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Utjecaj ekspresije proteina Tau na sinaptičku plastičnost posredovanu neuroliginom-1 u staničnom modelu Alzheimerove bolesti
No full textAlzheimer's disease (AD) is a progressive, neurodegenerative disorder characterized by the accumulation
of Tau neurofibrillary tangles that correlate with cognitive and memory decline. Synaptic dysfunction and
disruption of synaptic plasticity are among the earliest changes observed in the brains of patients and animal
models of AD. Recent evidence suggests a potential role for neuroligin-1 during the initial stages of AD in
the hippocampus. Neuroligin-1 is a postsynaptic adhesion molecule with a key role in stabilizing glutamatemediated long-term potentiation (LP) and memory formation. Despite the overlapping mechanisms, the
pathological influence of Tau protein on neuroligin-1 is not yet known. The aim of this work is to investigate
whether there is a direct relationship between the change in Tau protein expression and the level of
expression and function changes of neuroligin-1. This pilot study implies that the overexpression of Tau
protein and increased Tau phosphorylation at serine residue Ser262 are related to disruption in neuroligin1 levels in hippocampal neurons, providing valuable insight into their potential roles and interaction in early
AD pathogenesis.Alzheimerova bolest (AB) je progresivan, neurodegenerativni poremećaj karakteriziran nakupljanjem Tau
neurofibrilarnih snopića koji koreliraju s padom kognicije i memorije. Sinaptička disfunkcija i poremećaj
sinaptičke plastičnosti su među najranijim promjenama primijećenim u mozgu oboljelih i animalnim
modelima AB. Nedavni dokazi ukazuju na potencijalnu ulogu neuroligina-1 tijekom početnih stadija AB u
hipokampusu. Neuroligin-1 je postsinaptička adhezijska molekula s ključnom ulogom u stabilizaciji
dugotrajnog potenciranja (DP) posredovanog glutamatom i formiranju pamćenja. Unatoč preklapajućim
mehanizmima, još nije poznat patološki utjecaj proteina Tau na neuroligin-1. Cilj ovog rada je istražiti
postoji li direktna povezanost između promjene ekspresije proteina Tau i promjena razina ekspresije i
funkcije neuroligina-1. Ovo pilot istraživanje ukazuje na povezanost između povećane ekspresije proteina
Tau, hiperfosforilacije proteina Tau na serinskom ostatku Ser262 i poremećenih razina neuroligina-1 u
hipokampalnim neuronima, pružajući uvid u njihovu potencijalnu ulogu i interakciju u patogenezi ranog
stadija AB
Improvement of cinnarizine solubility in the presence of β-cyclodextrin and its derivatives
No full textCinarizin je derivat piperazina koji posjeduje antihistaminsku aktivnost te djeluje kao blokator
kalcijevih kanala. Prema biofarmaceutskom sustavu klasifikacije djelatnih tvari cinarizin se
svrstava u drugu klasu koju karakteriziraju lijekovi slabe topljivosti i visoke permeabilnosti
kroz biološku membranu. Topljivost ovakvih djelatnih tvari poboljšava se ukoliko se
kompleksiraju s ciklodekstrinima. U sklopu ovog diplomskog rada provođenjem
solubilizacijskih ispitivanja istražen je utjecaj β-ciklodekstrina te njegova tri derivata
(hidroksipropil-β-ciklodekstrina, sulfobutileter-β-ciklodekstrina i nasumično metiliranog β-
ciklodekstrina) na topljivost cinarizina u vodenom mediju pri vrijednosti pH od 4,5 (simulirani
duodenalni medij). Razvijena je UV/Vis spektrofotometrijska metoda za kvantitativno
određivanje cinarizina u prisutnosti navedenih ciklodekstrina. Metoda je validirana prema
smjernicama koje propisuje Međunarodna konferencija o usklađivanju tehničkih zahtjeva za
registraciju farmaceutskih proizvoda za primjenu kod ljudi.Cinnarizine is a piperazine derivative that acts as an antihistaminic and calcium channel
blocker. According to the biopharmaceutical classification system of active substances,
cinnarizine belongs to the second class, which is characterized by drugs with low solubility and
high permeability through the biological membrane. The solubility of these active substances
