Isolation of the Main Biologically Active Substances and Phytochemical Analysis of Ginkgo biloba Callus Culture Extracts.

The work reveals the results of studying the content of biologically active substances in samples of extracts of Ginkgo biloba callus cultures. Callus cultures grown in vitro on liquid nutrient media were the objects of the study. Considering various factors affecting the yield of the target components during extraction, the volume fraction of the organic modifier in the extracting mixture, the temperature factor, and the exposure time were identified as the main ones. The maximum yield of extractive substances (target biologically active substances with a degree of extraction of at least 50%) from the samples of callus culture extracts was detected at a ratio of extragent of 70% ethanol, a temperature of 50 °C, and exposure time of 6 h. Flavonoids, such as luteolin, quercetin, isoramentin, kaempferol, and amentoflavone, were isolated in the extract samples. As a result of column chromatography, fractions of individual biologically active substances (bilobalide, ginkgolide A, B, and C) were determined. The proposed schemes are focused on preserving the nativity while ensuring maximum purification from associated (ballast) components. Sorbents (Sephadex LH-20, poly-amide, silica gel) were used in successive stages of chromatography with rechromatography. The degree of purity of individually isolated substances was at least 95%.

Highly Efficient Bioconversion of trans-Resveratrol to δ-Viniferin Using Conditioned Medium of Grapevine Callus Suspension Cultures.

δ-Viniferin is a resveratrol dimer that possesses potent antioxidant properties and has attracted attention as an ingredient for cosmetic and nutraceutical products. Enzymatic bioconversion and plant callus and cell suspension cultures can be used to produce stilbenes such as resveratrol and viniferin. Here, δ-viniferin was produced by bioconversion from trans-resveratrol using conditioned medium (CM) of grapevine (Vitis labruscana) callus suspension cultures. The CM converted trans-resveratrol to δ-viniferin immediately after addition of hydrogen peroxide (H2O2). Peroxidase activity and bioconversion efficiency in CM increased with increasing culture time. Optimized δ-viniferin production conditions were determined regarding H2O2 concentration, incubation time, temperature, and pH. Maximum bioconversion efficiency reached 64% under the optimized conditions (pH 6.0, 60 °C, 30 min incubation time, 6.8 mM H2O2). In addition, in vitro bioconversion of trans-resveratrol was investigated using CM of different callus suspension cultures, showing that addition of trans-resveratrol and H2O2 to the CM led to production of δ-viniferin via extracellular peroxidase-mediated oxidative coupling of two molecules of trans-resveratrol. We thus propose a simple and low-cost method of δ-viniferin production from trans-resveratrol using CM of plant callus suspension cultures, which may constitute an alternative approach for in vitro bioconversion of valuable molecules.

The Callus of Phaseolus coccineus and Glycine max Biotransform Flavanones into the Corresponding Flavones.

In vitro plant cultures are gaining in industrial importance, especially as biocatalysts and as sources of secondary metabolites used in pharmacy. The idea that guided us in our research was to evaluate the biocatalytic potential of newly obtained callus tissue towards flavonoid compounds. In this publication, we describe new ways of using callus cultures in the biotransformations. In the first method, the callus cultures grown on a solid medium are transferred to the water, the reaction medium into which the substrate is introduced. In the second method, biotransformation is carried out on a solid medium by growing callus cultures. In the course of the research, we have shown that the callus obtained from Phaseolus coccineus and Glycine max is capable of converting flavanone, 5-methoxyflavanone and 6-methoxyflavanone into the corresponding flavones.

Comparative analysis of the effects of chemically and biologically synthesized silver nanoparticles on biomass accumulation and secondary metabolism in callus cultures of Fagonia indica.

