Naphthaleneacetic Acid Research Articles

Tissue Culture Response and In Vitro Plant Regeneration of Malus 'Baiyun' (a New Cultivar of Ornamental Crabapple).

Malus 'Baiyun' (registration no. 20210210), a new crabapple cultivar, was registered in 2021 by the Nanjing Forestry Unversity. However, the difficult rooting has greatly limited the production of high-quality M. 'Baiyun' in industrialization development. There is thus a pressing need to develop an organogenesis protocol for the in vitro propagation of M. 'Baiyun' to alleviate a shortage of high-quality M. 'Baiyun' seedlings. The results showed that choosing the apical bud in mid-March was an excellent explant material. To promote proliferation, the highest proliferation (6.27) of apical shoots was cultured on Murashige and Skoog (MS) medium supplemented with 0.5 mg·L-1 6-benzylaminopurine(6-BA) + 0.05 mg·L-1 indole-3-butyric acid (IBA). Subsequently, a 100% rooting rate, average number of roots per shoot of 6.2 and maximum length of roots of 4.96 cm were obtained on half-strength Murashige and Skoog (1/2 MS) medium with the application of 0.5 mg·L-1 naphthaleneacetic acid (NAA) or 0.6 mg·L-1 NAA + 0.7 mg·L-1 IBA. Additionally, thick and lateral roots were obtained with 0.6 mg·L-1 NAA + 0.7 mg·L-1 IBA. Our study is the first to establish an effective organogenesis protocol for new crabapple cultivars using stem segments.

Chitosan (CTS) induced secondary metabolite production in Canscora decussata Schult. - An endangered medicinal plant

Bioengineering has excellent potential in plant-based in vitro cultures for efficiently synthesizing desired plant products. Due to the rapid destruction of natural habitats, in vitro methods might save endangered species. Winged-Stem Canscora is an endangered medicinal herb used in traditional medicine for brain disorders. This study investigates the induction of in-vitro metabolite production using chitosan. Among the various concentrations of auxins tested, 82% of callus was obtained from Murashige and Skoog (MS) medium with 3 mg L−1 Kinetin (KIN) + 1 mg L−1 1-Naphthaleneacetic acid (NAA). The highest callus content was after the 3rd subculture at 200 mg L−1 chitosan treatment. On the 70th day of culture, levels of total phenolics and flavonoids are increased in the elicited and normal callus. However, antioxidant activity was higher in elicited callus compared to normal callus and wild plants. Higher secondary metabolite concentrations of elicited callus have superior anti-bacterial and anti-fungal activity. Secondary metabolites from elicited callus have a lower inhibitory concentration than those from wild plant and normal callus. High-performance thin layer chromatography (HPTLC) analysis showed that the elicited callus had higher amounts of mangiferin (0.178 μg mL−1) and scopoletin (0.133 μg mL−1). An efficient approach was employed to ensure that sufficient amounts of the widely used plant secondary metabolites from medicinal plants, such as mangiferin and scopoletin, were accessible to meet demand.

Facile preparation of covalent-organic framework composites for magnetic solid-phase extraction of naphthaleneacetic acid in food prior to HPLC-UV analysis

Novel magnetic covalent organic frameworks (COFs) were prepared by one-pot synthetic strategy and employed as an efficient adsorbent for magnetic solid-phase extraction (MSPE) of naphthaleneacetic acid (NAA) in food samples. Depending on the predesigned the hydrogen bonding, π-π and hydrophobic interactions of magnetic COFs, the efficient and selective extraction process for NAA was achieved within 15 min. The magnetic COFs adsorbent combined with HPLC-UV was devoted to develop a novel quantitative method for NAA in complex food. The method afforded good coefficient in range of 0.002–10.0 µg mL-1 and low limit of detection was 0.0006 µg mL-1. And the newly established method afforded less adsorbent consumption, wider linearity and lower LODs than the reported analytical methods. Ultimately, the method was successfully applied to determine NAA in fresh pear, tomato and peach juice. The magnetic COFs based MSPE coupled with HPLC-UV method provided a simple, efficient and dependable alternative to monitor trace NAA in food samples.