improves if they are complexed with cyclodextrins. The influence of β-cyclodextrin and its
three derivatives (hydroxypropyl-β-cyclodextrin, sulfobutylether-β-cyclodextrin, and randomly
methylated β-cyclodextrin) on the solubility of cinnarizine in aqueous medium at pH 4.5
(simulated duodenal medium) was investigated by conducting phase solubility analyses. An
UV/Vis spectrophotometry method was developed for the quantitative determination of
cinnarizine in the presence of the mentioned cyclodextrins. The method was validated
according to the guidelines prescribed by the International Conference on the Harmonization
of Technical Requirements for the Registration of Pharmaceutical Products for Human Use
Analysis of Dolines on the Southwestern Istrian Planation Surface
No full textPonikve su analizirane na temelju digitalnog modela reljefa razlučivosti 5×5 m. Evidentirani su morfološki parametri ponikava te je provedena morfometrijska analiza poligona ponikava i analiza točkastog uzorka ponikava, primjenom GIS alata. Obodi ponikava su izdvojeni poluautomatskom metodom. Ukupno je ustanovljeno 34 476 ponikava na području istarskog poluotoka te 17 478 na području zaravni jugozapadne Istre. Analizom litostratigrafskih jedinica i morfometrijskih parametara reljefa određen je obuhvat zaravni. Na području zaravni većina ponikava je kružnog oblika, prosječne dubine 5,7 m. Gustoća ponikava iznosi 10,02 pon/km2, a indeks okršenosti 14,33. Najviše ponikava je razvijeno u donjokrednim vapnencima i dolomitima te rudistnim vapnencima. Područja najveće gustoće ponikava se nalaze na nadmorskim visinama iznad 200 m. Uspoređeni su digitalni modeli reljefa različite razlučivosti pri analizi gustoće ponikava i nagiba padina te gustoće i vertikalne raščlanjenosti reljefa. Analizirana je mjera disperzije na više razina. Uspoređeni su rezultati ručne digitalizacije i poluautomatske metode.Dolines were analysed on the basis of a digital relief model with a resolution of 5×5 m. The morphological parameters of the dolines were recorded, and a morphometric analysis of the doline polygons and point pattern analysis of the dolines were performed using GIS. The edges of the dolines were extracted using a semi-automatic method. A total of 34 476 dolines were found in the area of the Istrian peninsula and 17 478 in the area of the Southwestern Istrian Planation Surface. Based on the analysis of lithostratigraphic units and morphometric parameters of the relief, the extent of the Planation Surface was determined. In the area of the Planation Surface, most of the dolines are circular and on average 5,7 m deep. The doline density is 10,02 dolines/km2, and the pitting index 14,33. Most of the dolines are formed in limestones and dolomites of the Lower Cretaceous and in rudist limestones. The areas with the highest doline density are located at altitudes above 200 m above sea level. Digital relief models of different resolutions were compared in the analysis of doline density and slope as well as in doline density and relative relief. The dispersion measure was analysed on several levels. The results of the manual digitisation and the semi-automatic method were compared
Thermodynamics of inclusion reactions of amphiphilic compounds with cyclodextrins and cucurbiturils
No full textRazumijevanje termodinamike inkluzije nepolarnih funkcionalnih skupina u ciklodekstrine i kukurbiturile
otvara put za dizajn novih makrocikličkih receptora, omogućuje njihovu širu industrijsku primjenu te pruža
uvid u hidrataciju lipofilnih gostiju i šupljina navedenih receptora. S ciljem rasvjetljavanja utjecaja strukture
gosta, temperature i otapala na kompleksiranje amfifilnih alifatskih i aromatskih spojeva te njihovih
hidrofobnih analoga s ciklodekstrinima i kukurbiturilima, pripadne su reakcije karakterizirane
mikrokalorimetrijski u širokom temperaturnom rasponu (278 ≤ T / K ≤ 338), a struktura nastalih kompleksa
u otopini istražena je spektroskopijom NMR.
