Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
New Sequential Model for Human Hemoglobin: Alpha Subunit as Cooperativity Inducer
148
153
EN
Mohammad Reza
Dayer
0000-0003-0459-3133
Department of Biology, Faculy of Science, Shahid Chamran University of Ahvaz, Ahvaz, Iran
mrdayer@scu.ac.ir
Mohammad Saaid
Dayer
Department of Parasitology and Medical Entomology, Tarbiat Modares University, Tehran, Iran
dayer@modares.ac.ir
Bahador
Taheri
Department of Biology, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, Iran
b.taheri92@yahoo.com
Hemoglobin is a tetrameric oxygen transport protein in animal bodies. However, there is a paucity of information regarding differences between alpha and beta subunits of hemoglobin in terms of oxygen affinity. The sequential model of Koshland, Nemthy and Filmer (KNF model) has attributed similar affinities to both alpha and beta subunits. The main purpose of the present study is to construct a new sequential model for hemoglobin oxygenation based on higher oxygen affinities for alpha subunits. To this end, coordinate files of 19 oxy and 41 deoxy hemoglobin structures were used as starting structures. These files were processed using Microsoft Excel and SPSS software in order to calculate Euclidean distances between each pair of proximal and distal histidine Fe<sup>2+</sup> as well as other pairs of atoms of interest. The calculated distances were then compared for either set of hemoglobin conformations, <em>i.e.</em> oxy and deoxy conformations. Our results showed that α<sub>2 </sub>subunit show higher structural changes that could be related to oxygen affinity. This subunit could be introduced as initiator of hemoglobin oxygenation and cooperativity. Subunit α<sub>2</sub> in our sequential model induces relaxed conformation in α<sub>1</sub>, β<sub>2</sub> and β<sub>1</sub> respectively. The order of oxygen affinity in our model is as follow: α<sub>2</sub> > α<sub>1</sub> > β<sub>1</sub> > β<sub>2</sub>.
hemoglobin,Tens,Relax,Conformation,Cooperativity
https://www.bmmj.org/article_18574.html
https://www.bmmj.org/article_18574_70c9c5f368d3ad0059092913e75ff6cc.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Studies of Interaction between Propranolol and Human Serum Albumin in the Presence of DMMP by Molecular Spectroscopy and Molecular Dynamics Simulation
154
166
EN
Fatemeh
S.
Mohseni-Shahri
Department of Chemistry, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran
fmohsenishahri@gmail.com
Mohammad
R.
Housaindokht
0000-0002-5428-2512
Department of Chemistry, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran
housain@um.ac.ir
Mohammad
R.
Bozorgmehr
Department of Chemistry, Mashhad Branch, Islamic Azad University, Mashhad, Iran
mr_bozorgmehr@yahoo.com
Ali
A.
Moosavi-Movahedi
Institute of Biochemistry & Biophysics, University of Tehran, Tehran , Iran
moosavi@ut.ac.ir
The interaction between propranolol (PROP) and human serum albumin (HSA) was studied in the presence of dimethyl methylphosphonate (DMMP). DMMP is usually considered as a simulant for chemical warfare agents (CWAs). For this purpose fluorescence quenching, resonance light scattering (RLS), synchronous, three-dimensional fluorescence spectroscopy and molecular dynamics (MD) simulation were employed under physiological conditions. Fluorescence spectroscopy showed that DMMP could quench, and PROP increased intensity of the HSA fluorescence spectra. The presence of DMMP remarkably decreased binding constant of PROP to HSA. Therefore, by decreasing the amount of drugs transported to its target, the free drug concentration of the target would be raised, increasing the efficacy of the drug. The results of synchronous fluorescence and three-dimensional fluorescence spectra showed that the binding of PROP and DMMP to HSA induced conformational changes of HSA. According to molecular dynamics simulation results proposed that these ligands could interact with the HSA, with affecting the secondary structure of protein and with a modification of its tertiary structure.