Biotechnological strategies are needed to produce larger quantities of biomass and phytochemicals. In this study, callus cultures of Fagonia indica were elicited with different concentrations of chemically and biologically synthesized silver nanoparticles (chem- and bioAgNPs) to compare their effects on biomass, total phenolic content (TPC), total flavonoid content (TFC) and antioxidant activity of the extracts from callus. The results revealed that bioAgNPs being more biocompatible produced the highest biomass initially on day 10 (FW = 4.2152 ± 0.13 g; DW = 0.18527 ± 0.01 g) and day 20 (FW = 7.6558 ± 0.10 g; DW = 0.3489 ± 0.01 g) when supplemented in media as 62.5 µg/mL and 250 µg/mL, respectively. Initially, the highest TPC (319.32 ± 8.28 µg GAE/g of DW) was recorded on day 20 in chemAgNPs (31.25 µg/mL) induced callus as compared to TPC = 302.85 ± 3.002 µg GAE/g of DW in bioAgNPs-induced callus. Compared to the highest values of TFC (108.15 ± 2.10 µg QE/g of DW) produced in 15.6 µg/mL chemAgNPs-induced callus on day 20, TFC produced in bioAgNPs (62.5 µg/mL) was 168.61 ± 3.17 µg GAE/g of DW on day 10. Similarly, chemAgNPs-induced callus (62.5 µg/mL) showed the highest free radical scavenging activity (FRSA) i.e. 87.18% on day 20 while bioAgNPs (125 µg/mL) showed 81.69% FRSA on day 20 compared to highest among control callus (63.98% on day 40). The highest total antioxidant capacity of chemAgNPs-(125 µg/mL) induced callus was 330.42 ± 13.65 µg AAE/g of DW on day 20 compared to bioAgNPs-(62.5 µg/mL) induced callus (312.96 ± 1.73 µg AAE/g of DW) on day 10. Conclusively, bioAgNPs are potent elicitors of callus cultures of F. indica.

[Flavonoid glycosides from callus cultures of Dysosma versipellis].

Various chromatographic techniques, including silica gel column chromatography, Sephadex LH-20, preparative thin-layer chromatography, and preparative HPLC, were employed to isolate the chemical constituents from callus cultures of Dysosma versipellis. Structures of the compounds were elucidated based on UV, IR, MS and NMR spectroscopic data analysis. Totally, seven flavonoid glycosides were isolated from the 95% ethanol extract of the callus cultures and identified as kaempferol-3-O-[6″-(3″'-methoxy)-malonyl]-ß-D-glucopyranoside(1), kaempferol-3-O-(6″-O-acetyl)-ß-D-glucopyranoside(2), kaempferide-3-O-ß-D-glucopyranoside(3), kaempferol-3-O-ß-D-glucopyranoside(4), isoquercitrin(5), quercetin-4'-O-ß-D-glucopyranoside(6) and kaempferol-3-(6″-malonyl)-ß-D-glucopyranoside(7), respectively.All these compounds were isolated from callus cultures of D. versipellis for the first time.Compounds 1, 2, 3, 6 and 7 were firstly obtained from plant materials of D. versipellis, and compound 1 was a new compound.

Establishment of callus cultures and quantification of Caffeic acid using HPTLC from Desmodium gangeticum (L.).

Desmodium gangeticum (L.) belonging to family Fabaceae is an economically important medicinal plant which isutilised in Dashmoolarishta. Various bioactive compounds have been isolated from whole plant and roots, and one of them is an important phenolic compound - caffeic acid (CA). This phenolic acid and its derivatives have antioxidant, anti-inflammatory, anticarcinogenic and hepatocarcinoma, a highly aggressive and causing considerable mortality across the world. In the present study, leaf explants were placed on MS medium fortified with different concentration of cytokinin (BA/Kn) and auxin (IAA/NAA) for establishing callus cultures. MS medium fortified with BA (20 µM) and IAA (2 µM) was optimised for the same. Methanolic extracts of in vivo leaf sample (DG1) and in vitro sample (leaf derived callus) (DG2) were assessed for CA quantification using HPTLC. Thus, the chemical fingerprint that was obtained, confirmed that DG 2 of D. gangeticum exhibited the potency to synthesise more amount of CA (316 ± 7.5 µg/g DW) in comparison to DG1 which was 194 ± 2.3 µg/g DW.