Tissue-Specific Natural Synthesis of Galanthaminein Zephyranthes Species and Its Accumulation in Different In Vitro-Grown Organs Following Methyl Jasmonate Treatment.

Galanthamine is an immensely valuable alkaloid exhibiting anti-cancer and antiviral activity. The cultivation of plant tissues in in vitro conditions is a good source for the synthesis and enrichment of secondary metabolites of commercial interest. In this study, the Amaryllidaceae alkaloid galanthamine was quantified in three Zephyranthes species, such as Zephyranthes candida, Zephyranthes grandiflora, and Zephyranthes citrina, and the impact of the methyl jasmonate (MJ) signaling molecule on galanthamine accumulation was monitored in in vitro-derived plant tissues. This is the first ever study of the MJ-regulated accumulation of galanthamine in in vitro-grown Zephyranthes tissues. Shoot regeneration was obtained in all three Zephyranthes species on Murashige and Skoog (MS) medium containing 2.0 mgL-1 benzylaminopurine (BAP) + 0.5 mgL-1 naphthalene acetic acid (NAA). The regenerated shoots were rooted on a medium containing 2.0 mgL-1 indole butyric acid (IBA). A GC-MS study of Zephyranthes extracts revealed the presence of 34 phyto-compounds of varied levels with therapeutic activities against diseases. The galanthamine content was quantified in plant parts of the three Zephyranthes species using high-performance thin layer chromatography (HPTLC); the maximum was found in Z. candida bulb (2.41 µg g-1 dry wt.), followed by Z. grandiflora (2.13 µg g-1 dry wt.), and then Z. citrina (2.02 µg g-1 dry wt.). The galanthamine content showed bulb > leaf > root source order. The in vitro-generated plantlets were treated with different MJ concentrations, and the galanthamine yield was measured in bulb, leaf, and root tissues. The highest galanthamine content was recorded in bulbs of Z. candida (3.97 µg g-1 dry wt.) treated with 150 µM MJ, showing an increase of 64.73% compared to the control. This accumulation may be attributed to MJ-induced stress, highlighting the potential commercial synthesis of galanthamine in vitro.

Assessment of genetic uniformity in micro propagated plantlets of turmeric (Curcuma longa L.) through DNA markers

Turmeric is herbaceous plant, characterized by long lance-shaped leaves sprouting from its rhizomatous underground stem. Notably, its rhizomes contain the bioactive compounds curcuminoids, renowned for its medicinal and culinary significance. However, meeting the surging demand for turmeric, particularly during off- seasons, presents a formidable challenge due to the sluggish vegetative propagation rate in Curcuma longa. Given the scarcity of sexual reproduction in turmeric, micropropagation emerges as the convenient method for obtaining disease-free seeds. In addressing the challenge of direct regeneration of the native Erode local cultivar of turmeric, this study endeavors to establish a protocol for in vitro plantlet production. Results indicate that multiple shoots were successfully induced, notably with 13.32 μM of 6-benzylaminopurine (BAP), yielding a response rate of 73.2±4.7% and an average shoot count of 6.95±0.81 per explant. Subsequently, a concentration of 17.76 μM of BAP demonstrated a response rate of 56.5%±4.7%, with an average shoot count of 5.94±0.81 per explant. Additionally, a combination of 13.32 μM BAP and 2.68 μM Naphthaleneacetic acid (NAA) resulted in 8.65±0.47 shoots per explant, with a response rate of 73.66±1.25%. Similarly, 13.32 μM BAP combined with 5.37 μM NAA yielded 7.32±0.47 shoots per explant, with a response rate of 72.33±0.47%. The acclimatization of plantlets in a greenhouse exhibited a remarkable survival rate, ranging from 90% to 98%. Importantly, all regenerated plantlets closely resembled the mother plants morphologically. Genetic uniformity assessment, employing 10 ISSR and 4 DAMD markers, indicated more than 90% uniformity among one mother plant and regenerants. This indicates a significant genetic uniformity, ensuring consistency in desired traits across the regenerated plantlets.