Human Serum Albumin,Dimethyl methylphosphonate,Propranolol,Molecular Dynamics Simulation,Flourescence quenching
https://www.bmmj.org/article_18578.html
https://www.bmmj.org/article_18578_c0c8a85ab26d8abd1cfb4e5ee3c4f7f3.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
A Comparative Study of Activity and Stability of the Free and the Immobilized Endoglucanase from Alicyclobacillus Acidocaldarius
167
176
EN
Parastou
Rahimizadeh
Department of Cellular and Molecular Biology, Faculty of Science, Azarbaijan Shahid Madani University, Tabriz, Iran.
parastou.biology@gmail.com
Saeed
Najavand
Department of Cellular and Molecular Biology, Faculty of Sciences, Azarbaijan Shahid Madani University, Tabriz, Iran
saeed.najavand@gmail.com
Mohammad
Pazhang
Department of Cellular and Molecular Biology, Faculty of Science, Azarbaijan Shahid Madani University, Tabriz, Iran.
mpazhang@yahoo.com
AaCel9A [β-1,4-endoglucanase, (E.C:3.2.1.4)], was immobilized onto glutaraldehyde activated chitosan macrosphere by covalent attachment. The properties of the immobilized AaCel9A were investigated by determining the optimum pH and optimum temperature for activity, thermal stability, and kinetic parameters. The immobilization process shifted the enzyme’s optimum temperature from 65 °C for the free enzyme towards a wider temperature range from 60-80 °C by the immobilized enzyme. The optimum pH of immobilized AaCel9A shifted to basic pH (pH 8) relative to free AaCel9A (pH 6.5). The immobilization on chitosan macrosphere enhanced half-life of AaCel9A enzyme. After 60 min, the immobilized and the free enzyme retained 75% and 40% their activity at 65 °C, respectively. The immobilized enzyme showed higher thermal stability than the free form. Km value of immobilized AaCel9A (17.05 mg ml<sup>-1</sup>) was higher than free AaCel9A (7.75 mg ml<sup>-1</sup>). Also, CMC hydrolysis by immobilized and free AaCel9A in the presence of SDS detergent was investigated. The results showed that the immobilized enzyme maintained its activity more than the free form in different concentrations of SDS.
AaCel9A,Immobilization,Chitosan,thermal stability
https://www.bmmj.org/article_18599.html
https://www.bmmj.org/article_18599_7ed111c98c75b42a3d31cf755e530ee0.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Selective Determination of Dopamine in the Presence of Ascorbic Acid and Uric Acid at Neutral pH Using a Silver Nanoparticles-modified Carbon Paste Electrode
177
186
EN
Sedigheh
Hashemnia
75169
Department of chemistry, Persian Gulf University, Bushehr 75169, Iran
shashemnia@pgu.ac.ir
Fatemeh
Nourmohammad
Persian Gulf University, Bushehr 75169, Iran
fnourmohammad@yahoo.com
Developing simple, sensitive and selective sensing systems for dopamine is important due to its biological significance. In this work, a silver nanoparticles-modified carbon paste electrode (AgNPs-CPE) has been constructed and used to detect of dopamine (DA) in the simultaneous presence of ascorbic acid (AA) and uric acid (UA) at neutral pH 7.0 by cyclic voltammetry. The modified electrode showed good performance toward the oxidation and determination of DA in the presence of AA and UA. In a mixture of the three compounds, DA showed a pair of redox peaks at about 182.0 and 116.0 mV for anodic and cathodic peaks potential, respectively, while AA and UA exhibited an oxidation peak at about 320.0 mV. Under these circumstances DA more easily oxidized than AA and UA at the surface of modified electrode and precisely determined by differential pulse voltammetry. A sensitivity of 0.074 μA/μM with a wide linear range of 12.5-300.0 μM and detection limit of 0.61 μM were obtained. The modified electrode was applied successfully for DA quantification in dopamine hydrochloride injection sample in the presence of AA and UA (100 μM).
