL/min; split ratio 10:1; injection volume 1 micro L; injection temperature 280°C; Initial oven temperature was 50°C which progress to 180°C at 5°C/min, from 180 to 260°C at 5°C/min, from 260 to 280°C at 6°C/min and holding at 280°C for 40 min; the ionisation mode used was electronic impact at 70 eV. The detector used was Single Quadrupole mass analyzer. Identification of the components was achieved from their linear retention indices on GsBP-5MS column, determined with reference to an homologous series of C8–C22 n-alkanes, and by a comparison of their mass spectral fragmentation patterns with those stored in the data bank NIST (National Institute of Standards and Technology) Library and FFNSC 3 (Flavors and Fragrances of Natural and Synthetic Compounds) Wiley Library and the literature. [11, 12]

RESULTS AND DISCUSSION

Extraction Yields

The yield of ethanolic extracts and essential oils obtained is represented in Figure 1 and Figure 2 respectively. Different spices and herbs were used to prepare the ethanolic extracts and the yield of phytochemicals in all spices and herbs were compared. Use of ethanol gave good yield for ajwain seeds (42%), followed by coriander seeds (38%), dill leaves (34%), bay leaves (31%), dill seeds (27%), kalonji seeds, (22%), anise seeds (18%) and scallions (14%).   The yield of essential oil obtained after hydrodistillation was highest for kalonji seeds (11%) and ajwain seeds (4.3%).  Dill seeds (2.6%), bay leaves (2.5%), anise seeds (1.8%) and coriander seeds (1.3%) gave an average yield, whereas very poor yield of essential oils was obtained with scallions (0.9%) and dill leaves (0.6%). The total yield of the essential oils of different spices and herbs coincide with the findings of literature papers. [13] The phytochemical yield of spices and herbs is influenced by a variety of factors, including species, cultivation and harvesting practices, and extraction techniques. Previously carried out research consistently shows that optimal growing conditions, correct harvest timing, dying methods and suitable extraction techniques can enhance the yield of bioactive compounds such as flavonoids, phenolics, terpenoids, and essential oils.[13]

       
           
       
Fig 1: Yield of Phytochemicals in ethanolic extract

The qualitative phytochemical screening results for eight ethanolic extracts prepared using commercial spices and herbs is outlined in Table 1

The results of the qualitative screening indicates that all the tested spices and herbs namely,  Anise seeds,  Ajwain seeds,  Bay leaves, Coriander seeds, Dill leaves, Dill seeds, Kalonji seeds,  and Scallions are rich in phytochemicals such as alkaloids, flavonoids, terpenoids, phenols and tannins. Kalonji seeds extract showed an abundance of fixed oils and fats. The extracts of coriander seeds and kalonji seeds revealed the absence of phytosterols; whereas absence of terpenoids was reported in the extracts of coriander seeds and dill seeds.

Table 1: Qualitative screening of Phytochemicals in ethanolic extracts

Key- + positive/ presence of test component, - = Negative/ Absence of test component

Similar results were reported for phytochemical screening of spices and herbs.^[14] The present study showed abundance of tannins, organic acids and phenolic acid derivatives in the spices and herbs..  In general, phenolic acids and their derivatives show good anti-oxidation potential, anti-inflammatory and antimicrobial properties.^[15] The phytochemical constituents of plants can vary based on soil conditions (pH, acidity, fertility, microbiome and moisture), environmental factors (temperature, humidity) during growth or variation in cultivars. These plants contain a wide variety of complex bioactive chemicals that have anti-inflammatory, anti-cancer, anti-oxidant, and antibacterial qualities, among other important health advantages. ^{[15, 16]}

GC- MS analysis of phytochemicals and essential oils

The phytochemicals and essential oils of the eight plants were found to contain a wide variety of chemical constituents. The major compounds in each of them were identified as listed in Table 2. Ajwain exhibited a high presence of thymol. Similarly, Bay leaves showed the presence of anethole. Anise was found to contain linalool and pinene, a potent compound known for its aromatic properties and digestive benefits. Coriander seeds contained cymene, while Dill seeds and leaves exhibited a high content of apiole and carvone, both of which have antimicrobial properties. Scallions demonstrated the presence of diallyl sulfide and kalonji seeds contained apiole, both of which are responsible for the aromatic and medicinal properties of these plants.

Table 2 GC-MS Analysis of ethanolic extracts and essential oils