This document describes the development and validation of a gas chromatography method for determining fatty acids and sterols in total diet samples. Key points:
- A method was developed using accelerated solvent extraction to obtain lipid extracts from samples, followed by saponification to isolate fatty acids and sterols without derivatization.
- The method was validated and found to be accurate (mean recovery 80.8-98.2%) and precise (RSD <8.6%) for quantifying the most abundant fatty acids and sterols.
- The method was then applied to analyze 28 total diet samples. Results showed the most abundant fatty acid was monounsaturated C18:1 at 34.5% of total fatty
In this work a new prodrug polymer was
prepared with two attachment groups (amid-ester), using di
functional spacer such as ethanol amine, which could react with
polyacrylic acid producing amide group, with remain ethanol
terminal group which could react with captopril acyl chloride,
producing ester group with extended the arm substituted drug to
improve the hydrolysis and to prevent the steric effect of polymer
chains. Many advantages enhanced the prodrug of polymer. The
prepared polymers were characterized by FTIR, 1H –NMR
spectroscopies. Controlled drug release was studied in different
pH values at 37℃, using UV. Spectra with comparing with
calibration curve. The modification percentage test was studied,and swelling percentage was calculated and all physical properties were observed.
The present paper reports the kinetics of soybean and sunflower oils’ ethanolysis. The transesterification reaction was carried out using a molar ratio of ethanol to oil of 9:1 and 1.0 wt% of sodium ethoxide as catalyst under stirring of 400 rpm. The reactions were performed in a stirred batch reactor at three different temperatures (308.15, 323.15 and 338.15 K) over a period of 120 min. The concentration of compounds was analyzed by High-Performance Size Exclusion Chromatography (HPSEC). The kinetic model assumed that ethanolysis occurs in a sequence of three reversible steps with the production of di- and monoacylglycerols as intermediate components. Based on the modeling approach it was possible to determine the rate constants of reaction and activation energies for the transesterification process of soybean and sunflower oils. Despite the phase splitting, no mass transfer control was observed and the proposed mathematical model fitted well the experimental data.
EFFECT OF MICROBIAL TRANSGLUTAMINASE (MTGASE) AND ISOLATED SOY PROTEIN (ISP) ...Hazmi Y
This study was carried out to determine the effects of Microbial Transglutaminase (MTGase) and isolated soy protein (ISP) on tilapia (Oreochromis niloticus) surimi gels. MTGase at 0, 0.3, 0.6 percent and ISP at 0, 3 and 6 percent were added to surimi gels of tilapia.
In this work a new prodrug polymer was
prepared with two attachment groups (amid-ester), using di
functional spacer such as ethanol amine, which could react with
polyacrylic acid producing amide group, with remain ethanol
terminal group which could react with captopril acyl chloride,
producing ester group with extended the arm substituted drug to
improve the hydrolysis and to prevent the steric effect of polymer
chains. Many advantages enhanced the prodrug of polymer. The
prepared polymers were characterized by FTIR, 1H –NMR
spectroscopies. Controlled drug release was studied in different
pH values at 37℃, using UV. Spectra with comparing with
calibration curve. The modification percentage test was studied,and swelling percentage was calculated and all physical properties were observed.
The present paper reports the kinetics of soybean and sunflower oils’ ethanolysis. The transesterification reaction was carried out using a molar ratio of ethanol to oil of 9:1 and 1.0 wt% of sodium ethoxide as catalyst under stirring of 400 rpm. The reactions were performed in a stirred batch reactor at three different temperatures (308.15, 323.15 and 338.15 K) over a period of 120 min. The concentration of compounds was analyzed by High-Performance Size Exclusion Chromatography (HPSEC). The kinetic model assumed that ethanolysis occurs in a sequence of three reversible steps with the production of di- and monoacylglycerols as intermediate components. Based on the modeling approach it was possible to determine the rate constants of reaction and activation energies for the transesterification process of soybean and sunflower oils. Despite the phase splitting, no mass transfer control was observed and the proposed mathematical model fitted well the experimental data.
EFFECT OF MICROBIAL TRANSGLUTAMINASE (MTGASE) AND ISOLATED SOY PROTEIN (ISP) ...Hazmi Y
This study was carried out to determine the effects of Microbial Transglutaminase (MTGase) and isolated soy protein (ISP) on tilapia (Oreochromis niloticus) surimi gels. MTGase at 0, 0.3, 0.6 percent and ISP at 0, 3 and 6 percent were added to surimi gels of tilapia.
Industrial fermentation-Does Fermentation Really Increase the Phenolic Conten...ShreyaMandal4
Decortication leads to a reduction in minerals, fibers, and antioxidants as phenolic compounds located in the peripheral parts of the grains. Fermentation can be applied to treat the whole nondecorticated flour to prevent functional loss. Bringing out the nutritional benefits of millets upon fermentation will serve us to include millets at a proportion in our meals along with traditional cereals.
Fatty alcohol. Define fatty alcohols Describe the production processes of fatty alcohols and its derivatives Draw the flow chart of fatty alcohol production Explain the uses and application of fatty alcohols.
3. Definitionof Fatty Alcohols Fatty alcohols are the workhorse raw materials that facilitate the existence of products such as shampoos, shaving creams, laundry detergents, etc, and are produced at a rate of about one-and-a-half million tonnes per year and growing. Fatty alcohols are oleochemicals derived from vegetable feedstocks. The feedstock raw materials include coconut and palm kernel oils. These refined vegetable oils are first converted to a methyl ester or fatty acid. This reaction generates crude glycerine. The intermediate methyl ester or fatty acid are then fractionated and hydrogenated to produce fatty alcohol. Sources : http://www.pgchemicals.com/products/fatty-alcohols/
4. Chemical Equation for Fatty Alcohol Production Sources : http://www.pgchemicals.com/products/fatty- alcohols/
5. Block diagram of Fatty Alcohol production process
6. Fatty acids are converted into methyl ester and hydrogenated into fatty alcohols.
7. Sources : http://www.abq.org.br/workshop/11/ADRIANO- SALES-%20FIRJAM_Oleochemicals-from-Palm-Kernel- Oil.pdf
8. Hydrogenation All natural fatty alcohol processes are based on renewable fats and oils like coconut, palm oil,palm kernel,rope seed and soya bean oil. It has been proven that hydrogenation of methyl esters are preferred alternatives than hydrogenating the oils directly. Using fixed bed hydrogenation process offers the advantage of lower hydrogenation temperatures and pressures. Using special catalysts, this process is able to produce unsaturated fatty alcohols too. To produce fatty alcohols, there are three routes which is acid route,ester route and wax ester route that are shown in the following block diagrams.
9. - Acid route - Ester route - Wax ester route
10. Acid Route
Analysis of Sugars in Honey Using the PerkinElmer Altus HPLC System with RI D...PerkinElmer, Inc.
Honey consumption has grown significantly during the last few decades due to its high nutritional value and unique flavor. The price of natural bee honey is much higher than other sweeteners making it susceptible to adulteration with cheaper sweeteners, primarily sucrose. Besides lower levels of nonsugar ingredients, natural honey primarily consists of glucose and fructose and may contain low levels of sucrose and/or maltose. However, according to the international regulations, any commercially available “pure”-labeled honey products that are found to have in excess of 5% by weight of sucrose or maltose are considered to be adulterated. With the focus on possible honey adulteration, this application highlights the LC separation of various sugars found in honey and the analysis of these components in four store-bought honey samples. Method conditions and performance data, including linearity and repeatability, are presented.
Study on Fructooligosaccharide (Fos) Production By Enzyme Pectinex Ultra Sp-l...IJAEMSJORNAL
Fructooligosaccharide (FOS) is a new alternative sweetener with its characteristics such as low energy and safety for people with diabetes. Pectinex Ultra SP-L which has fructosyltransferase, catalyzes the reaction to produce short chain fructooligosaccharides. The research was conducted to enhance the high FOS content in process of FOS production by immobilized enzyme. Results achieved by Empirical planning Design Expert 7.0 - central composite method (CCD) identified at optimum conditions for process of immobilized enzyme with alginate 3.3 (%), ratio of enzyme: alginate was 0.79 (w/w), CaCl2 3.75 (%). Efficiency of loading protein reached 73.32 (%). Reaction conditions: 60 (oC), shaking velocity 90 (rpm), the initial sucrose concentrations of 50 (%), pH 5.75. When produced in 20 (h), the reaction obtained the highest level of FOS with column reaction system. The FOS for producted by immobilized enzyme achieved 47.87 (%), of which 1-kestose obtained 37.06 (%), the remaining was 10.81 (%) including nystose and fructofuranosylnystose. Efficiency of producing FOS by using immobilized enzyme compared to the free enzyme was high up to 89.49 (%).