Dopamine,Ascorbic acid,Uric acid,Silver nanoparticles,Carbon paste electrode
https://www.bmmj.org/article_18649.html
https://www.bmmj.org/article_18649_6d9cd431be12a139036c7dc9ee7f15c6.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Change in Oxygen Absorption of Human Adult and Fetal Hemoglobin Due to 940 MHz Electromagnetic Field Radiation Exposure
187
195
EN
Aghdas
Banaei
Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran
banaei@ibb.ut.ac.ir
Hedayatollah
Ghorchian
Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran
hadi@ibb.ut.ac.ir
Reza
Faraji Dana
School of Electrical and Computer Engineering, University of Tehran, Tehran, Iran
reza@ut.ac.ir
Ali Akbar
Moosavi Movahedi
Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran
moosavi@ut.ac.ir
H
Naghavi
School of Electrical and Computer Engineering, University of Tehran, Tehran, Iran
S.M.
Saviz
School of Electrical and Computer Engineering, University of Tehran, Tehran, Iran
R
Amjadi
Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran
R
Hosseinzadeh
Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran
The effects of electromagnetic fields (EMFs) radiation at the frequency of 940 MHz on the structure and function of human adult and fetal hemoglobin (HbA and HbF) were studied. After extraction and purification of HbA and HbF, the oxygen absorption values for exposed and unexposed HbA and HbF to EMF were compared. The slope of oxygen absorption curve for exposed HbA was increased while that for HbF was decreased compare to those before EMF exposing. Furthermore, the oxygen absorption saturation values were changed from 3.4-5.1 and from 5.1-3.1 mg l<sup>-1</sup>, respectively for HbA and HbF after exposing to EMF. The UV-Vis, circular dichroism and fluorescence spectroscopy confirmed the quaternary structural changes of both proteins after EMF exposure. So that, the structural transition of HbA from tense to relaxed state caused an increasing in oxygen absorption; whilst in HbF, transition from relaxed to tense state was occurred and therefore oxygen absorption was decreased.
Electromagnetic fields radiation,Fetal hemoglobin,Conformational changes,Structural transition
https://www.bmmj.org/article_18650.html
https://www.bmmj.org/article_18650_8ceedbb5ee08be3be09b0fa59d737d02.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Evaluation of Immobilized Bacteriorhodopsin’s Function by Laser Irridiation
196
203
EN
Ahmad
Molaeirad
0000-0001-9787-6911
1- Department of bioscience and biotechnology, Malek-Ashtar university of technology, Tehran, Iran
molaeirad@gmail.com
Monireh
Besharati Vineh
2- Department of Biology, College of Basic Science, Tehran Science and Research Branch, Islamic Azad University, Tehran, Iran
besharati_85_m@yahoo.com
Mahdi
Alijanianzadeh
1- Department of bioscience and biotechnology, Malek-Ashtar university of technology, Tehran, Iran
alijanianzadeh_m@yahoo.com
Maryam
Khayati
1- Department of bioscience and biotechnology, Malek-Ashtar university of technology, Tehran, Iran
khayati293@yahoo.com
Bacteriorhodopsin (BR) is a retinal protein that is a light-driven proton pump and has an important role in photosynthesis in archaebacterium Halobacterium salinarum. The BR molecule absorbs light and photochemical changes occur in it, and different intermediates will be produced in its photochemical cycle that some of them like P and Q intermediates have a long half-life. There have been many efforts to immobilize BR for constructing data storage devices. In this study the BR suspension-contained film has been immobilized on a polycarbonate and the effects of green and red lasers on it (in different times and temperatures) have been investigated by UV spectrophotometer method, The modified surfaces were characterized by ATR-FTIR and AFM techniques. The results show that with used condition for immobilization of BR on polycarbonate, the 0 and 1 bite that relate to BR and P, Q intermediate, are formed. The red and green lasers convert BR to O and, P or Q intermediates respectively that could be used instead of 0 and 1 bites in popular compact disks.