Afinitet kukurbiturila prema svim gostima, s iznimkom 1‑naftola u vodi, zamjetno je veći u
usporedbi s ciklodekstrinima s jednakim brojem monomernih podjedinica. To je u svim slučajevima
posljedica izrazito egzotermnog kompleksiranja s glikolurilnim receptorima, a sustavna kalorimetrijska
istraživanja inkluzije također su ukazala na entropijski nepovoljniju dehidrataciju šupljine kukurbiturila.
Reakcijski toplinski kapaciteti su za veliku većinu reakcija kompleksiranja u vodi negativni, što dovodi do
izražene temperaturne entalpijsko-entropijske kompenzacije. Opažene ovisnosti ΔrH°(T) i ΔrS°(T) u najvećoj
su mjeri uzrokovane promjenama u organizaciji vode oko nepolarnih funkcionalnih skupina gostiju i mogu
se protumačiti njihovim kozmotropnim utjecajem na otapalo pri nižim, odnosno kaotropnim pri višim
temperaturama. Navedena činjenica znatno utječe na termodinamiku inkluzije u ciklodekstrine koja je u
većini slučajeva pri 278 K entropijski kontrolirana. Inkluzija alifatskih derivata u β-ciklodekstrin entropijski
je favorizirana, a u kukurbit[7]uril entalpijski, pri čemu je afinitet oba receptora prema tim gostima veći u
usporedbi s aromatskim analozima. Za razliku od vode, reakcije kompleksiranja u formamidu i etilen‑glikolu
su entalpijski kontrolirane, a ΔrH° gotovo neovisna o temperaturi. Posljedično, iako solvofobna inkluzija nije
ograničena na vodu, temperaturna ovisnost pripadnih termodinamičkih parametara kompleksiranja
posljedica je njezinih specifičnih solvatacijskih svojstava.Understanding the thermodynamics of non‑polar functionalities’ inclusion within cyclodextrins and cucurbiturils paves the way for the design of new macrocyclic receptors, enables their wider industrial application, and provides insights into the hydration of lipophilic guests and cavities of these receptors. To shed light on the influence of guest structure, temperature, and solvent on the complexation of amphiphilic aliphatic and aromatic compounds and their hydrophobic analogs with cyclodextrins and cucurbiturils, the corresponding reactions were characterized microcalorimetrically in a wide temperature range (278 ≤ T / K ≤ 338), and the structure of the resulting complexes was investigated by NMR spectroscopy.
The affinity of cucurbiturils towards all guests, with the exception of 1‑naphthol in water, is noticeably higher compared to cyclodextrins with an equal number of monomeric subunits. This is in all cases due to extremely exothermic complexation with glycoluryl receptors, whereby systematic calorimetric studies of inclusion also indicate the entropically less favorable dehydration of the cucurbituril cavity. Reaction heat capacities are negative for most complexation reactions in water, which leads to pronounced temperature enthalpy-entropy compensation. The observed ΔrH°(T) and ΔrS°(T) dependencies are largely due to changes in the organization of water around nonpolar guest functionalities and can be interpreted by their cosmotropic effect on the solvent at lower, and kaotropic at higher temperatures. This significantly affects the thermodynamics of inclusion within cyclodextrins, which is in most cases entropically controlled at 278 K. The inclusion of aliphatic derivatives is entropically favored compared to their aromatic analogues in the case of β-cyclodextrin, and enthalpically favored in the case of cucurbit[7]uril. This results in higher affinity for aliphatic guests in both cases. Unlike in water, the complexation reactions in formamide and ethylene glycol are enthalpically controlled, and ΔrH° almost independent of temperature. Consequently, although solvophobic inclusion is not a water-limited phenomenon, the temperature dependence of the corresponding thermodynamic complexation parameters is due to its specific solvation properties
Analysis of raw cow's milk sampled in different stages of mastitis by applying 1H and DOSY NMR techniques
No full textGoveđi mastitis je upala mliječnih žlijezda koju najčešće uzrokuju razne patogene bakterije u
kombinaciji s ozljedom žljezdanog tkiva kravljeg vimena. Mastitis znatno utječe na sastav
mlijeka, jer dolazi do promjene u kvaliteti proteina, sastavu masnih kiselina, broju i vrsti
metabolita malih molekulskih masa te koncentraciji laktoze i minerala. Cilj ovog rada je
identificirati metabolite u kravljem mlijeku i usporediti sastav sirovog mlijeka zdravih krava
(2 skupine) te krava oboljelih od subkliničkog (4 skupine) i kliničkog mastitisa (1 skupina)
pomoću tehnika 1H i DOSY (engl. diffusion-ordered spectroscopy) NMR uz supresiju signala
otapala primjenom pulsnog slijeda NOESY (engl. nuclear Overhauser effect spectroscopy).