Profiling of Polyphenolic Compounds of Leontopodium alpinum Cass Callus Cultures Using UPLC/IM-HRMS and Screening of In Vitro Effects.

Leontopodium alpinum Cass. (edelweiss) is recognized as a frequent constituent of anti-aging skin care products, providing increased antioxidant and anti-inflammatory defense. Considering the growing demand and the protected status of edelweiss in many countries, alternative methods of production have been developed, one of them being callus culturing. This study reports the phytochemical composition of a methanolic extract of L. alpinum callus cultures, characterized by liquid chromatography coupled to ion-mobility high resolution mass spectrometry (UPLC/IM-HRMS). The methanolic extract exhibited strong free radical scavenging activity (122.19 ± 7.28 mg AAE/g dw), while the quantitative evaluation revealed that four major constituents (phenylpropanoid derivatives) represent 57.13% (m/m) of the extract. Consequently, a screening of antiproliferative effects was performed on ten cancer cell lines, representative of prostate, colon, lung and breast cancer, showing inhibition of colony formation in all cases. These results provide a comprehensive phytochemical characterization of L. alpinum callus cultures using advanced IM-HRMS, while the in vitro explorations confirmed the potent antioxidant properties of edelweiss which are worth exploring further in cancer prevention.

In vitro metabolism of tebuconazole, flurtamone, fenhexamid, metalaxyl-M and spirodiclofen in Cannabis sativa L. (hemp) callus cultures.

BACKGROUND: Cannabis sativa L. (hemp) is a medicinal plant producing various cannabinoids. Its consumption is legalized for medical use due to the alleged positive health effects of these cannabinoids. To satisfy the demand, C. sativa plants are propagated in contained growth chambers. During indoor propagation, pesticides usually are used to ensure efficient production. However, pesticide registration and safe application in C. sativa has not been investigated in detail. RESULTS: With this study the metabolic degradation of pesticides in recently established C. sativa callus cultures was examined. Tebuconazole, metalaxyl-M fenhexamid, flurtamone and spirodiclofen were applied at 10 µm for 21 days. Results were compared with metabolism data obtained from Brassica napus L., Glycine max (L.) Merr., Zea mays L. and Tritium aestivum L. callus cultures as well as in metabolism guideline studies. The successfully established C. sativa callus cultures were able to degrade pesticides by oxidation, demethylation, and cleavage of ester bonds in phase I, as well as glycosylation and conjugation with malonic acid in phase II and III. Initial metabolites were detected after Day (D)7 and were traced at D21. CONCLUSION: The resulting pathways demonstrate the same main degradation strategies as crop plants. Because metabolites could be the main residue, the exposure of consumers to these residues will be of high importance. We present here an in vitro assay for a first estimation of pesticide metabolism in C. sativa. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Evaluation of the Conditions for the Cultivation of Callus Cultures of Hyssopus officinalis Regarding the Yield of Polyphenolic Compounds.

The cultivation of plants in the form of callus cultures constitutes a renewable source of secondary plant metabolites. The conditions for the cultivation of callus cultures affect the yield of target compounds. Callus cultures of Hyssopus officinalis were chosen for study. Nutrient media of various compositions were used for Hyssopus officinalis callus culture. For each culture, data on the quantitative contents of saponins, flavonoids and polyphenolic compounds, as well as antioxidant activity, were obtained. It was found that Murashige and Skoog medium supplemented with 1-naphthylacetic acid and kinetin led to the highest yield of secondary metabolites.

Evaluation of Callus Cultures to Elucidate the Metabolism of Tebuconazole, Flurtamone, Fenhexamid, and Metalaxyl-M in Brassica napus L., Glycine max (L.) Merr., Zea mays L., and Triticum aestivum L.

