Top PDF In-depth proteomic analysis of a mollusc shell: acid-soluble and acid-insoluble matrix of the limpet Lottia gigantea

In-depth proteomic analysis of a mollusc shell: acid-soluble and acid-insoluble matrix of the limpet Lottia gigantea

In-depth proteomic analysis of a mollusc shell: acid-soluble and acid-insoluble matrix of the limpet Lottia gigantea

models at present (http://genome.jgi-psf.org/Lotgi1/Lotgi1. download.html) [24], together with the corresponding reversed database and the sequences of common contami- nants, including human keratins from IPIhuman. Carba- midomethylation was set as fixed modification. Variable modifications were set as oxidation (M), N-acetyl (protein) and pyro-Glu/Gln (N-term). Initial peptide mass tolerance was set to 7 ppm and fragment mass tolerance was 20 ppm. Two missed cleavages were allowed and the minimal length required for peptide identification was seven amino acids. The peptide and protein false discovery rates (FDR) were both set to 0.01. The maximal posterior error probability (PEP) for peptides, which is the probabil- ity of each peptide to be a false hit considering identifica- tion score and peptide length [28,29], was set to 0.01. The Re-quantify and Second Peptide [30] options were enabled. At least two MaxQuant group sequence-unique peptides with a score >100 were required for protein identification. Furthermore, identifications were only accepted if the peptides were identified in at least two replicates within the respective group A, B or C. Identifications with only two unique peptides were manually validated considering the assignment of major peaks, occurrence of uninter- rupted y- or b-ion series of at least 4 consecutive amino acids, preferred cleavages N-terminal to proline bonds, the possible presence of a2/b2 ion pairs and immonium ions, and mass accuracy. The ProteinProspector MS- Product program (http://prospector.ucsf.edu/) was used to calculate the theoretical masses of fragments of identified peptides for manual validation. BLAST and FASTA searches against non-redundant databases (all organisms) were performed using the programs provided by NCBI (http://www.ncbi.nlm.nih.gov/blast) and EBI http:// www.ebi.ac.uk/Tools/sss/. Domains were predicted with InterProScan (http://www.ebi.ac.uk/Tools/pfa/iprscan/) and PROSITE (http://prosite.expasy.org/). For sequence alignments we employed Kalign (http://www.ebi.ac.uk/ Tools/msa/kalign/) and ClustalW (http://www.ebi.ac. uk/Tools/msa/clustalw2/). Sequence repeats were predicted using RADAR (http://www.ebi.ac.uk/Tools/ Radar/index.html). The abundance of proteins was estimated by calculating the exponentially modified protein abundance index (emPAI) [33]. Observable peptides were determined and counted with Protein Prospector (http://prospector.ucsf.edu/prospector/cgi-bin/ msform.cgi? form = msdigest) using zero miss-cleavages, a peptide mass of 700–2800, and a minimal peptide length of seven amino acids. Observed unique parent ions with a minimal length of seven amino acids and a mass between 700 – 2800 used for emPAI calculation included ions with up to two miss-cleavages, modifications specified for MaxQuant analysis (see above), different charges, and neutral losses [33]. Proteins with emPAI ≥ 9 were referred to as major proteins in this report.
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In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