Asignirani su najintenzivniji signali u spektrima 1H NMR te su identificirani najvažniji
metaboliti u mlijeku. Analizom spektara DOSY NMR izračunate su vrijednosti translacijskih
difuzijskih koeficijenata za identificirane metabolite. Na temelju dobivenih podataka, pojedini
uzorci klasificirani su prema stadiju i uzročniku mastitisa. Na kraju su izračunate i uspoređene
vrijednosti translacijskih difuzijskih koeficijenata sastojaka mlijeka nakon pohrane uzoraka
pri sobnoj temperaturi u različitim vremenskim intervalima.Bovine mastitis is an inflammation of mammary glands usually caused by various pathogenic
bacteria combined with the glandular tissue injury of the cow’s udder. Mastitis significantly
affects the milk composition, since it changes the protein quality, the fatty acid content, the
number and types of small molecule metabolites and the concentrations of lactose and
minerals. The goal of this thesis is to identify metabolites in cow’s milk and to compare the
composition of raw milk from healthy cows (2 groups) and from cows suffering from
subclinical (4 groups) and clinical (1 group) mastitis using 1H and DOSY (diffusion-ordered
spectroscopy) NMR techniques with NOESY (nuclear Overhauser effect spectroscopy)-type
solvent suppression. The most intense signals in 1H NMR spectra were assigned and the most
important metabolites in milk were identified. By analysing DOSY NMR spectra, the values
of translational diffusion coefficients for identified metabolites were calculated. Based on the
obtained data, individual samples were classified according to the stage and causative agent of
mastitis. Finally, the translational diffusion coefficients of milk constituents were calculated
and compared after storage of the samples at room temperature in different time intervals
Atomska orbitala
No full textAtomska orbitala je jednoelektronska valna funkcija koja predstavlja rješenje Schrödingerove
jednadžbe za atome slične vodikovom atomu. Pristup rješavanju je separacija radijalne od
angularne Schrödingerove jednadžbe što vodi do rješivih diferencijalnih jednadžbi. Konačno,
atomska orbitala dobije se kao produkt radijalne funkcije i sfernih harmonika. Sva svojstva
atomskih orbitala određena su s četiri kvantna broja: glavni kvantni broj , orbitalni kvantni
broj , magnetski kvantni broj
i spinski kvantni broj koji daju direktnu poveznicu s
mjerljivim fizikalnim veličinama (
se ne dobije rješavanjem Schrödingerove jednadžbe već
se uvodi naknadno)
Laser ablation-inductively coupled plasma-mass spectrometry method in iron nanoparticles analysis
No full textU ovom radu sintetizirane su nanočestice neutralnog željeza (nZVI) za analizu sorpcije iona neodimija, europija, gadolinija, erbija i uranija iz vodene otopine na njihovu površinu. Sadržaj iona zaostao u vodenoj otopini određen je metodom spektrometrije masa uz induktivno spregnutu plazmu (ICP-MS), a sadržaj vezan na nanočestice željeza metodom spektrometrije masa uz induktivno spregnutu plazmu s laserskim otparavanjem (LA-ICP-MS). Ispitana je mogućnost uporabe polimliječne kiseline kao nosača čvrste faze za analizu metodom LA-ICP-MS. Zamijećena je loša reproducibilnost mjerenja te velika standardna odstupanja mjerenih signala. Za potrebe analize učinkovitosti sorpcije odabranih iona na