An Efficient In Vitro Shoot Organogenesis and Comparative GC-MS Metabolite Profiling of Gaillardia pulchella Foug

Gaillardia pulchella Foug. is a widely studied plant because of its high pharmacological and ornamental value. The leaves of G. pulchella were used for inducing callus and subsequent plant regeneration as it is the primary source of phytocompounds. The purpose of the present investigation was to formulate an in vitro propagation method for Gaillardia by using leaf explants in MS (Murashige and Skoog) medium. The best callus induction was observed on high (2.0 mg/L) α-naphthalene acetic acid (NAA) and a low (0.5 mg/L) 6-benzylaminopurine (BAP) with callus induction frequency of 91.66%. The leaf callus also demonstrated high caulogenesis ability (95.83%), with an average 5.2 shoots/callus mass at 0.5 mg/L BAP and 2.0 mg/L NAA. Indole Acetic acid (IAA) at 1.0 mg/L had the maximum rooting percentage (79.17%) with 12.4 roots per shoot. Rooted plantlets were later transferred to greenhouse conditions, showing a survivability rate of 75–80%. The physiological parameters, i.e., phenolic compounds and the flavonoids’ level, in the DPPH assay were higher in leaves obtained in vitro compared to callus formed from leaves and field-obtained (mother) leaves. Gas chromatography–mass spectrometry (GC–MS) analysis of methanol extracts of leaves (in vivo and in vitro) and leaf callus presented a wide array of compounds. In callus extract, some 34 phytocompounds were identified. Some of them were 3-hydroxy-2,3-dihydromaltol (25.39%), isoamyl acetate (11.63%), palmitic acid (11.55%), 4-methyloxazole (7.54%), and 5-methoxypyrrolidin-2-one (7.49%). Leaves derived in vivo and in vitro had 45 and 28 phytocompounds, respectively, belonging to different classes like lignans, phenols, terpenoids, alkaloids and fatty acids, etc. Those findings demonstrated that the leaf derived callus and the leaves are the potential stable source of several compounds with medicinal importance. The developed protocol may provide an alternative source of compounds without affecting wild flora.

Establishment of Physalis alkekengi cell suspension culture: time-dependent behavior of genes related to the steroidal compounds, key enzymes, and physalins under static magnetic field.

Cell suspension culture has the potential to be a valuable source for the bioactive compound productions. In this study, an optimized procedure was established for callus and cell suspension culture of Physalis alkekengi for the first time, and the impact of static magnetic field (SMF, 6 mT) was studied on the high-value metabolic compounds through investigation of signaling molecules and gene expressions at the late log-to-stationary phase. Results showed that the growth regulators of 6-benzyl amino purine (BAP, 1.5mg-1 L) and 1-naphthaleneacetic acid (NAA, 0.4mg-1 L) induced the highest fresh weight, callus rate, callus index, and total withanolides. Cell suspension culture was established in the liquid MS medium supplied with BAP (1.5mg-1 L) and NAA (0.1mg-1 L). SMF application decreased slightly the cell growth and viability and enhanced the number of round-shaped cells. The hydrogen peroxide (H2O2) and nitric oxide (NO) levels increased at an all-time series after SMF exposure, and their maximum contents were observed after 12h. A significant alteration of malondialdehydecontent was also identified after 12h of SMF exposure. The expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), 1-deoxyD-xylulose 5-phosphate synthase (DXS), squalene synthase (SQS), sterol Δ7-reductase (DWF5), and C-7,8 sterol isomerase (HYD1) genes was upregulated significantly after 24 and 48h. An increase in the total withanolides was related to more activity of HMGR and DXS enzymes in SMF-exposed cells and the maximum physalin A (12.8mgg-1 DW) and physalin B (1.92mgg-1 DW) obtained after 24h compared to controls. Findings suggest that SMF can play a supportive factor in inducing steroidal compounds in P. alkekengi through modulating H2O2 and NO levels and the related-gene expressions.