Plant cell cultures can be used to identify the metabolic degradation of pesticides in crops. Therefore, Brassica napus L., Glycine max (L.) Merr., Zea mays L. and Triticum aestivum L. were used to elucidate the metabolic degradation of the following pesticides: tebuconazole, flurtamone, fenhexamid, and metalaxyl-M. Callus cultures were treated with 10 µM of the named pesticides by passive diffusion out of the nutrition agar while young plants were hydroponically exposed to it. After 14 days, the comparison of in planta and in vitro experiments showed that the metabolic degradation is well described by in vitro callus cultures. The intracellular uptake of all pesticides and a broad spectrum of exemplarily hydroxylated and conjugated metabolites were detectable. Overall, the comparability of the nature of residues out of both experiments with the regulatory guideline metabolism studies could be demonstrated. Therefore, we recommend it as a potential screening tool to elucidate the metabolism of pesticides in crops.

Large-scale production of recombinant miraculin protein in transgenic carrot callus suspension cultures using air-lift bioreactors.

Miraculin, derived from the miracle fruit (Synsepalum dulcificum), is a taste-regulating protein that interacts with human sweet-taste receptors and transforms sourness into sweet taste. Since miracle fruit is cultivated in West Africa, mass production of miraculin is limited by regional and seasonal constraints. Here, we investigated mass production of recombinant miraculin in carrot (Daucus carota L.) callus cultures using an air-lift bioreactor. To increase miraculin expression, the oxidative stress-inducible SWPA2 promoter was used to drive the expression of miraculin gene under various stress treatments. An 8 h treatment of hydrogen peroxide (H2O2) and salt (NaCl) increased the expression of miraculin gene by fivefold compared with the untreated control. On the other hand, abscisic acid, salicylic acid, and methyl jasmonate treatments showed no significant impact on miraculin gene expression compared with the control. This shows that since H2O2 and NaCl treatments induce oxidative stress, they activate the SWPA2 promoter and consequently up-regulate miraculin gene expression. Thus, the results of this study provide a foundation for industrial-scale production of recombinant miraculin protein using transgenic carrot cells as a heterologous host.

Influence of photoperiod on growth, bioactive compounds and antioxidant activity in callus cultures of Basella rubra L.

Basella rubra L. is an important green leafy vegetable vine and is known for its health benefits in traditional medicine. Light is a basic physical factor essential to the development and bioactive secondary metabolite production in in vitro callus cultures. The present study researched the impact of different photoperiods on biomass, bioactive compounds, and antioxidant activity in callus cultures of B. rubra. The in vitro seedling based cotyledonary leaf explants responded differently, when cultured on Murashige and Skoog (MS) medium with varying concentrations and combination of auxins and cytokinins. The best callus proliferation was found in MS medium with 0.1 mg.L-1 1-naphthaleneacetic acid (NAA) and 6 mg.L-1 6-benzylaminopurine (BAP), with greenish callus inception by about 2 weeks. The growth curve recorded for 6 weeks of culturing revealed that the photoperiod effect was found to be pivotal for acquiring biomass. At the fifth week, the continuous light supported maximum biomass (12.42 g) production followed by the 16:8 h photoperiod (9.02 g) and continuous darkness (4.28 g). The 80% ethanol extract of 1-week-old callus that grows under the 16:8 h photoperiod showed the highest total phenolic content (TPC) (74 mg.100 g-1 fresh weight, FW) when compared to all other extracts at different stages. The ferric reducing antioxidant power assay showed the highest (336.23 mg.100 g-1 FW) activity in methanol extractions of first-week callus cultures maintained in the continuous light condition. HPLC-UV identification and quantification of individual phenolics and flavonoids, such as gallic, trans-cinnamic, quercetin, protocatechuic and rutin, were highest in the callus cultures. The outcome of this study is significant to this plant, as B. rubra is familiar for its important health constituents with high-value bioactives and applications in the pharma and nutraceutical industries.