Acid-soluble and acid-insoluble matrix components were separated by ultracentrifugation (Optima LE 80 K, 45Ti rotor, Beckman Coulter, Krefeld, Germany) at 4 °C and 146,900 x g for 60 min. The fractions were then lyophi- lized for concentration and storage. Matrix proteins were separated by SDS-PAGE in pre-cast 4–12% Novex Bis-Tris gels using the MES buffer system with reagents and protocols supplied by the manufacturer (Invitrogen, Carlsbad, CA) except for the reducing agent, which was β-mercaptoethanol added to a final concentration of 2%. The sample buffer contained lithium dodecyl sulphate (LDS, final concentration 1%) while pre-cast gels and run- ning buffer contained SDS (0.1%). Samples were sus- pended in 30 μl sample buffer/200 μg of organic matrix, boiled for 5 min, and centrifuged at 13000 rpm for 5 min in an Eppendorf bench-top centrifuge before SDS-PAGE analysis. Separated proteins were stained with colloidal Coomassie blue (Invitrogen). Gels containing acid-soluble nacre matrix and acid-insoluble prismatic layer matrix were cut into 12 slices and identical slices of three lanes were used for in-gel digestion with trypsin [27]. All slices were treated equally irrespective of staining intensity or presence of visible bands. The eluted peptides were cleaned with C18 Stage Tips before MS analysis [28]. The acid-soluble fraction of the prismatic layer and the acid-insoluble matrix of nacre, PAGE analysis of which showed no or only few and week protein bands, respect- ively, were cleaved using a filter-aided sample preparation (FASP) method [29, 30] modified as follows. Matrix com- ponents were dissolved in 0.1 M Tris buffer, pH 8, con- taining 6 M guanidine and 0.01 M dithiothreitol (DTT) and heated to 56 °C for 60 min. Aliquots containing 200, 400 and 800 μg of matrix were then loaded onto Microcon YM-30 centrifugal filter devices (Millipore) and DTT was removed by centrifugation at 13000 rpm in a Eppendorf bench top centrifuge model 5415D for 10 min and wash- ing with 2 x 1vol of the same buffer. Carbamidomethyla- tion was performed in the device using Tris-guanidine buffer containing 0.05 mM iodoacetamide and incubation for 45 min in the dark. Carbamidomethylated proteins were washed with 0.05 M ammonium hydrogen carbonate buffer, pH 8, containing 2 M urea, and centrifugation as before. Trypsin (2 μ g, Sequencing grade, modified; Pro- mega, Madison, USA) was added in 40 μl of the same buf- fer and the devices were incubated at 37 °C for 16 h. Peptides were collected by centrifugation and the filters were washed twice with 40 μl of buffer. The peptide solu- tion was acidified to pH 1–2 with trifluoroacetic acid and peptides were cleaned and concentrated using C18 Stage Tips [28].
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The Lottia gigantea shell matrix proteome: re-analysis including MaxQuant iBAQ quantitation and phosphoproteome analysis

The Lottia gigantea shell matrix proteome: re-analysis including MaxQuant iBAQ quantitation and phosphoproteome analysis

In search of the reasons for apparent differences in previ- ously published Lottia shell proteomes [17,18] we noticed that database searches were done using the AllModels database in [18] while [17] used the FilteredModels data- base containing entries supported by EST sequences. Therefore we re-analyzed the raw-files produced previ- ously for acid-soluble and acid-insoluble matrix prepared according to method B [17] (also used to identify phos- phoproteins in the present report) using a combination of both databases and a subset of Uniprot containing Lottia + gigantea entries. Furthermore, to determine the approximate abundances of the identified proteins, the iBAQ (intensity-based absolute quantification) method [19] as implemented in more recent MaxQuant versions was enabled in this search. The previously used [17] emPAI method [41] belongs to the spectral count methods based on counting the number of identified unique parent ions per protein. In contrast, iBAQ and similar algorithms are called intensity-based because they calculate the sum of parent ion intensities of identified peptides per protein. In both types of methods, the numbers of theoretically possible peptides per protein for the protease used in sam- ple preparation enter the equation to account for different protein lengths and distribution and frequency of cleavage sites. Comparison of the two different types of methods show a higher accuracy of the intensity-based methods, including iBAQ (for instance [42]), indicating that they
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PHARMACEUTICO- ANALYTICAL STANTARDIZATION OF MRUDDARASRUNGA BHASMA .......

PHARMACEUTICO- ANALYTICAL STANTARDIZATION OF MRUDDARASRUNGA BHASMA .......