FATS AND OILS- Fats and oils are triglycerides(triesters).They are made from glycerol and fatty acids.Fats are solids at room temperature whereas oils are liquids.Fatty acids present in fats and oils can be saturated or unsaturated.Fats and oils store energy and help to insulate the body, cushion and protect organs.
SYNTHETIC DETERGENTS- Synthetic detergents or soapless soaps are synthetic substances that are being increasingly employed as cleansing agents these days.
The objective of the current investigation is to formulate sustained release microspheres, containing Metformin hydrochloride and Glipizide as model drugs. Eudragit RSPO, Eudragit RLPO, Ethyl cellulose and Hydroxy propyl methyl cellulose, polymers of different permeability characteristics were used in combination to prepare different microspheres. Metformin and Glipizide both are type II antidiabetic agents when administered together shows synergetic effect in their action. Microspheres were prepared by emulsion solvent evaporation method with different stabilizer concentration and at different speeds of emulsification while maintaining constant amounts of Metformin and Glipizide. Drug excipients compatibility study was performed prior to formulation development and only compatible excipients were used in the fabrication of microspheres. Prepared microsphere formulations were characterized by percentage yield, particle size analysis, entrapment efficiency, in-vitro release behavior, FTIR, differential scanning colorimetry (DSC) and scanning electron microscopy (SEM). SEM studies showed that the microspheres were spherical with rough surface morphology. The drug loaded microspheres showed 29-90% entrapment capacity for Metformin and Glipizide. The in-vitro release profile showed a slow and steady release pattern for both Metformin and Glipizide. A 100% Metformin was releases within a period of 12 hrs while only 30% Glipizide was released during this time. The Metformin HCl release was found to be best fitted with Higuchi and Zero order and for Glipizide best fitted with zero order and first order respectively in single and combined polymeric loaded microspheres. DSC results indicated that the physical state of the drug was changed upon fabrication. As a result of these experiments, it was conclude that, novel sustained release oral microspheres comprising a combination of Metformin and Glipizide were successfully prepared using ethyl cellulose, HPMC 15CPS, Eudragit RSPO and Eudragit RLPO as the polymer and using emulsion solvent evaporation technique.
Industrial fermentation-Does Fermentation Really Increase the Phenolic Conten...ShreyaMandal4
Decortication leads to a reduction in minerals, fibers, and antioxidants as phenolic compounds located in the peripheral parts of the grains. Fermentation can be applied to treat the whole nondecorticated flour to prevent functional loss. Bringing out the nutritional benefits of millets upon fermentation will serve us to include millets at a proportion in our meals along with traditional cereals.
Fatty alcohol. Define fatty alcohols Describe the production processes of fatty alcohols and its derivatives Draw the flow chart of fatty alcohol production Explain the uses and application of fatty alcohols.
3. Definitionof Fatty Alcohols Fatty alcohols are the workhorse raw materials that facilitate the existence of products such as shampoos, shaving creams, laundry detergents, etc, and are produced at a rate of about one-and-a-half million tonnes per year and growing. Fatty alcohols are oleochemicals derived from vegetable feedstocks. The feedstock raw materials include coconut and palm kernel oils. These refined vegetable oils are first converted to a methyl ester or fatty acid. This reaction generates crude glycerine. The intermediate methyl ester or fatty acid are then fractionated and hydrogenated to produce fatty alcohol. Sources : http://www.pgchemicals.com/products/fatty-alcohols/
4. Chemical Equation for Fatty Alcohol Production Sources : http://www.pgchemicals.com/products/fatty- alcohols/
5. Block diagram of Fatty Alcohol production process
6. Fatty acids are converted into methyl ester and hydrogenated into fatty alcohols.
7. Sources : http://www.abq.org.br/workshop/11/ADRIANO- SALES-%20FIRJAM_Oleochemicals-from-Palm-Kernel- Oil.pdf
8. Hydrogenation All natural fatty alcohol processes are based on renewable fats and oils like coconut, palm oil,palm kernel,rope seed and soya bean oil. It has been proven that hydrogenation of methyl esters are preferred alternatives than hydrogenating the oils directly. Using fixed bed hydrogenation process offers the advantage of lower hydrogenation temperatures and pressures. Using special catalysts, this process is able to produce unsaturated fatty alcohols too. To produce fatty alcohols, there are three routes which is acid route,ester route and wax ester route that are shown in the following block diagrams.
9. - Acid route - Ester route - Wax ester route
10. Acid Route
Analysis of Sugars in Honey Using the PerkinElmer Altus HPLC System with RI D...PerkinElmer, Inc.
Honey consumption has grown significantly during the last few decades due to its high nutritional value and unique flavor. The price of natural bee honey is much higher than other sweeteners making it susceptible to adulteration with cheaper sweeteners, primarily sucrose. Besides lower levels of nonsugar ingredients, natural honey primarily consists of glucose and fructose and may contain low levels of sucrose and/or maltose. However, according to the international regulations, any commercially available “pure”-labeled honey products that are found to have in excess of 5% by weight of sucrose or maltose are considered to be adulterated. With the focus on possible honey adulteration, this application highlights the LC separation of various sugars found in honey and the analysis of these components in four store-bought honey samples. Method conditions and performance data, including linearity and repeatability, are presented.
Study on Fructooligosaccharide (Fos) Production By Enzyme Pectinex Ultra Sp-l...IJAEMSJORNAL
Fructooligosaccharide (FOS) is a new alternative sweetener with its characteristics such as low energy and safety for people with diabetes. Pectinex Ultra SP-L which has fructosyltransferase, catalyzes the reaction to produce short chain fructooligosaccharides. The research was conducted to enhance the high FOS content in process of FOS production by immobilized enzyme. Results achieved by Empirical planning Design Expert 7.0 - central composite method (CCD) identified at optimum conditions for process of immobilized enzyme with alginate 3.3 (%), ratio of enzyme: alginate was 0.79 (w/w), CaCl2 3.75 (%). Efficiency of loading protein reached 73.32 (%). Reaction conditions: 60 (oC), shaking velocity 90 (rpm), the initial sucrose concentrations of 50 (%), pH 5.75. When produced in 20 (h), the reaction obtained the highest level of FOS with column reaction system. The FOS for producted by immobilized enzyme achieved 47.87 (%), of which 1-kestose obtained 37.06 (%), the remaining was 10.81 (%) including nystose and fructofuranosylnystose. Efficiency of producing FOS by using immobilized enzyme compared to the free enzyme was high up to 89.49 (%).
FATS AND OILS- Fats and oils are triglycerides(triesters).They are made from glycerol and fatty acids.Fats are solids at room temperature whereas oils are liquids.Fatty acids present in fats and oils can be saturated or unsaturated.Fats and oils store energy and help to insulate the body, cushion and protect organs.
SYNTHETIC DETERGENTS- Synthetic detergents or soapless soaps are synthetic substances that are being increasingly employed as cleansing agents these days.
The objective of the current investigation is to formulate sustained release microspheres, containing Metformin hydrochloride and Glipizide as model drugs. Eudragit RSPO, Eudragit RLPO, Ethyl cellulose and Hydroxy propyl methyl cellulose, polymers of different permeability characteristics were used in combination to prepare different microspheres. Metformin and Glipizide both are type II antidiabetic agents when administered together shows synergetic effect in their action. Microspheres were prepared by emulsion solvent evaporation method with different stabilizer concentration and at different speeds of emulsification while maintaining constant amounts of Metformin and Glipizide. Drug excipients compatibility study was performed prior to formulation development and only compatible excipients were used in the fabrication of microspheres. Prepared microsphere formulations were characterized by percentage yield, particle size analysis, entrapment efficiency, in-vitro release behavior, FTIR, differential scanning colorimetry (DSC) and scanning electron microscopy (SEM). SEM studies showed that the microspheres were spherical with rough surface morphology. The drug loaded microspheres showed 29-90% entrapment capacity for Metformin and Glipizide. The in-vitro release profile showed a slow and steady release pattern for both Metformin and Glipizide. A 100% Metformin was releases within a period of 12 hrs while only 30% Glipizide was released during this time. The Metformin HCl release was found to be best fitted with Higuchi and Zero order and for Glipizide best fitted with zero order and first order respectively in single and combined polymeric loaded microspheres. DSC results indicated that the physical state of the drug was changed upon fabrication. As a result of these experiments, it was conclude that, novel sustained release oral microspheres comprising a combination of Metformin and Glipizide were successfully prepared using ethyl cellulose, HPMC 15CPS, Eudragit RSPO and Eudragit RLPO as the polymer and using emulsion solvent evaporation technique.