The influence of light quality on the production of bioactive metabolites - verbascoside, isoverbascoside and phenolic acids and the content of photosynthetic pigments in biomass of Verbena officinalis L. cultured in vitro.

In vitro callus cultures of Verbena officinalis L. were maintained on solid Murashige and Skoog medium, enriched with 1 mg dm-3 BA and 1 mg dm-3 IBA under LED lights (red, blue, red/blue 70%/30%), in darkness and under control fluorescent lamps. The measurements of 2 phenylpropanoid glycosides (verbascoside and isoverbascoside) and 23 phenolic acids were performed in methanolic extracts from the biomass collected after 2-, 3- and 4-week growth cycles using the HPLC-DAD method. The presence of verbascoside, isoverbascoside and additionaly 7 phenolic acids (protocatechuic, chlorogenic, vanillic, caffeic, ferulic, o-coumaric and m-coumaric acids) was confirmed in all extracts. Blue and red/blue lights stimulated the accumulation of verbascoside (max. of 6716 and 6023 mg 100 g-1 DW after a 4-week growth cycle) and isoverbascoside (max. 333 and 379 mg 100 g-1 DW also after 4 weeks). The maximum amounts of verbascoside and isoverbascoside were respectively 1.8- and 7.0-fold higher than under the control conditions. Phenolic acids were accumulated in different amounts, and the maximum total amounts ranged from 36 to 65 mg 100 g-1 DW. LED lights also stimulated their accumulation in comparison with darkness and control. The main phenolic acids included: m-coumaric acid (max. 39 mg 100 g-1 DW), ferulic acid (max. 12 mg 100 g-1 DW), and protocatechuic acid (max. 13 mg 100 g-1 DW). Additionally, the quantities of photosynthetic pigments (chlorophyll a, b and carotenoids) were estimated in acetonic extracts using spectrophotometry. Red/blue light stimulated the biosynthesis of pigments (max. total content 287 µg g-1 FW after 4-week growth cycles). This is the first study describing the effect of LED lights on the production of phenylpropanoid glycosides and phenolic acids in V. officinalis callus cultures. Very high amounts of verbascoside and isoverbascoside are interesting from a practical point of view.

Strategy for early callus induction and identification of anti-snake venom triterpenoids from plant extracts and suspension culture of Euphorbia hirta L.

Euphorbia hirta L. from the family of Euphorbiaceae is an annual herb, which grows as a roadside weed in most tropical countries. It is prominently used by the traditional healers in rural India for the treatment of snakebites. However, the mechanisms and the major bioactive compounds behind its inhibition activity are relatively unknown. From our preliminary in silico studies, it was found that a group of pentacyclic triterpenoids from this plant are playing a major role in inhibiting the snake venom proteins. The present study was aimed at standardizing methods for obtaining callus from this medicinal plant at a much faster rate by hormone pretreatment of explants and, thus, by developing suspension cultures to obtain bioactive secondary metabolites in vitro. The results were promising that longer incubation of explants with hormone treatment showed early induction of callus. The major bioactive compounds responsible for the anti-snake venom activity were characterized from natural plant material as well as from suspension cultures, and the efficiency was found to be relatively high. The secondary metabolite analysis from suspension culture and natural plant extracts revealed that a major compound 'Taraxerol' and its derivatives was found abundant along with few other triterpenoids. This compound showed high inhibitory activity against pit viper snake venoms from our in silico studies with molecular docking tools. Hence, this study with identification of potential bioactive compounds against snake venom with standardization of In vitro culture methods would help in developing natural alternative medicine for snakebites in near future.

New triterpenic acids produced in callus culture from fruit pulp of Acca sellowiana (O. Berg) Burret.