Bacteriorhodopsin,Polycarbonate,Laser,Immobilization
https://www.bmmj.org/article_18686.html
https://www.bmmj.org/article_18686_7b7ef4a7c1f8ac46fd50402035681c8a.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Mathematical Analysis of Drug Release for Gastrointestinal Targeted Delivery Using β-Lactoglobulin Nanoparticle
204
211
EN
Behafarid
Ghalandari
Department of Medical Nanotechnology, Science and Research Branch, Islamic Azad University, Tehran, Iran
Applied Biophotonics Research Center, Science and Research Branch, Islamic Azad University, Tehran, Iran
behafarid.gh@gmail.com
Adeleh
Divsalar
Department of Biological Sciences, Kharazmi University, Tehran, Iran
divsalar@khu.ac.ir
Ali
Komeili
Department of Nutrition, Science and Research Branch, Islamic Azad University, Tehran, Iran
Applied Biophotonics Research Center, Science and Research Branch, Islamic Azad University, Tehran, Iran
ali_komeili43@yahoo.com
Mahbube
Eslami-Moghadam
Chemistry and Chemical Engineering Research Center of Iran, Tehran, Iran
eslami_moghadam@ccerci.ac.ir
Ali Akbar
Saboury
Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
Center of Excellence in Biothermodynamics, University of Tehran, Tehran, Iran
saboury@ut.ac.ir
Kazem
Parivar
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran
kazem_parivar@yahoo.com
To answer challenge of targeted and controlled drug release in oral delivery various materials were studied by different methods. In the present paper, controlled metal based drug (Pd(II) complex) release manner of β‑Lactoglobulin (β-LG) nanoparticles was investigated using mathematical drug release model in order to design and production of a new oral drug delivery system for gastrointestinal (GI) tract. The β-LG nanoparticles containing Pd(II) complex were fabricated in the presence of low methoxyl pectin (LMP) at different conditions. Characterization of β-LG nanoparticles using dynamic light scattering (DLS) and atomic force microscopy (AFM) were performed. The <em>in vitro</em> drug release studies were carried out at 37 °C during 8 h in the simulation conditions of GI fluid. The obtained results were fitted in various kinetically release models. The Korsmeyer-Peppas model was evaluated the best describe of each simulation conditions such results indicated an anomalous diffusion manner for drug release. The release data were fitted to the Kopcha model; then, using statistically evaluation revealed that β-LG nanoparticles-LMP complex contain Pd(II) complex dramatically sensitive to pH. In addition, results indicated that for drug release from β-LG nanoparticles delivery system erosion is predominate. So, the erosion-controlled is drug release mechanism of this delivery system. We concluded that β-LG nanoparticles complex with LMP based on mathematical drug release model would be a targeted and practical promising device for GI drug delivery.
β‑LG nanoparticle,Drug release,Gastrointestinal tract,mathematical modelling,Kopcha model
https://www.bmmj.org/article_18687.html
https://www.bmmj.org/article_18687_34b7ee1c9e2e9ecbbc30f843db18b45c.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Fructation Induces Hemin Degradation in Methemoglobin
212
219
EN
Mehran
Habibi Rezaei
School of Biology, College of Science, University of Tehran, Tehran, Iran
mhabibi@ut.ac.ir
Mostafa
Bakhti
School of Biology, College of Science, University of Tehran, Tehran, Iran
mostafa.bakhti@helmholtz-muenchen.de
Ali Akbar
Moosavi-Movahedi
Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
moosavi@ut.ac.ir
Seyed Jalal
Zargar
chool of Biology, College of Science, University of Tehran, Tehran, Iran
zargar@khayam.ut.ac.ir
Hedayatollah
Ghorchian
0000-0003-0541-9209
Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
ghourchian@ut.ac.ir
Under prolonged diabetic conditions, HbA<sub>1c</sub> is produced from normal hemoglobin (HbA<sub>0</sub>) through a non-enzymatic glycosylation or glycation, a process which enhances the hemoglobin susceptibility to be auto-oxidized to methemoglobin (metHb). Here we are reporting that the non-enzymatic reaction between fructose and metHb (metHb fructation) induces significant changes in the globin structure and degrades the hemin moiety. Moreover, glycation of metHb by fructose brings about the extensive hypochromic effect and a small bathochromic effect in the Soret region. The products of hemin degradation were shown to be dialyzable species. Cathodic peaks of the cyclic voltammogram (CV) of methemoglobin during incubation with fructose resulted positive potential shift and the declined current at the peaks due to a decrease in the number of metHb molecules with the preserved hemin groups. Moreover, we found a significant increase in the hemin oxidation products such as ferryl/oxoferryl-Hb upon fructation of metHb. The production of such species during metHb fructation and hemin degradation might have a key role not only in the induction of hypoxic stress, but also in the reduced protective function of neural hemoproteins in Alzheimer’s disease (AD).