Abstract: While designing a cellular network, the main issue for the network planning is to achieve maximum
capacity while maintaining an acceptable grade of service and good speech quality. Planning an immature
network does not allow future growth and expansion. Wise & calculative re-use of site location in the future
network structure will save money for the operator. For this reason, digital maps are one of the most essential
elements to the network engineers while they have to think about expanding their business. However, the digital
maps cost a lot of money. This problem can be mitigated if Google Earth is used.
In this paper, the procedure of how to design a cellular digitized map on Google Earth is shown. By
calculating the cell radius, implementing the single cell site, forming the 7-cell cluster and all the cells a low
cost digitized map is designed. It is necessary to have a digitized map in mobile communication because
ultimate goal includes efficient usage of RF wave, frequency reuse, total use of BW and last but not the least
cost reduction.
Keywords: Cellular digitized map, Cell radius, Google Earth.
Content Curation : Open Educational Resources to Support Flexible Learning fo...Sarah Parker
This presentation was created as a summary of the library's contribution in its partnership developing a content curation process for OER resources for the UBC Biology department.
(download the presentation to see the notes for each slide)
Isolation and Purification of Secoisolariciresinoldiglucoside oligomers (Lign...IOSR Journals
The present study aimed to extract and purify the compound of Secoisolariciresinoldiglucoside
oligomers (lignan) from flax seed (Linumusitatissimum) and its antioxidant activity. The Lignan was extracted
by solvents which gave the best results were ethanol : 1,4 dioxane (1:1, v:v).SDG release after alkaline
hydrolysisby using a methanolicNaOH , 20 mM, pH=8 at 50 ºC.followed by using following chromatographic
techniques: Liquid-liquid, Sephadex LH-20 column chromatography, thin layer chromatographic (TLC), high
performance liquid chromatographic (HPLC) and Fourier Transform Infra-Red(FTIR) . The EC50 values of
Pure lignan extract (9 μg/ml) was shown possess DPPH radical scavenging activity compared to reference
substances BHT and vitamin C (EC50= 3 and 4.2 μg/ml) respectively, and this was higher than partial pure
lignan component (EC50= 25.5 μg/ml).The total phenolic content of the pure lignanwas higher than partial
pure lignan which gave 22.312 and 14.85 g/ml respectively.
Lipid profiling and corresponding biodiesel quality of mortierella isabellina...zhenhua82
Four lipid extraction methods (Bligh & Dyer, hexane & isopropanol, dichloromethane & methanol, and hexane) were evaluated to extract lipid from freeze- and oven-dried fungus Mortierella isabellina ATCC42613. The highest lipid yield (41.8%) was obtained from Bligh & Dyer extraction on the oven-dried fungal biomass with a methanol:chloroform:water ratio of 2:1:0.8. Other lipid extraction methods on both freeze- and oven-dried samples had lipid yields ranging from 20.7% to 35.9%. Non-polar lipid was the main lipid class (more than 90% of total lipid) in M. isabellina. Regarding fatty acid profile, there was no significant difference on fatty acid concentration between different drying and extraction methods. Estimation of biodiesel fuel properties using correlative models further demonstrated that the fungal biodiesel is a good alternative to fossil diesel.
Development and method validation for determination of Deltamethrin residue i...IOSR Journals
Olive oil is the most important commodities produced in the Mediterranean region. Due to its significant economical importance, the usage of pesticides in its production is systematic, by using a wide range of plant protection products with a variety of modes of action. As a consequence, monitoring of their residue levels in these products is a necessity. In the present study a reversed-phase high performance liquid chromatography method, with a short sample preparation step, based on acetonitrile extraction is developed and validated in olive oil, with a large scope that includes Deltamethrin as pesticide. Good sensitivity and selectivity of the method were obtained with limits of quantification at 0.2 mg kg-1. Deltamethrin has recovery rate which is of about 80℅. We confirm also the efficiency of alumina, used as adsorbent in the clean up step, to remove triglycerides and to get a pure extract. The agronomic implementation of this protocol allows us to determine the influence of some parameters on the dose and the period of treatment affecting the detected quantities of Deltamethrin residues in the produced olive oil. Indeed, we prove that the treatment dose should be specific for each case considering the olive variety, the geography of the orchard, and the predicted harvest time to determine the convenient dose of treatment. In addition, the results show that the preventive treatment at the blooming phase, does not lead to the concentration of Deltamethrin residues in the oil as it happens at the lipogenesis phase.
Congenital Agenesis Of The Corpus Callosum With Intracerebral Lipoma And Fron...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
“Hemodynamic and recovery profile with Dexmedetomidine and Fentanyl in intrac...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Correlation of Estrogen and Progesterone Receptor expression in Breast Canceriosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Analytical Study of Urine Samples for Epidemiology of Urinary Tract Infection...iosrphr_editor
The current study was carried out in District Abbottabad aimed to determine the common urinary
tract infections in local community to determine the epidemiology of significant diseases in asymptomatic patients
of renal disorder. In this study a total of 1000 urine samples were examined during 3rd February to 1st April 2015
from patients attending Ayub Teaching Hospital Abbottabad by using dipstick and microscopic analysis of urine.
There were 638 females and 362 males patients examined during this period. The range of age groups is between
1.5 years to 80 years. Results of this study was reported as Pyuria 11%, Proteinuria 21.1%, Hematuria 10.4%,
Epithelial Cells 8.2%, pH 7.8 %, Granular casts 7.3%, Triple phosphate 6.6%, Calcium oxalate 6.4%, Glycosuria
6.3%, Bacteria 6.2% and mucous 4.1%. This study concludes that routing urinalysis should be performed for all
individuals to diagnose the asymptomatic diseases that will help in simple therapeutic measurements as urinalysis
is a simple step to determine the root of Urinary tract disorders.
Chest sonography images in neonatal r.d.s. And proposed gradingiosrphr_editor
BACKGROUND : Lung sonography has been used to monitor the patients of R.D.S. in
N.I.C.U. in recent times.
AIMS : To Describe and Grade the changes of R.D.S. by lung sonography.
SETTING & DESIGN : Tertiary care institutional set up in a rural medical college.
STUDY DURATION : September 2014 to May 2015. Follow-up variable, upto 2 weeks.
PROSPECTIVE, ANALYTICAL STUDY.
MATERIALS AND METHODS -This was a single institute study approved by the institutional ethics
committee. Prior informed consent was obtained from the parents. 100 consecutive patients admitted in
N.I.C.U. WITH gestational age < 36 weeks with respiratory complaints were enrolled. Chest x-ray was
obtained within few hours of admission and lung sonography was performed within 24 hours. Follow – up
sonography was performed as and when necessary. Sonography image was graded and correlated with chest
xray and clinical picture
The Comprehensive Review on Fat Soluble Vitaminsiosrphr_editor
This review article deals with brief description of fat soluble vitamins with figures and tables
showing statistical analytical data duly quoting the references wherever necessary. The word “soluble” actually
means “able to be dissolved.” Whether a vitamin is classified as 'fat-soluble' or 'water-soluble' has to do with
how the vitamin is absorbed, stored and removed from the body. Vitamins are tiny organic compounds with a
huge impact on the health and well-being of the body. The body needs a small amount of fat soluble vitamins in
order to stay in optimal health. Fat soluble vitamins play an important role in keeping the body healthy and
functioning from immune system and muscle and heart function, easy flow and clotting of blood as well as eye
health. They are critical to health and wellness–particularly reproductive health and wellness. Low-fat, no-fat
and vegan diets are woefully lacking in fat soluble vitamins. However a diet based on traditional foods can
naturally provide these vitamins. Science is still learning about many of the functions of vitamins. "Too much
vitamin A, D, or K can lead to increased levels that are unhealthy and can cause serious health consequences.