The aim of this work was the study of the best conditions for obtaining a callus culture from the pulp of Acca sellowiana, and to perform a quali-quantitative analysis of the secondary metabolites yielded by the in vitro callus culture. To this end, callus was induced on both Murashige and Skoog and Gamborg B5 media containing various combinations of growth regulators. Three previously undescribed ursane-type triterpenoids, 2α,3ß,6α,23-tetrahydroxy-18α,19α-urs-20-en-28-oic acid, 2α,3ß,23-trihydroxy-18α,19α-urs-20-en-28-oic acid and 2α,3ß,6ß,23-tetrahydroxy-18α,19α-urs-20-en-28-oic acid were isolated from the methanolic extract of A. sellowiana culture and characterized by 1D and 2D NMR experiments. Moreover, the quali-quantitative analysis (ESI-MSn and GC-MS) also showed the presence of ß-sitosterol, phloridzin, oleanolic, ursolic, 3ß-hydroxy-18α,19α-urs-20-en-28-oic, maslinic, corosolic, 2α,3ß-dihydroxy-18α,19α-urs-20-en-28-oic, and tormentic acid.

Effect of in vitro morphogenesis on the production of podophyllotoxin derivatives in callus cultures of Linum album.

The anticancer compound podophyllotoxin and other related lignans can be produced in Linum album in vitro cultures, although their biosynthesis varies according to the degree of differentiation of the plant material. In general, L. album cell cultures do not form the same lignans as roots or other culture systems. Our aim was to explore how the lignan-producing capacity of organogenic cell masses is affected by the conditions that promote their formation and growth. Thus, L. album biomass obtained from plantlets was cultured in darkness or light, with or without the addition of plant growth regulators, and the levels of podophyllotoxin, methoxypodophyllotoxin and other related lignans were determined in each of these conditions. The organogenic capacity of the cell biomass grown in the different conditions was studied directly and also with light and scanning electronic microscopy, leading to the observation of.several somatic embryos and well-formed shoots. The main lignan produced was methoxypodophyllotoxin, whose production was clearly linked to the organogenic capacity of the cell biomass, which to a lesser extent was also the case for podophyllotoxin.

Influence of cold pretreatment on shoot regeneration from callus in date palm (Phoenix dactylifera L.) cv. 'Barhee'.

Mass propagation of date palm through indirect somatic embryogenesis or organogenesis has attracted the interest of commercial producers. But, this technique still faces some problems that hindered the production of date palm plantlets in vitro. Tissue browning is one of the serious problems that reduce callus growth and shoot regeneration. So the objective of the present study is to investigate the effect of cold pretreatment on callus growth, shoot regeneration, and polyphenol oxidase (PPO) activity during the callus culture. Results showed that a high survival rate of callus cultures (100%) were obtained when cultures were incubated in low temperature (cold treatment) for 45 and 75 days. On the other hand, total amount on phenolic compounds was also reduced to 0.47 and 0.53 mg GAE/g after same period of incubation (45 and 75 days respectively) at low temperature. In additional, our results showed that the highest frequency of shoot formation (66.67 and 73.34, %) and the highest shoot numbers (7.8 and 8.6 shoots/100 mg) were obtained from callus treated with low temperature for 45 and 75 days, respectively.

Phytochemical and biological study of callus cultures of Tulbaghia violacea Harv. Cultivated in Egypt.

As in vitro plant cultures are used extensively to produce bioactive metabolites, our goal was to establish calli from Tulbaghia violacea Harv. flowers and assess the tissue phytochemically and biologically. Murashige & Skoog medium(MS) + 22.6 µM 2,4-dichlorophenoxyacetic acid +2.2 µM benzylaminopurine induced callus from flowers. Gas chromatography/mass spectrometry(GC/MS) analyses of n-hexane extracts of calli(HC) and flowers(HF) revealed 33 and 32 components(92.6 and 98.5%, respectively). Hydrocarbons were predominant in HC (55.0%), whereas a higher percentage of oxygenated compounds was found in HF(74.6%). Trans(E)-anethole(39.1%) and 16-hentriacontanone (30.3%) dominated in HF and HC, respectively. However, sulphur compounds were only detected in HF. Quantitative estimation of thiosulphinates, phenolics, flavonoids and saponins in ethanolic extracts of calli(EC) and flowers(EF) showed much higher contents in EF. Antioxidant, antimicrobial and cytotoxic screening of extracts demonstrated that EF was the most potent, followed by HF and EC; conversely, HC was inactive. Although HC and EC were less biologically active, these calli could be an alternative source of bioactive metabolites.