Hemin,Fructation,Reactive Oxygen Species,Methemoglobin,Cyclic voltammetry,HbA1c
https://www.bmmj.org/article_18688.html
https://www.bmmj.org/article_18688_1a5f8160b2ca56b4c0609562a8210cca.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Tumor Suppressor p53 Can Protect Normal Cells Against Dendrosomal Curcumin-Induced Apoptosis
220
229
EN
Alemeh
Heidarzadeh
Department of Genetics, School of Biological Sciences, Tarbiat Modares University, Tehran, Iran
heidarzade.al@gmail.com
Ali Mohammad
Ahadi
Department of Genetics, School of Sciences, Shahrekord University, Shahrekord, Iran
ahadi_al@sci.sku.ac.ir
Alireza
Panahi
School of Basic Sciences,University of Mohaghegh Ardabili, Ardabil, Iran
arpanahi@uma.ac.ir
Majid
Sadeghizadeh
Department of Genetics, School of Biological Sciences, Tarbiat Modares University, Tehran, Iran
sadeghma@modares.ac.ir
Curcumin is a natural substance with anti-cancerous properties without many disadvantages of currently-used anticancer drugs. Its toxicity is significantly higher in tumor cells compared with normal cells. We hypothesized the difference of p53 function between normal and tumor cells as one of the presumable causes of this phenomenon. We knocked down the expression of p53 in normal fibroblasts using anti-p53 siRNA and subsequently explored the effects caused by dendrosomal curcumin- a novel nanoformulation of curcumin- on these cells in terms of apoptosis induction and gene expression analysis. The results of MTT assay demonstrated dendrosomal curcumin is selectively cytotoxic for melanoma cancer cells without any considerable effects on normal fibroblasts. Knocking-down of p53 in normal fibroblast cells caused increase of NF-κB1 and decrease of p21 expression level. Treating p53-suppressed normal fibroblast cells with dendrosomal curcumin led to a robust increase in apoptosis rate of the cells. Taken together, these results imply the fact that p53 can protect normal cells from dendrosomal curcumin-induced apoptosis.Therefore, dendrosomal curcumin- in addition to being a chemotherapeutic compound-represents potential capacities to be used as an effective chemopreventive agent.