Biocontrol potential of endophytic fungi against phytopathogenic nematodes on potato (Solanum tuberosum L.)

Root-knot nematodes (RKNs) are a vital pest that causes significant yield losses and economic damage to potato plants. The use of chemical pesticides to control these nematodes has led to environmental concerns and the development of resistance in the nematode populations. Endophytic fungi offer an eco-friendly alternative to control these pests and produce secondary metabolites that have nematicidal activity against RKNs. The objective of this study is to assess the efficacy of Aspergillus flavus (ON146363), an entophyte fungus isolated from Trigonella foenum-graecum seeds, against Meloidogyne incognita in filtered culture broth using GC–MS analysis. Among them, various nematicidal secondary metabolites were produced: Gadoleic acid, Oleic acid di-ethanolamide, Oleic acid, and Palmitic acid. In addition, biochemical compounds such as Gallic acid, Catechin, Protocatechuic acid, Esculatin, Vanillic acid, Pyrocatechol, Coumarine, Cinnamic acid, 4, 3-indol butyl acetic acid and Naphthyl acetic acid by HPLC. The fungus was identified through morphological and molecular analysis, including ITS 1–4 regions of ribosomal DNA. In vitro experiments showed that culture filtrate of A. flavus had a variable effect on reducing the number of egg hatchings and larval mortality, with higher concentrations showing greater efficacy than Abamectin. The fungus inhibited the development and multiplication of M. incognita in potato plants, reducing the number of galls and eggs by 90% and 89%, respectively. A. flavus increased the activity of defense-related enzymes Chitinas, Catalyse, and Peroxidase after 15, 45, and 60 days. Leaching of the concentrated culture significantly reduced the second juveniles’ stage to 97% /250 g soil and decreased the penetration of nematodes into the roots. A. flavus cultural filtrates via soil spraying improved seedling growth and reduced nematode propagation, resulting in systemic resistance to nematode infection. Therefore, A. flavus can be an effective biological control agent for root-knot nematodes in potato plants. This approach provides a sustainable solution for farmers and minimizes the environmental impact.

Regeneration of a Multipurpose Woody Legume - Acacia auriculiformis through Mature Tree Nodal Explants

Multiple shoots have been induced in nodal explants of 30-year-old Acacia auriculiformis trees on Nitsch's medium (N medium) supplemented with 1.0 mg/l N6-benzyladenine. Addition of 0.1 mg/l adenine sulfate to N medium containing BA, enhanced the caulogenic response whereby the number of shoots developing on each node increased from three to fifteen. Plantlets were obtained by rooting axenically raised shoots on N medium (half strength) with 1.0 mg/l naphthalene acetic acid. Use of activated charcoal (0.01%) in the medium completely suppressed callus formation at the root-shoot junction. Plant Tissue Cult. & Biotech. 34(1): 1-10, 2024 (June)

Effect of Plant Growth Regulators on Mass Propagation of Strawberry (Fragaria sp.)