Diseased conditions leading to decreased fat absorption leads to decreased absorption of vitamins. The fatsoluble
vitamins work most safely and effectively when obtained them from natural foods within the context of a
diet rich in all their synergistic partners. If fat soluble vitamins are stored for lengthy time they generate threat
for toxicity than water soluble vitamins and such situation even aggravated, provided they are consumed in
excess. Vitamin products, above the legal limits are not considered food supplements and must be registered as
prescription or non-prescription (over-the-counter drugs) due to their potential side effects. Vitamin A and E
supplements do not provide health benefits for healthy individuals, instead they may enhance mortality, and it is
held proved that beta-carotene supplements can be harmful to smokers
Sulphasalazine Induced Toxic Epidermal Necrolysis A Case Reportiosrphr_editor
Toxic Epidermal Necrolysis (TEN) is a rare and life threatening mucocutaneous reaction
characterized by extensive necrosis and detachment of epidermis. The Worldwide incidence of TEN is 0.9 to 1.4
per million populations per year [1]. Here we have discussed a case of Toxic Epidermal Necrolysis secondary
to Sulfasalazine managed with fluid replacement, analgesics, anti-infective therapy aggressive nutritional
support and intravenous high dose steroid therapy.
Keywords- Toxic Epidermal Necrolysis, Sulfasalazine
Evaluation the efficacy of IVIgG in treatment of Hemolytic Disease of Newborniosrphr_editor
Hemolytic disease of newborn (HDN) is an important cause of hyperbilirubinemia in the
neonatal period,and delayed diagnosis and treatment may lead to permanent brain damage. Traditional
neonatal treatment of HDN is intensive phototherapy and exchange transfusion.Intravenous
immunoglobulin(IVIgG) has been introduced as an alternative therapy to exchange transfusion. This study was
conducted to assess the effect of IVIG in HDN .
FIBROLIPOMATOUS HAMARTOMA OF ULNAR NERVE: A RARE CASE REPORT.iosrphr_editor
Nervous fibrolipomatous hamartoma is said to be a rare tumor-like condition involving the peripheral
nerves,in which the epineurium and perineurium are enlarged and distorted by excess of fatty and fibrous tissue
s that infiltrate between and around nerve boundaries. The median nerve is more likely to develop a hamartoma
than other nerves with a predilection for the carpal tunnel.
A fibrolipomatous hamartoma – is a rare, benign, congenital lesion most commonly found in the median nerve,
usually at the level of the wrist or hand.
We report a case of this rare condition in ulnar nerve.
SELF MEDICATION PRACTICES FOR ORAL HEALTH PROBLEMS AMONG DENTAL PATIENTS IN B...iosrphr_editor
Introduction: Self‑ medication is commonly practiced all over the world. Self-medication is defined as the use
of medication by a patient on his own initiative or on the advice of a pharmacist or a lay person instead of
consulting a medical practitioner. The present study was aimed to estimate the prevalence of self-medication for
oral health problems among dental patients in Bengaluru city; to identify triggering factors that could influence
self-medication practices; to identify sources of medications used; to identify sources of information about
medications used; and to identify reasons for self-medication.Study Design: A Cross sectional Study.Methods:A
survey was conducted among 175 subjects among dental patients in Bengaluru city. Data were collected
through a specially designed proforma using a closed‑ ended, self‑ administered questionnaire containing 15
questions, in five sections.
Results: The prevalence of
Clinico-haematological Profile of Falciparum Malaria in a Rural Hospital of T...iosrphr_editor
Aim: To study the clinico-haematological profile malaria in a rural hospital of Tripura.
Material and methods: A cross-sectional hospital-based study was done from at Kulai District
Hospital,Tripura. This hospital based cross sectional study was done on 60 confirmed cases of falciparum
malaria (either by peripheral smear or rapid diagnostic test) admitted in Kulai District Hospital. A case sheet
proforma was prepared and data (demographic profile,clinical feature, investigation, treatment, and
complication) from all indoor patients was collected and analyzed.
Result: Out of 60 patients, 40(66.6%) were males and 20 (33.4%) were females. Most of the patients were
between the age group 21-40 years with the highest prevalence between the age group of 21-30. Fever was the
most common symptom. Anemia was present in 42(70%) patients, out of which 6(10%) patients had severe
anemia. Thrombocytopenia was present in 36(60%) patients.Abnormal liver function tests were observed in
26(43.3%) subjects while abnormal kidney function tests were observed in16(26.6%) patients. All the 60
patients received Artemisinin based antimalarial drugs.
Conclusion: Early detection, prompt management, and adequate supportive therapy may reduce mortality due
to falciparum cerebral malaria.
Indonesian Wild Ginger (Zingiber sp) Extract: Antibacterial Activity against ...iosrphr_editor
Lempuyang gajah (Zingiber zerumbet (L.) Smith), lempuyang pahit (Zingiber amaricans BL.), and
lempuyang wangi (Zingiber aromaticum Vahl.) are used as traditional medicine (jamu) in Indonesia. It is also
used for treatment of microbial infections, helps to increase appetite and stimulate digestion in chickens.
Information on their uses are available, but only limited in the scientific data on their bioactivity. The study was
conducted on the antibacterial effect of organic extracts of these plants with Mycoplasma gallisepticum as the
agent of chronic respiratory disease in chickens. Juice and extracts of fresh and dried rhizome are evaluated
through the disc diffusion assay and minimum inhibitory concentration. Oxytetracyclin (30 µg) are used as
standards. All extracts are individually exhibited as antibacterial activity against Mycoplasma gallisepticum (7
± 0.11 mm to 21 ± 0.86 mm). The minimum inhibitory concentration (MIC) determination of plants extracts are
ranged from 7.8 mg/ml to 31.2 mg/ml. The preliminary results suggested promising antibacterial properties of
wild ginger from Indonesia, and probably could be used in management of chronic respiratory disease in
chickens.
A case of allergy and food sensitivity: the nasunin, natural color of eggplantiosrphr_editor
Abstract: Allergies and food sensitivities can both be considered as "adverse reactions individualistic" to food.
Are pathological and individual forms because they affect a few individuals in way rather serious; immediate
or delayed reactions occur instead with simple effects histamine, or, in severe cases with respiratory and
anaphylactic shock
The eggplant (Solanum melongena L.) is known to cause food allergies in some Asian countries, but detailed
studies on allergies caused by eggplant are lacking, however, it was highlighted the presence of allergens in
edible parts of eggplant with preponderance in the peel .
The purpose of this study was to propose an extraction method rapid, efficient and cost of natural dye from
waste products from the food industry, such as the peels of eggplant, from which it was extracted, isolated and
purified the nasunin,a colored molecule in red-fuchsia.
Nasusin was tested on 58 patients to evaluate the potential sensitizing effect on the skin. The results demonstrate
that allergenic effects are negligible and therefore the nasunin can be used as a colorant in various industrial
sectors with a certain safety margin
Complete NMR Assignment of MogrosidesII A2, II E andIII A1Isolated from Luo H...iosrphr_editor
NMR analysis allowed complete assignments of three known mogrol glycosides, Mogroside IIA2 (1),
II E (2)and IIIA1 (3), isolated from the extracts of Luo Han Guo. Herein, complete 1H and 13C NMR
assignmentsof all threemogrosidesare described based on NMR experiments (1H NMR, 13C NMR, COSY,
HSQC-DEPT, HMBC, NOESY and 1DTOCSY) and mass spectral data.
Nanoemulsion and Nanoemulgel as a Topical Formulationiosrphr_editor
: Nanoemulsion is referred type of emulsion with uniform and extremely small droplet size in the range
of 20-200 nm. Nanoemulsion provides numerous advantages over other carrier such as polymeric nanoparticle
and liposomes, including low cost preparation procedure, high hydrophilic and lipophilic drug loading system
to enhance the longer shelf live upon preserving the therapeutic agents. Incorporating the preparation of
nanoemulsion with hydrogel matrix to produce nanoemulgel exhibited by the two separate systems that forming
it. Nanoemulgel possesses the properties of thixotropic, non-greasy, effortlessly spreadable, easily be removed,
emollient, not staining, soluble in water, longer shelf life, bio-friendly, translucent and agreeable appearance.
Pharmacokinetics of High-Dose Methotrexate in Egyptian Children with Acute Ly...iosrphr_editor
Aim:Since several factors have been shown to influence the clearance of methotrexate, the purpose of this study
was to identify potential relationships between patient covariates and the methotrexate clearance estimates and
deduce a pharmacokinetic model for the estimation of methotrexate clearance in Egyptian pediatric ALL
patients that may help dosage adjustment and achieve target steady-state plasma concentrations in a similar
sittings.
