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Phytochemical analysis of different species of gentiana from Nepal
(2017) Shakya, Niroj; Deepak Raj Pant
Nepal houses 44 different species of Gentiana. Plants belonging to genus Gentiana are
very well-known for their pharmacological properties. Ethnopharmacological data show
that various species of Gentiana have been effectively used in herbal medicine to treat
various ailments. They are known to possess potent therapeutic compounds like iridoids,
xanthones and glucoflavones. This study attempts to evaluate phytochemical,
antioxidant and antibacterial activities and estimate three bioactive compounds:
swertiamarin, amarogentin and mangiferin in different species of Gentiana collected
from different parts of Nepal.
Methods: Total phenolic and flavonoid contents were quantified spectrophotometrically
and in vitro DPPH free radical scavenging assay was measured. Agar well diffusion
method was employed for antibacterial assays. Thin Layer Chromatography was
performed on TLC Aluminium plates pre-coated with silica gel 60 for identification and
estimation of swertiamarin, amarogentin and mangiferin. Semi-quantitative estimation
was done using GelQuant.NET software using the standard compounds.
Results: The quantitative phytochemical analysis showed presence of higher amounts of
polyphenol and flavonoid in methanol extracts as compared to the aqueous extract of
various Gentiana species. Among methanol extracts highest concentration of polyphenol
was observed in G. depressa (79.2±19.19mgGAE/gm) while G. capitata (44.6±3.97
mgGAE/gm) showed the lowest content. Similarly, G. capitata recorded the highest total
flavonoid content (19.09±0.97mgQE/gm) and G. ornata (11.31±0.49 mgQE/gm) showed
the least content. Likewise, the methanol extracts showed more promising antioxidant
activity compared to the aqueous extracts. The methanol extract of G. depressa showed
the best antioxidant activity among the different Gentiana sps with an IC
50 value of39.57±0.95 µg/ml. Semi-quantitative analysis showed that swertiamarin was present in
higher quantities than amarogentin and mangiferin. Highest concentration of
swertiamarin and mangiferin was identified in G. ornata (0.109±0.013 mg/gm DW and
0.018±0.001 mg/gm DW respectively) while G. capitata possessed highest concentration
of amarogentin (0.075±0.005 mg/gm DW). Antibacterial assay showed varying degree of
sensitivity to various pathogenic microorganisms. G. ornata was showed highest activity
among all plant species against K. pneumonia and E. feacalis while G. depressa showed
maximum sensitivity to S. aureus and moderate sensitivity to K. pneumonia. These
results clearly show that crude methanol and aqueous extract of various Gentiana
species possess potent pharmacological compounds, and justified the folkloric use of
these plants in digestive and respiratory ailments.
Keywords: Gentiana, antioxidant, antibacterial, amarogentin, swertiamarin, mangiferi
Validation of Saliva Biospecimen; an Alternative to Nasopharyngeal Swab detection of Sars- Cov 2using TR- PCR
(2022) Maharjan, Salin; Krishna Das Manandhar
Extensive and widespread testing is necessary to control the rapid spread of SARS-CoV-2
pandemic which has been rampant since its origin in 2019. The current gold standard for
COVID testing, nasopharyngeal sampling (NPS) method presents various obstacles for a
successful mass testing like need of highly trained personnel, transport media, personal
protective equipment (PPE). Hence, we have tried to establish saliva sample as an
alternative to NPS. In this study, patients visiting the KM biotech lab for PCR test were asked
to provide saliva samples in addition to NPS sample. The saliva sample is then heat killed
and directly processed for PCR amplification. The corresponding data from NPS and saliva
samples are compared to gauge efficiency of saliva against NPS. We observed that the saliva
is effective as NPS with sensitivity (80%) and specificity (91%) [P<0.01]. We found that 60%
(110/180) samples tested positive in either or both the NPS and saliva test, while 21(11%)
samples tested positive in NPS only and 7(4%) samples tested positive in saliva only. In
conclusion, saliva assay demonstrated significant results against the NPS samples. Saliva
assay could prove effective in controlling COVID pandemic when compared to painful and
resource intensive NPS sample. This study is relevant, especially in case of countries like
Nepal, where we are completely dependent on international market for transport mediums,
PPE, swabs etc. Saliva assay could reduce our dependence significantly as it has substantially
less resource requirements.
Keywords: SARS-CoV-2, COVID-19, Saliva assay, NPS, COVID testing, RT-PCR
Morphological and Molecular Charactrization of Gastrointestinal Parasites of wild water Buffalo of Chitwan National Park
(2021) Aryal, Menuka; Kriashna Das Manandhar
The microscopy-based identification of gastrointestinal parasites can be further
strengthened at species-level, which shows the importance of molecular based
identification.The main aim of this study is the molecular characterization of the
gastrointestinal parasites in Wild Water buffalo of Chitwan National Park. A wide range of
helminth and protozoan species can infect or colonize the gastrointestinal tract of humans
and animals. Intestinal parasitic infections caused mainly by protozoa and helminths are
most prevalent in Wild Water buffalo, which are of relevance to critically endangered
species. Twenty-five fecal samples were collected from groups of Wild Water buffalo. The
fecal samples were examined microscopically by direct wet mount method,
sedimentation method, floatation method, and molecularly by PCR, sequencing and,
analysis. An integrative taxonomic approach is followed, where conspicuous
morphological traits and phylogenetic trees based on DNA sequences are used to test the
validity of the species.
Nine intestinal parasites were detected microscopically, where the infection rate was
96%, of which 88% and 84% were due to protozoa and helminths respectively. The
identified protozoan parasites were Entamoeba (80%), Eimeria (60%), Balantidium (8%),
and Cryptosporidium (4%) and helminths were Oxyruid (64%) Strongyloides (28%),
Strongyle (36%), Eurytrema (8%), and Paramphistomum (4%). The PCR assays were
conducted using Entamoeba spp. 18S specific primers. Ribosomal genome sequence (550
bp) was obtained and DNA sequence analysis enabled more precise identification of E.
bovis. Entamoeba was successfully sequenced and it was separated from those of other
Entamoeba species/genotypes in phylogenetic analysis by the Neighbor Joining method.
Also the amplification of Eimeria was done by using genus-specific primers targeting 18S
ribosomal region. This investigation gives benchmark data of GI parasites and their
conveyance in wild buffalo. This is the primary report for the molecular discovery of E.
bovis in translocated wild buffalo in Nepal.
Keywords: Bubalus arnee, Chitwan National Park, Gastrointestinal Parasite, Protozoa,
Helminthes, 18S rDNA