Efficient in vitro regeneration protocols for strawberries are crucial for accelerating the propagation of superior cultivars and improving overall productivity through controlled manipulation of plant growth regulators. This study investigated the influence of different concentrations and combinations of plant growth regulators on in vitro regeneration process of strawberries. BARI Strawberry-1 runner tips were sterilized and placed on Murashige and Skoog medium (MS) supplemented with various combinations of cytokinins, including 6-benzylaminopurine (BAP) and kinetin (KIN), to initiate shoot growth. Notably, the combination of 1.5 mg/L BAP + 1.0 mg/L KIN led to the shortest duration for shoot initiation (9 days), resulting in the highest number of shoots per explants (6.33), longest shoot length (2.87 cm), and maximum number of leaves per shoot (4.00). Additionally, this combination also exhibited the highest rate of shoot regeneration (70.33%). Subsequently, the proliferated shoots were transferred to MS medium supplemented with various concentrations and combinations of auxins, specifically 3-indolebutyric acid (IBA) and 1-naphthaleneacetic acid (NAA), for root induction. The in vitro regenerated shoots cultured in MS medium supplemented with 1.5 mg/L IBA + 1.0 mg/L NAA showed the shortest duration for root formation (8 days), the highest number of roots per explants (4.00), and the longest root length (2.77 cm). Furthermore, this combination also resulted in the highest rate of root regeneration (40%). These findings contribute valuable insights for the mass production of healthy strawberry plantlets using in vitro culture techniques. J Bangladesh Agril Univ 22(2):178-184, 2024

Propagasi In Vitro Kaliandra Merah (Calliandra calothyrsus Meisn.) II: Induksi Perakaran Tunas dan Aklimatisasi Planlet

Red calliandra is one of woody plants that has been extensively developed as biofuel. Tissue culture techniques are employed to produce a large and uniform quantity of seedlings in a relatively short period. Previous stages of in vitro initiation and multiplication of shoot from calliandra explants have shown promising results. The shoots that were multiplied by in vitro technique need to be induced for rooting process to produce plantlets. These plantlets are then prepared for the subsequent acclimatization process. This research was conducted in two stages; root induction of shoots using a modification of MS half-strength media (MS ½) supplemented with 1-4 mg/L 1-naphthaleneacetic acid (NAA), and acclimatization of the plantlets into soil media. The results shown that MS ½ medium and media supplemented with 1 mg/L NAA were suitable for root induction, particularly to induce the root length. Furthermore, MS ½ media supplemented with 1-2 mg/L NAA stimulates the increment of plantlet height and leaves numbers. Plantlets which were transferred into soil exhibit robust growth during 3-weeks of acclimatization process, up to 100% plantlets survived during this stage. These results underscore the potential of tissue culture techniques in providing large scale calliandra seedlings production for future biofuel development.

Optimization of in vitro Regeneration of Cowpea (Vigna unguiculata L.) using Computational Models

Cowpea (Vigna unguiculata L.) is the most preferable cultivated crop in the World and can grow on poor soil with water scarcity. More than 80.0% of its cultivation is confined to the Sub-Saharan African continent. However, it is now one of the most preferable crops in Asia and Europe due to its low nutritional and water demand during cultivation under field conditions. In this study, the Bodrum Red cultivar of cowpea was used for in vitro regeneration. The 9-0 days old half cotyledonary node explant was used and placed on different concentrations of Benzylaminopurine (BAP), Indole-butyric acid (IBA), and Naphthalene acetic acid (NAA). The results were analyzed by ANOVA and response surface methodology (RSM). The ANOVA analysis revealed the maximum shoots from the medium enriched with 1.0 mg/L BAP with shot counts ranging from 1.17-3.11. The results were further analyzed by surface regression analysis which revealed the significant impact of NAA on callus formation. The results were also analyzed through Pareto charts and normal plots which confirmed the significant role of BAP and NAA on in vitro regeneration. The use of a response optimizer revealed the maximum shoot counts of 3.0 per explant from the medium supplemented with BAP. Results revealed the significance of RSM for optimizing input factors precisely for multiple input factors simultaneously. Advanced biotechnological approaches like genetic engineering and marker-assisted selection could enhance cowpea resilience to environmental conditions, ensuring its sustainability and adaptability across diverse agroecological zones with the aid of RSM

Efficient electrochemical synthesis of phenylacetic acid derivatives: Utilizing CO2 for sustainable production