Patients and methods: A total of 94 pediatric patients with B-cell ALL, of whom 70 were the studied population
and 24 were the test population, were treated with four courses of HDMTX doses 2.5 gm/m2
(low-risk arm) or 5
gm/m2
(standard-/high-risk arm) given every other week by intermittent intravenous infusions over 24 hours as
a part of their treatment protocol. Patients were monitored for the 24 hour MTX concentration and the systemic
methotrexate clearance was calculated for each methotrexate dose
Epidemiology of Tuberculosis (TB) in Albania 1998-2009iosrphr_editor
Abstract : In Albania, many people erroneously think that tuberculosis (TB) is a disease of the past-an illness
that no longer constitutes a public health threat. Surveillance is an integral part of tuberculosis (TB) control.
Albania has a highTB notification rate and there are doubts about underreporting. The evolution of the
incidence of tuberculosis is presented, together with more detailed figures over the period 1998-2009. These
figures were obtained by the monthly forms (called 14/Sh) compared with the individual notification data.
Objective: To examine the distribution and sources of increased tuberculosis (TB) morbidity and reporting
system deficiencies in the Albania from 1998 through 2009. Metodology: The study is descriptive one conductet
during the period 1998-2009. The statistical analysis is based on data reported from regional level (regional
epidemiological departments) to the central level (Public Health Institute). Results: The main findings were:
discordance between the collected data (individual form) and reported data (monthly form); tuberculosis
incidence rate shows little oscillations which ranges from 6.67 to 9.2 cases/100.000 population; 50% of the
regions show a lack of information on the confirmation of diagnosis and laboratory examination type used for
confirmation. Conclusion: TB disease in high-risk populations where it is difficult to detect, diagnose, and treat;
limitations of current control measures and the need for new tests and treatments, including an effective
vaccine; improving information system, regulation of individual form and personnel training.
Total Phenol and Antioxidant from Seed and Peel of Ripe and Unripe of Indones...iosrphr_editor
Study on total phenol and antioxidantactivity ofsugar apple fruits of various solvent, part of fruits, and level of ripening. Solvent extraction used were 80% (v/v) methanol, 50% (v/v) acetone, boiling water, and 50% (v/v) ethanol. Part of fruits thatbeen used for samples were seed and peel which are normally by products of sugar apple processing, level of ripening were unripe, and ripe sugar apple fruits. Total phenol was determined by Folin-ciocalteau method. Total antioxidant was quantified by 1,1-diphenyl-2-picrylhydrazyl(DPPH) method.Therewas a difference in type of solvent, part of fruits, and level of ripeningon total phenol and antioxidant concentration of sugar apple fruits. Seeds have higher total phenol concentration than peels of this fruits. Unripe sugar apple fruits have higher total phenol and antioxidant than ripe fruit. The best solvent for phenol extraction was ethanol 50%butthe best solvent for antioxidant extraction was acetone 50%.
A Review on Step-by-Step Analytical Method Validationiosrphr_editor
When analytical method is utilized to generate results about the characteristics of drug related samples it is essential that the results are trustworthy. They may be utilized as the basis for decisions relating to administering the drug to patients. Analytical method validation required during drug development and manufacturing and these analytical methods are fit for their intended purpose. To comply with the requirements of GMP pharmaceutical industries should have an overall validation policy which documents how validation will be performed. The purpose of this validation is to show that processes involved in the development and manufacture of drug, production and analytical testing can be performed in an effective and reproducible manner. This review article provides guidance on how to perform validation characteristics for the analytical method which are utilized in pharmaceutical analysis.
A Cross Sectional Study of Ethnic Differences in Occurrence and Severity of A...iosrphr_editor
Non-steroidal anti-inflammatory drugs are the most widely used "over the counter" medication all over the world despite their complications in different major organs. Present studies envisaged for knowing the occurrence and severity of adverse drug reactions from NSAIDs in different ethnic communities of Sikkim. A cross sectional study was undertaken in the medicine outpatients department of a secondary and tertiary care hospital. The patients belonging to Nepalese, Bhutias, Lepchas ethnic communities and others community (settlers from other parts of India) were included to analyzed the data based on the age and gender, ethnicity and ADRs, drugs and ADRs. Severity assessment was done using Hartwing and Siegel scale and causality assessment by Naranjo scale. Total 109 cases of ADRs, predominating in female were detected. Nepalese were the most affected and Gastrointestinal tract (GIT) being the most affected organ in them. Diclofenac showed maximum number of ADRs in all the communities. Maximum number of cases occurred on single day use (40.36%) of drugs. All the cases were belonging to the "possible category" and the maximum being the mild (72.48%) in nature. It is advisable to consider the ethnic/racial differences equally with other factors, to improve the safety and efficacy of a drug.
A Cross Sectional Study of Ethnic Differences in Occurrence and Severity of A...
C0532011019
1. IOSR Journal Of Pharmacy
(e)-ISSN: 2250-3013, (p)-ISSN: 2319-4219
www.iosrphr.org Volume 5, Issue 3 (March 2015), PP. 11-19
11
Validation and Application of a Gas Chromatographic Method
for the Determination of Fatty Acids and Sterols in the Total Diet
Vesna Kostik
Institute of Public Health of Republic of Macedonia, 50 Divizija No. 6, 1000 Skopje, Republic of Macedonia
ABSTRACT : Fatty acids (FAs) and sterols represent the most important fraction of edible oils and fats,
which play a significant role in human nutrition and health. A gas chromatographic method for the
identification and quantification of FAs and sterols based on the extraction of lipids by accelerated solvent
extraction (ASE), saponification of the obtained extract, and direct determination of the isolated FAs and sterols
without derivatization, was developed and validated. The proposed method was accurate and precise (mean
recovery in the range 80.8% to 98.2%, precision with RSD < 8.6%; LOQ < 0.86 mg/kg; measurement
uncertainty < 14.8%). This method was applied for the determination of the most abundant FAs and sterols in
the total diet. The obtained results showed that the most abundant FA in the total diet is monounsaturated C18:1
with 34.5% of the total FAs. The lowest presence was observed for the short chain FAs: C4:0 – C12:0 (6.4%).
Cholesterol was found to be the most abundant sterol in the diet (57.5%). The total amount of the plant sterols
in the diet was found to be 42.5%.
KEYWORDS: gas chromatography, fatty acid, sterol.
I. INTRODUCTION
Edible oils and fats are one of the major components of human diets, comprising as much as 25% of
average caloric intake. They play important functional and sensory roles in food products and they act as
carriers of fat-soluble vitamins (A, D, E, and K). They also provide an essential linoleic and linolenic acid,
responsible for growth [1]. Edible oils and fats are biological mixtures of plant origin consisting of ester
mixtures derived from glycerol with chain of fatty acids [2]. Both the physical and the chemical characteristics
of oils and fats are greatly influenced by the kind and proportion of the fatty acids (FAs) on the triacylglycerol
[3, 4]. FAs can be classified in classes as saturated, mono-unsaturated (MUFA) and polyunsaturated (PUFA)
fatty acids. The predominant fatty acids present in the diet are saturated and unsaturated compounds with
straight aliphatic chains. An even number of carbon atoms, from 16 to 18, with a single carboxyl group, is the
most common [5, 6]. High levels of saturated FAs in the diet are frequently considered to have influence in
increasing the concentration of low density lipoproteins (LDL), affecting the ratio of LDL to HDL (high density
lipoproteins), promoting clothing and vascular smooth muscle proliferation [7, 8]. Diet with increasing intake of
linoleic and linolenic acids increase HDL-cholesterol and decreases LDL-cholesterol, while higher intake of
oleic acid decreases LDL-cholesterol, but does not affect HDL cholesterol levels [9].
Therefore, the determination of FAs is an essential requirement in testing food material [10].Gas
chromatography (GC) coupled with mass spectrometer (MS) or flame ionization detector (FID) is the most
widely used technique for the determination of FAs [11-13]. Commonly, the most methodologies used for
determining FAs are lipid extraction followed by conversion of the FAs into corresponding methyl esters
(FAMEs) [14, 15]. Such methodologies are usually used for preparing FAMEs from lipids either by basic
hydrolysis followed by methylation of the free FAs (FFAs) or by trans-esterification of lipids using acid or base
catalyzed as rapid and simple methods [14,16].
The presence of non-lipids at the moment of the derivatization process may interfere with lipids
resulting to potential errors and high variable profiles [17].
Beside fatty acids, the most abundant components present in the edible oils and fats are sterols.