Isolation and quantification of antimalarial N-alkylamides from flower-head derived in vitro callus cultures of Spilanthes paniculata.

This is the pioneer work reporting on simple procedure for synchronized determination and quantification of two biologically active N-alkylamides, (2E,6Z,8E)-N-isobutyl-2,6,8-decatrienamide (spilanthol) and (2E,4Z)-N-isobutyl-2,4-undecadiene-8,10-diynamide (UDA), using in vitro callus cultures from flower-heads of Spilanthes paniculata. The extracts were purified using preparative thin layer chromatography (TLC) and finest separation of compounds was optimized using high performance liquid chromatography (HPLC). Eventually, N-alkylamides were validated by mass spectrometry. Linearity curve with its regression coefficients (R2) obtained for both these alkylamides was 0.99. While spilanthol was quantified using tentative standard dodeca-2(E),4(E)-dienoic acid due to the non-availability of commercial standard and the precision of a developed method was evaluated in terms of relative standard deviation by measuring inter- and intra-days variation 3.52% and 1.74%, respectively. Similarly, calibration curve was obtained for the compound UDA isolated from flower-head explants from field grown parental plant with its inter- and intra-day RSD values as 4.33% and 3.61%, respectively. With this protocol, a very high yield of 2.23 mg/g of spilanthol and 4.30 mg/g dry weight (DW) of UDA, was obtained, simultaneously, from callus cultures. Flower-heads from parent plants, used as control, showed negligible amount of spilanthol and quantity of UDA was marginally higher than that in callus cultures. The highly stable biotherapeutic spilanthol and UDA with m/z 222 and m/z 230, respectively, showed retardation of malaria parasite development through blockage at ring stage of erythrocytic schizogony and ultimately lead to parasite death. The effect on parasite was additive. This study signifies the utility of in vitro cell cultures for therapeutic compound production, throughout the year, at higher yield for down-stream applications.

Calcium pectinate gel beads obtained from callus cultures pectins as promising systems for colon-targeted drug delivery.

Low methyl-esterified pectins obtained from the cell walls of the campion (SV, SV>300), tansy (TV, TV>300) and duckweed (LM, LM>300) callus cultures and apple pectin (AP, Classic AU 701) were used as the carriers for colon delivery of prednisolone. The pectins with molecular weight more than 300kDa (SV>300, TV>300, LM>300) formed gels which exhibited the higher gel strength. The higher gel strength of these gels appeared to be related to the higher Mw and the lower degree of methylesterification (DE) of these pectins. Release aspects of prednisolone in the simulated gastric (pH 1.25), intestinal (pH 7.0) and colonic (pH 7.0+pectinase) media were investigated. The LM-5%, AP-3% and AP-5% beads destroyed in simulated intestinal medium probably due to the higher DE of the LM and AP pectins. The SV>300-3% and TV>300-3% prednisolone loaded bead systems showed a high stability at pH 1.25 and pH 7.0. Prednisolone release occurred in a larger extent in colonic medium due to the enzymatic erosion of the beads. The SV>300-3% and TV>300-3% particles showed a more controlled release that appeared to be related to the lower DE, rhamnogalacturonan content, rhamnogalacturonan I branching and the higher linearity and Mw of the TV>300 and SV>300 pectins, as well as to the higher gel strength. This in vitro study suggests that calcium pectinate gel beads obtained from callus cultures pectins can be proposed as potential systems for colon-targeted drug delivery.