Phenylacetic acid derivatives, including 1-naphthaleneacetic acid, hold crucial applications in various domains. Traditional synthesis methods often involve intricate steps or the use of expensive and polluting heavy metal catalysts. This paper presents an electrochemical approach for efficiently producing valuable compounds like 1-naphthaleneacetic acid and other phenylacetic acid analogs through the reduction of benzyl halides, such as 1-chloromethyl naphthalene, in the presence of CO2. At the same time as preparing phenylacetic acid, the resource utilization of CO2 is realized. Six kinds of carboxy analogs were synthesized by potentiostatic electrolysis of benzyl halides in acetonitrile with CO2 as a co-reactant and tetraethyl ammonium tetrafluoroborate as an electrolyte. The electrochemical reduction products were confirmed structurally through NMR, mass spectrometry, and other techniques. The Faraday electrolysis efficiencies ranged from 82 % to 94 %, with 72.5–92.9 % conversions. The study delves into the electron transfer mechanism using cyclic voltammetry and in situ FT-IR spectroelectrochemistry. The reduction process on silver electrodes under a CO2 atmosphere was identified as an irreversible two-electron process, generating a monovalent anion that attacks CO2 to yield the respective carboxyl analogs.

Androgenesis in Hot Pepper by Chilli Anther Culture

In vitro androgenesis techniques play a key role in the quick generation of fully homozygous lines. This work's goal was to put the procedure for producing androgenic embryos in hot pepper populations into practice. A number of variables can affect androgenetic success, including the types and quantities of carbohydrate sources and the nature of the nutritional medium. Different sugar types were tested at different doses in anther culture of pepper to elucidate their influence on the yield of androgenesis. In this study, sucrose was used as a source of sugar, Murashige & Skoog (1962-MS) medium, silver nitrate (AgNO3), naphthaleneacetic acid (NAA), BAP and one breeding lines of pepper were used to know the genotypic effects on haploid embryo formation. It was found that one of the key elements influencing pepper anther and microspore culture success is genotype. Also, microspore stage and the cold treatment of flower bud responsible for the success of embryo formation. This embryo formation frequency has been calculated according to the results that were generated in one month and the frequency of embryo formation is 0.7%.

Manganese toxicity elicits the degradation of auxin transport carriers to restrain arabidopsis root growth

Manganese (Mn) is an essential microelement for plant growth but can be toxic when in excessive. Mn toxicity-inhibited root growth possible involves in auxin biosynthesis and transport in plants, but the mechanism remains elusive. Our results have suggested that Mn-inhibited root growth was possible associated with a reduced auxin levels in roots, as shown in experiments with (1) transgenic DII-VENUS and DR5rev::GFP; (2) the application of L-kynurenine (an inhibitor of TAA1 activity), 1-naphthylacetic acid (NAA), 1-naphthylphthalamic acid (NPA) to manipulate auxin levels; (3) using taa1 (defection in auxin biosynthesis) mutants; and (4) determining IAA by enzyme-linked immunosorbent assay. Those results were explained via down-regulated transcriptional levels of auxin biosynthesis-related genes in plants, and reduced abundance of PM-localized auxin transport carriers in roots. Combination of auxin transport-related transgenic pPINs-GFP or pAUX1::AUX1-YFP lines and a real-time in vitro observation of PM-localized PIN2/3/4 carriers using transgenic Dendra2 lines have revealed that excessive Mn induced degradation of these proteins, which might involve in Mn toxicity-elicited up-regulation of partially ubiquitin-, clathrin-meidiated endocytosis (CME)-, and endosomal sorting complex required for transport (ESCRT)-related genes. Overall, our results suggest that Mn toxicity reduces auxin level in the root apex possible via down-regulation of auxin biosynthesis-related genes and post-translational stimulation of the degradation of auxin transport proteins, leading to root growth inhibition caused by Mn toxicity.