Cholesterol, sitosterol and stigmasterol are polycyclic steroid compounds with similar chemical structure.
Cholesterol is a typical animal sterol, e.g. its content in milk fat is 95–98%, stigmasterol and sitosterol are
referred to as phytosterols [18].
The determination of sterols in the total diet is carried out for the following objectives: to measure the
total cholesterol content to obtain nutritional information and to detect the presence of vegetable fats [18,19].
The most appropriate and frequently used method for the determination of total sterols content in foods
is the direct saponification, followed by the extraction of the unsaponificable residue into the nonpolar solvent,
derivatization of the isolated sterols and final gas chromatographic detection [20]. The aim of our study was to
develop and validate simple, reliable and accurate GC method for determination of the most abundant FAs and
2. Valdiation and application of a gas chromatographic method for the determination of fatty…
12
sterols in the total diet without derivatization of the isolated analytes. The method was applied for determination
of FAs and sterols in the total diet (28 samples).
.
II. MATERIALS AND METHODS
2.1. Instrumentation
The chromatographic analysis (GC) were performed on a Shimadzu 2010 GC system equipped with
flame-ionization detector (FID), and auto injector (AOC- 20i), and the Chrom- Solution software. The sample
extraction was performed in a DIONEX Accelerated Solvent extractor, ASE-100 (USA). Stainless steel
extraction cells (34 mL) were used for the extraction. Helium (purity 99.999 %) was used as a purge gas. The
sample saponification was performed with reflux condenser.
2.2. Reagents and standards
All chemicals and solvent were a special grade for gas chromatography. N-hexane, 96% (V/V) ethanol,
absolute ethanol and chloroform were obtained from Merck (Darmstadt, Germany). Anhydrous sodium
sulphate, (prepared 3 hours at 650 0
C), was purchased from Sigma-Aldrich/Fluka/Riedel-de-Haen (Zwijndrecht,
The Netherlands). The moisture absorbing polymer (MAP) and diatomaceous earth (DE) were purchased from
Thermo Fisher Scientific, Waltham, MA (USA). Water was deionized then distilled from glass apparatus.
Potassium hydroxide analytical grade was obtained from Merck (Darmstadt, Germany)
The analytical standards: butyric acid (C4:0) purity ≥98.5% GC, caproic acid (C6:0) purity ≥99.0% GC,
caprylic acid (C8:0) purity ≥98.5% GC, capric acid (C10:0) purity ≥99.5% GC, lauric acid (C12:0) purity ≥99.0%
GC, myristic acid (C14:0) purity ≥98.5% GC, palmitic acid (C16:0) purity ≥99.0% GC, palmitoleic acid (C16:1)
purity ≥98.5% GC, stearic acid (C18:0) purity ≥99.0% GC, oleic acid (C18:1) purity ≥98.5% GC, linoleic acid
(C18:2) purity ≥98.5%, linolenic acid (C18:3) purity ≥98.5%, cholesterol purity ≥98.5% GC, stigmasterol purity
≥98.5% GC, β- sitosterol purity ≥98.0% GC, campesterol purity ≥98.5% GC and 5-α-cholestane purity ≥98.0%
GC were obtained from Sigma-Aldrich/Fluka/Riedel-de-Haen (Zwijndrecht, The Netherlands). Blank samples
of dietary non fatty supplements used for fortification were purchased from the local market.
2.3. Preparation of the standard solutions
2.3.1. Preparation of FAs standard solutions
Stock solutions of individual fatty acid (FA) standards were prepared in n – hexane at 1000 mg/L and
were stored in a refrigerator at -20 0
C. Standard working solution mixture (i) was prepared by transferring 10
mL of each individual stock standard solution in a 50 mL volumetric flask and diluting with n – hexane to a
concentration of 200 mg/L. Stock standard solutions of individual FA and standard working solution mixture (i)
was used for the preparation of chromatographic standard solutions (ii) with different FA concentrations: 10
mg/L – 100 mg/L, i.e. 10 mg/L; 25 mg/L; 50 mg/L; 75 mg/L and 100 mg/L. Chromatographic standard
solutions were prepared in n-hexane. Stock solutions and standard working solution mixture (i) were used for
the fortification of the blank samples.
2.3.2. Preparation of sterols standard solutions
Stock solutions of individual sterols were prepared in chloroform at 1000 mg/L and stored in a
refrigerator at -200
C. Standard working solution mixture (i) was prepared by transferring 5 mL of each
individual stock standard solution in a 50 mL volumetric flask and diluting with chloroform to a concentration
of 100 mg/L. Standard working solution mixture (ii) was prepared by transferring 5 mL of standard working
solution mixture (i) to a 50 mL volumetric flask and diluting with chloroform to a concentration of 10 mg/L.
Standard working solution mixture (i) was used for the preparation of the chromatographic standard solutions
with different stigmasterol and campesterol concentrations: 1 mg/L – 10 mg/L, i.e. 1.0 mg/L; 2.5 mg/L; 5 mg/L;
7.5 mg/L and 10 mg/L. Stock solutions of cholesterol and β sitosterol were used for the preparation of the
chromatographic standard solutions with cholesterol and β sitosterol concentrations ranging from 10 mg/L to
100 mg/L, i.e. 10 mg/L; 25 mg/L; 50 mg/L; 75 mg/L and 100 mg/L. Chromatographic standard solutions were
prepared in chloroform. Stock solutions and standard working solution mixture (i) and mixture (ii) were used for
the fortification of blank samples. Internal standard stock solution of 5-α-cholestane was prepared in chloroform
at a 1000 mg/L. Working internal standard solution was prepared by transferring 100 μL of internal standard
stock solution (IS) in a 10 mL volumetric flask and diluting with chloroform to a concentration of 10 mg/L. 1
mL of internal standard solution (10 mg/L) was added into the sample prior the extraction.
3. Valdiation and application of a gas chromatographic method for the determination of fatty…
13
2.4. Sample preparation
An aliquot of 10 g of homogenized sample was mixed with 2 g of MAP and 2 g of DE. The extraction
cell was filled with the homogenized mixture, and placed in the ASE, which worked out under the conditions
shown in Table 1.
Table 1. ASE operating conditions
Solvent n-hexane
Temperature 100 0
C
Pressure 1500 psi
Static time 5 min
Flush volume 60 %
Purge time 140 sec
Static Cycle 1
The obtained extract (15 mL) was evaporated under the stream of nitrogen to dryness. The residue was
dissolved in 30 mL solution of KOH in absolute ethanol (0.5 mol/L). The saponification was performed using
reflux condenser within 30 min. The solution obtained after the saponification was quantitatively transferred
into a 50 mL volumetric flask and filled with absolute ethanol up to the mark. The solution was used for the
further analysis.
2.4.1. Extraction of FAs
5 mL of the solution was transferred to a 100 mL separatory funnel. 0.1 mL 25% (V/V) HCl was added
together with 25 mL of n-hexane and 25 mL of distilled water. The mixture was vigorously shaken for 5 min and
then allowed to stand for 15 min. The water portion was discarded. The hexane layer was collected through the
anhydrous sodium sulphate and removed by a rotary evaporator at 40 0
C. The residue was dissolved in n –
hexane and adjusted to the volume of 10 mL with the same solvent. The sample solution was used for GC-FID
determination.
2.4.2. Extraction of sterols
45 mL of the solution was transferred to a 250 mL separatory funnel together with 100 mL of n-hexane
and 50 mL of mixture (96%, V/V) ethanol: distilled water (1:1). The mixture was vigorously shaken for 5 min
and then allowed to stand for 15 min. The water portion was discarded. The hexane layer was collected through
the anhydrous sodium sulphate and removed by a rotary evaporator at 40 0
C. The residue was dissolved in
chloroform and adjusted to the volume of 1 mL with the same solvent. The sample solution was used for GC-
FID determination.
2.5. GC determination
Separation of FAs was achieved on a fused silica HP-FFAF capillary column (25 m x 0.32 mm i.d. x
0.52 μm film thickness), supplied by Agilent (USA). Operating conditions were as follows: injector port
temperature, 230 0
C; injection volume, 2 μL in split modе (1:10); detector temperature, 260 0
C (make up gas -
nitrogen flow 27.5 mL/min); nitrogen as carrier gas at flow rate at 1.5 mL0
C /min; oven temperature
programme, 180 0
C (1 min) increased with the rate of 2 0
C/min to 200 0
C and held for 19 min. The total run time
of the chromatographic analysis was 30 min. The column equilibration time was 3 min.