Cancer,Dendrosomal curcumin,Normal fibroblast,p53,Knockdown,Apoptosis
https://www.bmmj.org/article_14417.html
https://www.bmmj.org/article_14417_c71df3935574ea9a5597499dc998e17d.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Molecular Dynamics and Molecular Docking Studies on the Interaction between Four Tetrahydroxy Derivatives of Polyphenyls and Beta Amyloid
230
241
EN
Najmeh
Mahmoodabadi
School of Chemistry, Damghan University, Damghan, Iran
mahmoodabadi_n@yahoo.com
Davood
Ajloo
School of Chemistry, Damghan University, Damghan, Iran
ajloo@du.ac.ir
Interactions of 3,3',4,4'-tetrahydroxybiphenyl (BPT) and three isomeric 3,3",4,4"-tetrahydroxyterphenyls (OTT, MTT, PTT) with Alzheimer’s amyloid-β peptide (Aβ) were studied by molecular dynamics simulation and molecular docking. Structural parameters such as Root-mean-square derivations (RMSD), radial distribution function (RDF), helix percentage and other physical parameters were obtained. These inhibitors have been evaluated and compared for their activity against aggregation of Aβ. The results showed that all four compounds successfully inhibit association of Aβ and reduce aggregation of protein. For the tetrahydroxyterphenyls efficacy varies with linker geometry: the <em>ortho</em>-position affords the most successful inhibition and the <em>para</em>-geometry the least perhaps due to differing abilities of these inhibitors to bind amyloid-β peptide. Of the four small inhibitors studied 3,3',4,4'-tetrahydroxybiphenyl (BPT) is the most effective inhibitor. Molecular docking studies have been done to confirm the simulation results. Investigation of binding site and free energy confirmed that the efficiency of interaction with Aβ depends on differing abilities of these inhibitors to bind amyloid-β peptide. Binding energy of BPT is more negative than the other and it significantly decreases for PTT. Self-aggregation of this inhibitor decreases in comparison with BPT; therefore Aβ aggregation in the presence of biphenyl form is higher than terphenyls.
Amyloid-β,inhibitors,Molecular Dynamics Simulation,Molecular Docking
https://www.bmmj.org/article_19161.html
https://www.bmmj.org/article_19161_2ab512ec351980abadd1228ef7710696.pdf
Iran Society of Biophysical Chemistry (ISOBC)
Biomacromolecular Journal
7280-2423
1
2
2015
12
01
Novel Pt(II) Complex and Its Pd(II) Aanalogue. Synthesis, Characterization, Cytotoxicity and DNA-interaction
242
253
EN
Somaye
Shahraki
Department of Chemistry, University of Zabol, Zabol
somaye_shahraki@yahoo.com
Hassan
Mansouri-Torshizi
Department of Chemistry, University of Sistan & Baluchestan, Zahedan
hmtershizi@hamoon.usb.ac.ir
Maryam
Sadeghi
Department of Chemistry, University of Sistan & Baluchestan, Zahedan
sadeghi@yahoo.com
Adeleh
Divsalar
Department of Biological Sciences, Kharazmi University, Tehran
divsalar@khu.ac.ir
Ali-Akbar
Saboury
Institute of Biochemistry and Biophysics, University of Tehran, Tehran
saboury@ut.ac.ir
The ability of small molecules to perturb the natural structure and dynamics of nucleic acids is intriguing and has potential applications in cancer therapeutics. This work reports the synthesis, characterization, cytotoxicity and DNA-binding studies of two cytotoxic and intercalative [M(bpy)(pyrr-dtc)]NO<sub>3</sub> complexes (where M = Pt(II) and Pd(II), bpy = 2,2´-bipyridine and pyrr-dtc = pyrrolidinedithio-carbamate). Binding interaction of these complexes with calf thymus DNA (CT-DNA) was investigated by spectrophotometric, spectrofluorometric and gel filtration techniques. Gel filtration studies indicate that the binding of these complexes with CT-DNA is strong enough not to readily break. The binding constant and the thermodynamic parameters have been determined using absorption measurements. The fluorescence studies indicate that the two complexes bind to CT-DNA through an intercalative mode. The cytotoxic activity of these metal complexes has been tested against chronic myelocytic leukemia K562 cell lines and revealed much lower 50% cytotoxic concentration (C<sub>c50</sub>) than that of cisplatin. We hope that such spectroscopic studies to be indeed helpful in studying the pharmacological response of drugs and design of dosage forms.
Platinum(II)/palladium(II) complexes,DNA-binding,Intercalation,Cytotoxicity
https://www.bmmj.org/article_19297.html
https://www.bmmj.org/article_19297_72aa424aa9951185d608a2ba20be8098.pdf