In vitro morphogenesis and micro-morpho-anatomical developments in Moringa concanensis Nimmo.: An endemic tree of Indian sub-continent

Moringa concanensis Nimmo. is a regionally important and underutilized medicinal tree of the Indian subcontinent. An in vitro morphogenesis protocol for Moringa concanensis has been developed using seedling-derived explants. The cotyledonary node explants were found to be the most suitable exhibiting cent-percent bud break with each regenerating 2.33±0.76 shoots of 1.32±0.69 cm length within 15 days on 1.0 mg/L 6-benzyl aminopurine (BAP) supplemented modified Murashige and Skoog Medium (MMS) media. The rate of shoot multiplication increased up to 10.17±2.54 shoots/inoculum by sub-culturing of regenerated shoots on 0.5 mg/L BAP along with 0.1 mg/L Indole acetic acid (IAA). The in vitro cloned shoots were rooted using both in vitro and ex-vitro methods by applications of root-inducing auxins, indole butyric acid (IBA), and naphthalene acetic acid (NAA). IBA proved to be more effective and induced roots in all in vitro raised shoots. It produced 17.72±4.83 roots/shoot with 2.12±0.88 cm length on 1/4th strength MMS media fortified with 1.0 mg/L of IBA. Ex vitro rooted plantlets with concurrent acclimatization were achieved by pre-treating shoot bases with IBA at 250 ppm for 5 min. Comparative micro-morpho-anatomical evaluation of leaf and stem revealed differentiation of non-glandular trichomes, anomocytic stomata, and vascular tissue development under applied in vitro conditions, which suggests the survivability of plantlets. Sixty percent of the in vitro regenerated plants survived in the field. It is the foremost report on in vitro morphogenesis of Moringa concanensis protocol using cotyledonary nodal explant and ex vitro rooting method. The tissue culture methods defined and developed can be used for large-scale multiplications of Moringa concanensis as non-conventional approaches.

Establishment of a callus induction system of Saxifraga stolonifera Meerb. and its response to different elicitors

Elicitation is a well-known strategy for increasing the accumulation of secondary metabolites in callus cultures. In this study, a callus induction system for Saxifraga stolonifera Meerb. was developed, and elicitors were used to enhance secondary metabolite production. Tender leaves of S. stolonifera were used as explants for in vitro culture. We examined callus induction characteristics of S. stolonifera and its response to various elicitors by assessing the sterilization conditions of the explants, varying hormone ratios, and determining the optimal concentrations of salicylic acid (SA), Sodium nitroprusside (SNP) and methyl jasmonate (MeJA). The results revealed that sterilizing the explants with 70% ethanol for 40 s, followed by 0.1% mercury chloride for 10 min, resulted in a survival rate of 82.22% during tissue culture. Following these sterilization methods, callus formation was successfully induced on Murashige and Skoog medium supplemented with 2 mg/L 6-benzylaminopurine and 0.15 mg/L naphthaleneacetic acid, achieving an induction rate of 88.88%. Additionally, the study identified SA as the most effective elicitor for increasing the levels of gallic acid, protocatechuic acid, and quercitrin in the callus, with optimal concentrations of salicylic acid being 60, 40, and 60 μM, respectively. SNP at 40 μM, 80 μM, and 20 μM concentrations was identified as the best elicitor for promoting the accumulation of bergenin, chlorogenic acid, and quercetin, respectively. This study established appropriate conditions for cultivating S. stolonifera callus and identified the optimal elicitor concentrations for maximizing secondary metabolite production.

Effect of Plant Growth Regulators and Growing Media on Rooting of Cuttings in Punica granatum L. cv. Bhagwa

Background: When pomegranates are multiplied through seeds, germination leads to vigorous tree growth. However, the tree will not be exactly similar to its parent and the resultant fruit produced will be irregular in colour, size, juiciness and sweetness. Methods: Soft-wood cuttings of Pomegranate cv. Bhagwa were planted in different growing media that were entitled G1- Soil + FYM, G2- Soil + Vermiculite, G3- Soil + Sand + FYM and G4- Soil + FYM + Vermiculite. These cuttings were treated with different combinations of 1-Naphthaleneacetic acid (NAA), Indole-3-butyric acid (IBA), Hydrogen peroxide (H2O2), Ascorbic acid (ASC) and Boric acid. Studies were carried out in a two-factorial randomized block design (FRBD) with 28 treatment permutations. Result: Current research indicated that treatment T18 (NAA 500 ppm + H2O2 30 ppm + IBA 500 ppm, Soil + Vermiculite (1:1) worked out to be the best treatment as it exhibited superiority over other treatments. The results of this experiment can practically be implemented to propagate quality planting material by adopting these best results/treatments.