Sterols separation was achieved on a fused silica HP-FFAF capillary column (25 m x 0.32 mm i.d. x
0.52 μm film thickness), supplied by Agilent (USA). Operating conditions were as follows: injector port
temperature, 230 0
C; injection volume, 2 μL in split mode (1:3); detector temperature, 300 0
C (make up gas -
nitrogen flow 27.5 mL/min); nitrogen as carrier gas at flow rate at 1.5 mL0
C /min; oven temperature was set at
260 0
C and held for 40 min. The column equilibration time was 3 min.
The proposed method was validated in respect to the recovery, linearity, precision expressed as within
day repeatability and between day reproducibility of retention time and peak area, stability, limit of detection
(LOQ), limit of quantification (LOQ) and measurement uncertainty (Ux).
III. RESULTS AND DISCUSSION
3.1. Method optimization
In order to obtain the best separation for all tested FAs and sterols, a series of preliminary
investigations with capillary GC columns with a different polarity of the stationary phase were tested. The
optimum separation of components of interest was achieved when capillary column with a strong polarity
(Polyethylene glycol-TPA modified) and optimized temperature programmes are used (Fig. 1).
4. Valdiation and application of a gas chromatographic method for the determination of fatty…
14
Representative chromatograms for FAs and sterols separation with GC – FID method are shown in Fig.
1 and Fig. 2, respectively.
Figure 1. Chromatogram of standard solution of C16:0 (1), C18:0 (2), C18:1 (3), C18:2 (4) and C18:3 (5), at 10 mg/L
Figure 2. Chromatogram of the sterols extract from total diet sample [1. 5 α-Cholestan (I.S.); 2. Cholesterol; 3.
Stigmasterol; 4. Campesterol; 5. β - Sitosterol]
In our research accelerated solvent extraction (ASE) technique was used for samples extraction. The
elevated pressure increases the boiling temperature of the solvent and extraction can occur at relatively high
temperatures, which leads to faster extractions [21, 22]. The extraction process is therefore significantly faster
than traditional methods such as Soxhlet extraction or supercritical fluid extraction [23]. In a study conducted by
Omar and Salimon [20] the total time required for extraction of the lipids from the sample with Soxhlet
extraction was approximately 3 hours. Soxhlet extraction and SFE are also used as techniques for the isolation
of lipids prior to the determination of sterols [24]. In our investigations due to the use of elevated temperature
and pressure in the extraction cell the extraction time was approximately 15 min.
Up to our knowledge, no previous data was reported in the literature for direct determination of FAs
and sterols by GC. In our study, for the first time we introduce an analytical method for direct determination of
free FAs and sterols, avoiding the derivatization step and therefore the possible errors which could be due to this
demanding procedure.
3.2. Method Validation
3.2.1. Recovery, Precision and Linearity
The recovery, precision and linearity of all the FAs and sterols was determined using fortified samples
of dietary non fatty supplements which were previously tested on the presence of FAs and sterols. In each case,
5 replicates each at 5 levels were fortified into the samples. The samples (10 g) were fortified before the
extraction to yield 10 mg/kg; 25 mg/kg; 50 mg/kg; 75 mg/kg and 100 mg/kg, respectively (C4:0, C6:0, C8:0, C10:0,
C16:0, C16:1, C18:0, C18:1, C18:2, C18:3, cholesterol and β - Sitosterol); and to yield 1.0 mg/kg; 2.5 mg/kg; 5 mg/kg; 7.5
5. Valdiation and application of a gas chromatographic method for the determination of fatty…
15
mg/kg and 10 mg/kg, respectively (stigmasterol and campesterol). Gas tight glass syringes (10 µL, 50 µL and
100 µL) were use for the addition of spiking mixtures.
The recovery, precision and linearity of the tested method for the determination of FAs and sterols are
presented in Table 2 and Table 3, respectively.
From the obtained results, it can be noticed that the proposed method is accurate and precise enough for
the determination of all the tested FAs and sterols. The obtained values for multiple correlation coefficients,
ranged from 0.9904 to 0.9982, indicated that the method has a good linearity for all the tested compounds. High
analytical recoveries ranging from 80.8% to 98.2% were obtained for FAs determination, as well as, for sterols
determination (82.7% - 91.0%).
Table 2. Statistical data for mean recovery, precision data and linearity of the method for the
determination of FAs
FA
Fortification level
(mg/kg)
FA found - mean value
(mg/kg ± SD)
Recovery (%), n=5 RSD (%) Regression equation
C4:0
10 8.98 ± 0.55 89.8 6.2
y = 0.9912x – 2.2561
R2
= 0.9987
25 22.4 ± 1.2 86.2 5.3
50 45.6 ± 2.4 91.1 5.2
75 71.3 ± 4.0 95.1 5.6
100 98.2 ± 4.7 98.2 4.8
C6:0
10 9.12 0.23 91.2 2.5
y = 0.9832x – 1.6415
R2
= 0.9982
25 23.5 ± 1.05 94.0 4.5
50 44.8 ± 2.5 89.6 5.6
75 72.5 ± 3.4 96.7 4.7
100 97.5 ± 4.2 97.5 4.3
C8:0
10 8.79 ± 0.61 87.9 6.9
y = 0.969x – 2.7665
R2
= 0.9954
25 21.8 ± 1.2 87.2 5.5
50 41.5 ± 2.1 83.0 5.1
75 70.5 ± 2.3 94.0 3.3
100 95.5 ± 4.4 95.5 4.6
C10:0
10 8.65 ± 0.54 86.5 6.2
y = 0.9525x – 3.0378
R2
= 0.9951
25 20.5 ± 1.1 82.0 5.4
50 40.5 ± 1.9 81.0 4.7
75 69.6 ± 2.9 92.8 4.2
100 93.2 ± 5.2 93.2 5.6
C12:0
10 8.3 ± 0.40 83.0 4.8
y = 0.9136x – 1.4293
R2
= 0.9983
25 21.4 ± 1.4 85.6 6.8
50 42.3 ± 2.5 84.6 5.9
75 68.9 ± 3.1 91.9 4.5
100 89.5 ± 5.9 89.5 6.6
C14:0
10 8.95 ± 0.54 89.5 6.8
y = 0.9089x –1.9915
R2
= 0.9982
25 20.5 ± 1.6 82.3 7.8
50 41.4 ± 2.8 82.8 6.7
75 67.9 ± 3.6 90.5 5.3
100 88.6 ± 5.5 88.6 6.2
C16:0
10 8.15 ± 0.6 81.5 7.4
y = 0.9273x –1.8305
R2
= 0.9987
25 20.8 ± 1.8 83.2 8.6
50 43.2 ± 2.5 86.4 5.8
75 69.5 ± 3.6 92.7 5.2
100 90.3 ± 5.2 90.3 5.7
C16:1
10 8.8 ± 0.43 88.0 5.5
y = 0.9411x –2.4756
R2
= 0.9956
25 22.3 ± 1.3 89.2 5.8
50 40.5 ± 2.1 81.6 5.2
75 69.4 ± 2.9 92.5 4.2
100 92.3 ± 4.8 92.3 5.2
C18:0
10 8.6 ± 0.45 86.0 5.9
y = 0.952x –2.5634
R2
= 0.9926
25 23.4 ± 1.6 93.6 6.8
50 39.8 ± 2.5 89.6 6.3
75 70.5 ± 3.2 94.0 4.5
100 93.4 ± 5.1 93.4 5.5
C18:1
10 8.3 ± 0.40 83.0 5.4
y = 0.9464x – 3.3537
R2
= 0.9906
25 20.2 ± 1.4 80.8 6.9
50 39.6 ± 2.8 89.2 7.1
75 72.4 ± 3.0 96.5 4.1
100 89.8 ± 4.9 89.8 5.5
C18:2
10 8.1 ± 0.50 81.0 7.0
y = 0.9297x – 3.2634
R2
= 0.9912
25 20.5 ± 1.3 82.0 6.3
50 38.5 ± 2.1 82.5 5.4
6. Valdiation and application of a gas chromatographic method for the determination of fatty…
16
FA
Fortification level
(mg/kg)
FA found - mean value
(mg/kg ± SD)
Recovery (%), n=5 RSD (%) Regression equation
75 70.5 ± 3.2 94.0 4.5
100 88.8 ± 4.7 88.8 5.3
C18:3
10 8.34 ± 0.30 83.4 4.3
y = 0.9192x – 3.5561
R2
= 0.9904
25 19.8 ± 1.2 84.3 6.1
50 37.5 ± 2.4 85.0 6.4
75 69.5 ± 2.9 92.7 4.2
100 87.5 ± 3.6 87.5 4.1
Table 3. Statistical data for mean recovery, precision data and linearity of the method for the determination of
sterols
FA
Fortification level
(mg/kg)
FA found - mean value
(mg/kg ± SD)
Recovery (%),
n=5
RSD (%) Regression equation
Cholesterol