Moisture value of the final product was very less i.e. 1.1%w/w which indicates the preparation is almost free from the mois- ture, suggesting its long shelf life. Total Ash value ofMruddarasrungaBhas- mais77.7% w/w. The higher Total ash value indicative of presence of very high inorganic content..Acid insoluble Ash val- ue of the drug was 61.9 % w/w .which in- dicates the solubility of Lead oxide in HCl and insoluble nature of the complex com- pounds formed during the Marana and Bhavana. Water-soluble ash value 1.7% w/w was less due to presence of less quan- tity of water-soluble salts. pH value of Mruddarasrungabhasma is 6.67 suggests that is almost neutral in nature. The XRD report shows there is no Peak showing the presence of free lead. This supports the safety of Mruddarasrunga Bhasma. The Zeta Potential value of Mruddarasrun- gaBhasma was found 125.2mVwhich in-
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A study of the fatty acid profile in the muscle of Monopterus chuchia

A study of the fatty acid profile in the muscle of Monopterus chuchia

A common constituents of glycerides of human adipose tissue, found in high concentration in liver, is palmitoleic acid. It is biosynthesized from palmitic acid, which is also detected in this fish species. Palmitoleic acid is present in considerable amount in Monopterus chuchia. It is a beneficial fatty acid which increases insulin sensitivity by suppressing inflammation and inhibits the destruction of pancreatic beta-cells which are known to secrete insulin 27 . Linoleic acid, which is a precursor of n-6

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Comparative study on Physico & Phyto-Chemical analysis of Persea americana & Actinidia deliciosa

Comparative study on Physico & Phyto-Chemical analysis of Persea americana & Actinidia deliciosa

Abstract- The main aim of the present investigation is to determine the Moisture Content, Total ash, Acid insoluble ash, Water soluble ash, Alcohol soluble Extractive value and proximate analysis of Persea americana & Actinidia deliciosa fruit. The fruit samples were shade dried, powdered and used for further analysis. The phytochemicals were studied using the six different solvents extracts such as aqueous, ethanol, ethyl acetate, chloroform, petroleum ether and methanol. Among them aqueous, ethanol, ethyl acetate extract has shown the presence of potential components which act as antioxidants.
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Chromatographic estimation of maturity based phytochemical profiling of Ipomoea mauritiana

Chromatographic estimation of maturity based phytochemical profiling of Ipomoea mauritiana

High Performance Liquid Chromatography: The aqueous extracts of different samples of mature and immature tubers of I. mauritiana were subjected to HPLC analysis. Samples collected from different stages of maturity were used for analysis. The HPLC profile of mature and immature tubers remains same but the aqueous extracts of mature and immature tubers consistently exhibited a quantitative difference in the phytoconstituents particularly those eluting at 11 minute and 15 minute retention time (RT) in the HPLC chromatograms. It is interesting to note that the peak at 15 minute decreases with increasing girth quite significantly while at 11 minute peak increased. The ratio of 10 peaks at 11 minute (A) to 15 minute (B) RT increased with size (girth) of the tubers. Coupled HPLC chromatogram of stagewise tubers at
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Preliminary phytochemical and physicochemical investigation of Woodfordia fruticosa (linn) kurz root

Preliminary phytochemical and physicochemical investigation of Woodfordia fruticosa (linn) kurz root

for 5 min. The watch glass was rinsed with 5 ml of hot water, and the liquid was added to the crucible. The insoluble matter was collected on an ashless filter paper and washed with hot water until the filtrate was neutral. The filter paper containing the insoluble matter was transferred to the crucible, dried on a hot plate, and ignited to a constant weight. The crucible was cooled in desiccators and weighed. The content of acid-insoluble ash was calculated in mg per gm of air-dried material and expressed as a percentage. The experiment was carried out in triplicate. The content of acid-insoluble ash was calculated as a percentage of ash with reference to the air-dried root powder. The result showed in Table 5.
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Available online www.jocpr.com Journal of Chemical and Pharmaceutical Research, 2016, 8(1):7-12

Available online www.jocpr.com Journal of Chemical and Pharmaceutical Research, 2016, 8(1):7-12