In Vitro Micropropagation of the Vulnerable Chilean Endemic Alstroemeria pelegrina L.

The Chilean rhizomatous geophyte Alstroemeria pelegrina (A. pelegrina), a species endemic to the Coquimbo (31°45′ S) and Valparaíso (33°12′ S) regions, is currently classified as vulnerable and experiencing population decline due to loss of habitat. This heightened threat underscores the need to develop effective mass propagation techniques to support its conservation efforts. Consequently, the present study aimed to establish an optimized in vitro propagation protocol tailored specifically for A. pelegrina to facilitate its large-scale propagation and promote the preservation of this vulnerable geophyte. In our experiment, explants obtained from in vitro germination were used. These explants were then cultured on a Murashige and Skoog (MS) medium solidified with agar (6 g L−1) and supplemented with 2.22 μM 6-Benzylaminopurine (BAP) and 30 g L−1 sucrose. The results indicated an average of 4.6 new shoots produced per initial explant. The implementation of light-emitting diode (LED) illumination with a red:blue ratio of 3:1 and 2.22 μM BAP resulted in an average shoot length of 6.2 cm. For rhizome induction, the addition of either 5.37 μM 1-naphthaleneacetic acid (NAA) or 5.37 μM indole-3-butyric acid (IBA) resulted in average rhizome lengths of 1.9 cm and 1.7 cm, respectively, with fresh weights of the vitroplants ranging from 2.9 g to 3.1 g, and rooting percentages between 100% and 96%, respectively. Notably, the survival rate of rooted plants obtained through in vitro propagation was 90% after a 4-week acclimatization period in a cold greenhouse, indicating the efficacy of the developed protocol for mass propagation and conservation efforts of this vulnerable geophyte species.

Phytochemical characterization of callus cultures from the endangered plant Crocus scepusiensis (Rehm. & Woł.) Borbás ex Kulcz.

Crocus scepusiensis (Rehm. & Woł.) Borbás ex Kulcz., a critically endangered herbaceous plant which serves as a valuable source of bioactive compounds found across Europe and Asia. The aim of this study was to produce a calli from two different plant parts (leaf and shoot tip) for the critically endangered C. scepusiensis through tissue culture techniques, characterize the resulting calli through chemical profiling, with a focus on identifying key phytoconstituents, and lay the groundwork for future research on the biological activities of these calli extracts. Leaf disc and micro shoot tip explants were cultured on Murashige and Skoog (MS) medium supplemented with various concentrations of 6-benzylaminopurine (BA) and α-naphthaleneacetic acid (NAA) to induce organogenic calli. The resulting calli exhibited distinct biochemical profiles. Moreover, a phytochemical analysis was conducted to compare the metabolite composition of callus 1 (derived from leaf discs) and callus 2 (derived from shoot tips). Callus 1 displayed a higher total phenolic content (30.3558 ± 1.3564 mg (GAE)/g) compared to callus 2 (29.1543 ± 0.9754 mg (GAE)/g). Similarly, callus 1 exhibited a greater total flavonoid content (26.0089 ± 1.8029 mg (RE)/g) than callus 2 (18.4464 ± 1.4797 mg (RE)/g). Liquid chromatography-photodiode array-electrospray ionization-tandem mass spectrometry (LC-PDA-ESI-MS/MS) analysis revealed the presence of 26 and 25 constituents in callus 1 and 2, respectively. Fourteen and thirteen of these identified compounds have been previously reported in other Crocus species, with 22 constituents common to both calli. Twelve constituents were reported here in Crocus for the first time as far as we know.