10 8.3 ± 0.42 83.0 6.3
y = 0.8744x – 3.9683
R2
= 0.9953
25 18.6 ± 1.6 84.4 8.6
50 36.5 ± 2.7 83.0 7.4
75 60.5 ± 3.6 82.9 5.9
100 85.6 ± 4.5 85.6 5.2
β - Sitosterol
10 8.93 ± 0.30 89.3 4.3
y = 0.8421x – 2.4683
R2
= 0.9975
25 19.2 ± 1.4 86.8 7.3
50 37.6 ± 2.4 87.5 6.4
75 59.5 ± 3.4 86.3 5.7
100 83.4 ± 4.7 83.4 5.6
Campesterol
1.0 0.863 ± 0.03 86.3 4.3
y = 0.9274x – 0.3024
R2
= 0.9929
2.5 2.1 ± 0.15 84.5 7.1
5.0 3.9 ± 0.20 85.0 5.1
7.5 7.0 ± 0.30 83.7 4.3
10 8.9 ± 0.40 89.0 4.5
Stigmasterol
1.0 0.837± 0.04 83.7 5.5
y = 0.943x – 0.3378
R2
= 0.9975
2.5 2.0 ± 0.15 85.4 7.5
5.0 4.1 ± 0.22 83.6 5.4
7.5 6.9 ± 0.35 92.0 5.1
10 9.1 ± 0.30 91.0 3.3
3.2.2. Repeatability and Reproducibility
The within day repeatability of our method was determined by performing the analysis of 10 blank
samples fortified with FAs at 50 mg/g and sterols at 10 mg/kg, respectively. After the isolation of the lipid
fraction, saponification of the extracts and extraction of FAs and sterols, obtained extracts were analyzed by GC-
FID within the same day under the chromatographic conditions described in the section 2.5. The between day
reproducibility of the method was determined by performing the analysis of the extracts obtained from 5
fortified blank sample (50 mg/g of FAs, 10 mg/L of sterols). After the isolation of the lipid fraction,
saponification of the extracts and extraction of FAs and sterols, obtained extracts were analyzed by GC-FID
within the same day under the chromatographic conditions described in the section 2.5. The calculated RSD
values for the retention time (tR), the peak areas, within day repeatability and between day reproducibility are
shown in Table 4 and Table 5, respectively.
The calculated RSD values for within day repeatability in the case of FAs determination for tR values
ranged from 0.112% to 0.190%, whereas for peak areas RSD values ranged from 3.03% to 4.61%, indicating
good precision of the tR, as well as of the peak area, within the same day. Good precision of the tR, (RSD from
0.165% to 0.196%) and the peak area (RSD from 3.23% to 4.95%) were also observed within different days
(between day reproducibility).
The calculated RSD values for within day repeatability in the case of sterols determination for tR values
ranged from 0.182% to 0.192%, whereas for peak areas RSD values ranged from 3.82% to 4.27%, indicating
good precision of the tR, as well as of the peak area, within the same day. Good precision of the tR, (RSD from
0.196% to 0.205%) and the peak area (RSD from 4.11% to 4.94%) were also observed within different days
(between day reproducibility).
7. Valdiation and application of a gas chromatographic method for the determination of fatty…
17
Table 4. Statistical data for within day repeatability and between day reproducibility for the determination of
FAs
FA tR /min
Within day Repeatability (RSD,
%); n=10
Between day Reproducibility (RSD,
%); n=25
C4:0 4.872 0.112 4.17 0.165 4.11
C6:0 6.782 0.162 3.91 0.179 3.23
C8:0 8.450 0.190 3.40 0.198 3.78
C10:0 9.454 0.110 3.73 0.184 3.94
C12:0 12.345 0.121 4.61 0.194 4.97
C14:0 14.190 0.103 3.23 0.172 4.56
C16:0 15.104 0.105 3.52 0.198 4.46
C16:1 21.146 0.163 3.29 0.189 4.95
C18:0 21.890 0.142 3.79 0.177 4.65
C18:1 22.267 0.164 3.85 0.180 4.90
C18:2 23.467 0.161 3.21 0.192 4.34
C18:3 25.796 0.176 3.03 0.196 4.95
Table 5. Statistical data for within day repeatability and between day reproducibility for the determination of
sterols
FA tR /min
Within day Repeatability (RSD,
%); n=10
Between day Reproducibility (RSD,
%); n=25
Cholesterol 15.163 0.192 4.27 0.205 4.11
Stigmasterol 18.893 0.182 3.97 0.219 4.23
Campesterol 20.983 0.195 3.82 0.238 4.78
β -Sitosterol 29.124 0.185 3.93 0.196 4.94
3.2.3. Stability
Stock standard solutions and working standard solutions were found to be stable for at least 3 months,
respectively, when stored at -20 0
C. Moreover, the stability of a fortified blank sample at a concentration of 50
mg/g kept in the auto injector for 24 hours was assayed, and differences of < 5.0 % were obtained.
3.2.4. Limit of Detection, Limit of Quantification, Measurement Uncertainty
The limit of detection (LOD) and the limit of quantification (LOQ) were calculated according to the
formulas LOD = 3.3 · SD/slope and LOQ = 10 · SD/slope [25].
Table 6. Statistical data for LOD, LOQ, Ux for FAs and sterols
Compound LOD (mg/kg) LOQ (mg/kg) Ux (%)
C4:0 0.22 0.73 12.5
C6:0 0.18 0.59 14.8
C8:0 0.16 0.53 11.3
C10:0 0.19 0.63 11.9
C12:0 0.21 0.69 10.8
C14:0 0.15 0.49 9.6
C16:0 0.12 0.40 11.6
C16:1 0.15 0.49 10.5
C18:0 0.16 0.53 8.5
C18:1 0.10 0.33 9.5
C18:2 0.11 0.36 11.2
C18:3 0.12 0.40 9.8
Cholesterol 0.18 0.59 13.7
Stigmasterol 0.25 0.82 13.4
Campesterol 0.22 0.73 11.2
β -Sitosterol 0.26 0.86 12.4
The values for measurement uncertainty (Ux) were calculated according to a Eurachem/CITAC Guide
[26]. The obtained values showed in Table 5, ranged from 8.5% to 14.8%.
3.3. Application of the method
In order to demonstrate the applicability of the proposed method for the determination of FAs and
sterols in the total diet, the 2 week experiment was conducted. Sampling was done according to the
methodology of Thomas et al. [27]. The results of the investigation are presented in Fig. 3 and Fig. 4,
respectively.
8. Valdiation and application of a gas chromatographic method for the determination of fatty…
18
Figure 3. Distribution of the most abundant FAs in the total diet
Figure 4. Distribution of the most abundant sterols in the total diet
As can be seen from the obtained results, the most abundant FA in the total diet is monounsaturated
C18:1 with 34.5% of the total FAs. The lowest presence was observed for the short chain FAs: C4:0 – C12:0 (6.4%).
Cholesterol was found to be the most abundant sterol in the diet (57.5%). The total amount of the plant sterols in
the diet was found to be 42.5%.
IV. CONCLUSION
A reliable, accurate and precise GC-FID method, after extraction of the samples with ASE,
saponification of the lipid fraction and extraction of the FAs and sterols with n-hexane from the same solution
was developed and validated. With the usage of ASE technique, extraction of lipids from the samples was
performed within 15 minutes. The proposed extraction procedure of the FAs and sterols enables the single step
requiring the same organic solvent. The proposed method, reduce the amount of co- extractible lipids and
disables the appearance of the background peaks in the chromatogram, above signal to noise ratio of 3, at the
retention times of targeted compounds. FID was chosen for this analysis due to its selectivity for the compounds
of interest. Validation parameters obtained for determination of the FAs and sterols in the total diet demonstrate
that the developed analytical method meets the method performance acceptability criteria (mean recovery in the
range 80.8% to 98.2%, precision with RSD < 8.6%; LOQ < 0.86 mg/kg; measurement uncertainty < 14.8%).
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