The physicochemical constants of the plant Drymoglossum heterophyllum-(L)C.Chr. is as follows. Total ash value, Water soluble ash, Acid insoluble ash value of the shade dried leaf of was found to be 7.75% w/w, 4.42% w/w and 3.466% w/w successively. Ash values within fair limits, signifies the quality and purity of the Drymoglossum heterophyllum-(L)C.Chr and also gives idea about the total inorganic content. Alcohol soluble extractive value and Water soluble extractive value of the shade dried leaves was found to be4.157 % w/w. and 7.85% w/w. Moisture content of the shade dried leaves was found to be 8.65% w/w. Crude fibre content of the shade dried leaves was found to be 18.18% w/w. The chlorophyll content of the leaves was also estimated. The total chlorophyll content was 0.449/g tissue.
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Proteomic analysis of small acid soluble proteins in the spore core of Bacillus subtilis ΔprpE and 168 strains with predictions of peptides liquid chromatography retention times as an additional tool in protein identification

Proteomic analysis of small acid soluble proteins in the spore core of Bacillus subtilis ΔprpE and 168 strains with predictions of peptides liquid chromatography retention times as an additional tool in protein identification

Background: Sporulation, characteristic for some bacteria such as Bacillus subtilis, has not been entirely defined yet. Protein phosphatase E (PrpE) and small, acid soluble spore proteins (SASPs) influence this process. Nevertheless, direct result of PrpE interaction on SASPs content in spore coat of B. subtilis has not been evidenced so far. As proteomic approach enables global analysis of occurring proteins, therefore it was chosen in this experiment to compare SASPs occurrence in two strains of B. subtilis, standard 168 and ΔprpE, lacking PrpE phosphatase. Proteomic analysis is still a challenge, and despite of big approach in mass spectrometry (MS) field, the identification reliability remains unsatisfactory. Therefore there is a rising interest in new methods, particularly bioinformatic tools that would harden protein identification. Most of currently applied algorithms are based on MS- data. Information from separation steps is not still in routine usage, even though they also provide valuable facts about analyzed structures. The aim of this research was to apply a model for peptides retention times prediction, based on quantitative structure-retention relationships (QSRR) in SASPs analysis, obtained from two strains of B. subtilis proteome digests after separation and identification of the peptides by LC-ESI-MS/MS. The QSRR
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PHARMACOGNOSTICAL AND PRELIMINARY PHYTOCHEMICAL PROPERTIES OF AEGLE MARMELOS L  CORREA FRUIT PULP

PHARMACOGNOSTICAL AND PRELIMINARY PHYTOCHEMICAL PROPERTIES OF AEGLE MARMELOS L CORREA FRUIT PULP

Fluorescence Analysis: Fruit pulp powder were subjected to analyze fluorescence features under ultra violet light and day light after giving treatment for 48 hours with various chemical and organic solvents like ethanol, 50% sulphuric acid, 10% sodium hydroxide and dilute hydrochloric acid 10,11 .

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PHARMACOGNOSTICAL AND PHYTOCHEMICAL EVALUATION OF STEM OF CISSUS QUADRANGULARIS L

PHARMACOGNOSTICAL AND PHYTOCHEMICAL EVALUATION OF STEM OF CISSUS QUADRANGULARIS L

The stems of Cissus quadrangularis L. (Vitaceae) are reported to have great medicinal value. The present investigation was therefore undertaken to determine the requisite pharmacognostic standards for evaluating the plant material. The macroscopic features of the Cissus quadrangularis L. stem were observed under magnifying lens. Microscopic characters and powder analysis were determined under microscope. The physiochemical properties such as loss on drying, total ash value, acid insoluble ash value, water soluble ash value, pH, and extractive values of stem were carried out. The microscopic study showed abundant large cells of mucilage, clusters and bundles of acicular crystals of calcium oxalate scattered throughout the section. The powder microscopy showed the presence of annular, reticulated and boarded pitted thickening xylem vessels, cluster, rosette and acicular crystals of calcium oxalate and starch grains. Phytochemical analysis showed the presence of many important classes of phytoconstituents like alkaloids, flavonoids, cardiac glycosides and triterpenes. The determination of these characteristics will aid future investigators in their pharmacological analyses of this species.
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Quality control standards for the roots of Three Plumbago species

Quality control standards for the roots of Three Plumbago species

in P. zeylanica and 8.11% in P. capensis (P. rosea>P . zeylanica>P. capensis). Crude fi bre consists largely of cellulose and lignin (97%) plus some mineral matter. It represents only 60 to 80% of cellulose and 4 to 6% of lignin. It is useful as a measure of nutritive value of animal feeds and also in the analysis of various foods and food products to detect adulteration, quality and quantity. The fibre content was accounted to be 14.30% in P. zeylanica, 12.51% in P. capensis and 9.79% in P. rosea (P. zeylanica>P. capensis>P. rosea). The data evolved can be considered for laying down the pharmacopoeial standards for the roots of the three Plumbago species.
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Phytochemical Screening of Curculigo Orchioides Gaertn  Root tubers

Phytochemical Screening of Curculigo Orchioides Gaertn Root tubers

Phytochemical screening, mineral analysis and powder analysis was carried out on the root tubers of “Curculigo orchioides Gaertn” having short or elongated fleshy roots. Recommended as choice of drug for its aphrodiasiac, galactogogue and other miscellaneous uses in traditional system of medicine.

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Characterization of Acid Soluble Collagen from Skins of Surf Smelt (Hypomesus pretiosus japonicus Brevoort)

Characterization of Acid Soluble Collagen from Skins of Surf Smelt (Hypomesus pretiosus japonicus Brevoort)

2.7. Amino Acid Analysis of Acid-Soluble Collagen from the Skins of the Surf Smelt An amino acid analysis was performed to clarify the re- lationship between the contents of the imino acids and the denaturation temperature of the collagen sample. The collagens were hydrolyzed under reduced pressure in 6 M HCl at 110 ℃ for 24 h, and the hydrolysates were ana- lyzed using a JASCO liquid-chromatography system by on-line precolumn derivatization with o-phthalaldehyde. This system consisted of a JASCO PU-2080 plus intelli- gent HPLC-pump, a JASCO FP-2020 plus intelligent fluorescence detector, a JASCO CO-2060 plus intelligent column thermostat, a JASCO DG-2083-53 3-line degas- ser, a JASCO LG-2080-02 ternary gradient unit, a JASCO AS-2057 plus intelligent sampler, and a JASCO CrestPak C18S (φ4.6 × 150 mm) reversed-phase column. The excitation and emission wavelengths were set at 345 and 455 nm, respectively.
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 EXPLORING THE PHARMACOGNOSTIC CHARACTERISTICS AND ANTIMICROBIAL POTENTIAL OF LEAVES OF URENA LOBATA LINN.

 EXPLORING THE PHARMACOGNOSTIC CHARACTERISTICS AND ANTIMICROBIAL POTENTIAL OF LEAVES OF URENA LOBATA LINN.

Antibiotics are very effective against bacterial infections but pose a threat of developing allergies or resistant bacterial species. Herbal medicines are therefore comparatively safer for treating bacterial infections. Here an attempt was made to explore the antimicrobial activity of a less renowned herbal drug Urena lobata. Linn. According to the pharmacognostic studies performed; The values of total ash, water soluble ash and acid insoluble ash were found to be 11.67 %w/w, 3.50%w/w and 4.24%w/w respectively. All types of ash values of the crude drug were found to be in agreement with the earlier studies 21,30 .
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Chemical and spectroscopic characterization of insoluble and soluble humic acid fractions at different pH values

Chemical and spectroscopic characterization of insoluble and soluble humic acid fractions at different pH values

The elemental composition of the humic fractions is re- ported in Table 1 on a moisture- and ash-free basis. There- fore, the following considerations concern the organic component of each fraction. While the insoluble fractions at pH 1 and 3 were richer in C and poorer in O than the soluble fractions, the O increased at pH 5 and 7 indicating that the precipitation pH of the HA affected the distribu- tion of the functional groups between the precipitated and the liquid phase. The C content was lower at pH 5 and 7 than at pH 1 and 3 in the insoluble fractions, while the opposite was true for the soluble fractions. The O content and O/C ratio were lower in the insoluble fractions at pH 1 and 3 than at pH 5 and 7 indicating that polarity in- creased along with pH, while the opposite occurred in the soluble fractions. In the case of the N content, no clear trend could be identified indicating that pH did not affect the distribution of the N-containing moieties in the solid or liquid phase. The H content was higher at pH 5 and pH 7 than at pH 1 and pH 3 in both the insoluble and sol- uble fractions. This would appear to be an artifact, since the sum of each element present in the soluble and insol- uble fractions deriving from the same alkali solution should be the same at any pH. The highest H contents were found in the fractions exhibiting the highest ash con- tents. This suggests that the presence of inorganic compo- nents could induce an overestimation of the H content. One explanation for this result is that the elemental ana- lysis includes a high-temperature incineration step (up to 1,000°C) which could provoke the release of some
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“Pharmacognostic and Phytochemical Evaluation of Chrysophyllum cainito linn. Leaves” by Sunita Shailajan, Deepti Gurjar, India.

“Pharmacognostic and Phytochemical Evaluation of Chrysophyllum cainito linn. Leaves” by Sunita Shailajan, Deepti Gurjar, India.

The evaluation of pharmacognostical parameters may ensure the identity and authenticity of C. cainito leaves. The plant was found to be a good source of ursolic acid, β-sitosterol, lupeol and gallic acid by HPTLC analysis. The validated HPTLC methods can be applied to various herbal drugs for the quantitation of pharmacologically active markers- ursolic acid, β-sitosterol, lupeol and gallic acid. Data generated from the acute oral toxicity study may ensure an adequate safety margin of C. cainito leaves for their intended use. Thus, findings of this study could be useful for the compilation of a monograph in a suitable pharmacopoeia for its proper identification and quality control, which can be used globally.
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Pharmacognostic Evaluation of Leaves and Stem of Murraya koenigii

Pharmacognostic Evaluation of Leaves and Stem of Murraya koenigii

Murraya koenigii has been widely used as medicine in indigenous system of medicine. It is used as an analgesic, febrifuge, stomachic, carminative and for the treatment of dysentery skin eruption. The present study was planned for details pharmacognostical studies of Murraya koenigii leaves and stem. Macroscopical characters and microscopical characters of leaves and stem were perceived. The Physicochemical properties (total ash, acid insoluble ash, water soluble ash, water soluble extractive, methanol soluble extractive, moisture content and foaming index) of bark powder were studied. The values of physicochemical can also be used for standardization of Murraya koenigii. These studies provided referential information in regard to its identification parameters assumed significantly in the way of acceptability of herbal drugs in present scenario of lack of regulatory laws to control quality of herbal drugs.
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An assessment of Antioxidant potential and in-vitro SPF activity of Amaranthus tricolor and Curcuma longa

An assessment of Antioxidant potential and in-vitro SPF activity of Amaranthus tricolor and Curcuma longa

used to determine the moisture content in herbs as per standard. Ash value used to determine the quality and purity of herbs. Ash values are indicative to some extent of care taken in collection and preparation of drug for market and of foreign matter content of natural drug. The object of ashing is to remove all traces of organic material interfering in an analysis of inorganic constituents. Adhering dirt, sand as well as variation caused by calcium oxalate may be determined by acid-insoluble ash content. Total ash value usually consists of carbonates, phosphates, silicates & silica. The acid insoluble ash consists of mainly silica and indicates contamination with earthy matter. The water soluble ash is used to determination of the amount of inorganic elements present in drugs. The extractive values are useful to determination of the chemical constituents present in the crude drug and also help in estimation of specific constituents soluble in a particular solvent.
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