nanočestice željeza metodom LA-ICP-MS načinjene su pastile uzoraka s mikrokristalnom celulozom. Pripravljena su tri kalibracijska standarda s mikrokristalnom celulozom i kalibracija je provedena uz željezo kao unutarnji standard. Pri analizi opažen je izvrstan linearni odziv te relativno nisko standardno odstupanje pri određivanju odabranih elemenata u tragovima. Analizom vodenih otopina lantanoida metodom ICP-MS nakon sorpcije, utvrđeno je da nZVI uklanjaju ione odabranih elemenata s vrlo visokom učinkovitošću.In this experiment nanoparticles of zero-valent iron (nZVI) were synthesized and a sorption of neodymium, europium, gadolinium, erbium and uranium ions from aqueous solution onto their surface was analyzed. Amount of ions that had remained in aqueous solution after the sorption was determined by inductively coupled plasma-mass spectrometry (ICP-MS), and amount of ions adsorbed onto the surface of nZVI by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS). Applicability of polylactic acid as a solid carrier for LA-ICP-MS analysis was investigated, but poor measurement reproducibility and large standard deviation in measured signals were observed. Sample preparation for LA-ICP-MS analysis of bare nZVI included drying and homogenization by pressing nZVI with microcrystalline cellulose into pellets. Three calibration standards were prepared, and calibration was accomplished with iron as an internal standard. Highly linear response, and reasonably low standard deviations were observed during the analysis of the elements in ppm range. ICP-MS analysis of aqueous solutions after the sorption experiment showed very high sorption efficiency for all elements
Synthesis and biological evaluation of heterocyclic carbamates as human cholinesterases inhibitors
No full textAcetilkolinesteraza (AChE) i butirilkolinesteraza (BChE) su enzimi koji u ljudskom organizmu
hidroliziraju acetilkolin te se stoga istražuju kao terapijska meta u tretmanu
neurodegenerativnih bolesti povezanih s promjenom u koncentraciji tog prijenosnika živčanih
impulsa, naročito Alzheimerovom bolešću. U okviru ovog diplomskog rada sintetizirano je
osam karbamata s piperidinskim ili pirolidinskim supstituentom na dušiku karbamatne skupine.
Struktura spojeva potvrđena je snimanjem spektara 1H i 13C NMR. Za četiri karbamata čija je
čistoća bila veća od 85% određena je sposobnost inhibicije AChE i BChE te je in silico
procijenjena sposobnost prolaska kroz krvno-moždanu barijeru. Svi spojevi pokazali su se
brzim ili jako brzim inhibitorima obiju kolinesteraza s konstantama brzine inhibicije u rasponu
od 103 - 106 mol-1 dm3 min-1 Najbržim inhibitorima AChE pokazali su se spojevi 8 i 5,
piperidinski karbamati s feniletilnim, odnosno piperidinskim, supstituentom na 1-hidroksi-2-
aminoetilnom lancu, redom, dok su najbrži inhibitori BChE bili spojevi 3 (pirolidinski
piperidinski monokarbamat s cikličkim pirolidinskim supstituentom na 1-hidroksi-2-
aminoetilnom lancu na dušiku karbamatne skupine) i 5. Svi ispitivani karbamati pokazali su se
selektivnim inhibitorima BChE u odnosu na AChE. Za ispitivane karbamate je procijenjeno da
mogu proći kroz krvno-moždanu barijeru.Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are enzymes that, in humans,
are responsible for the hydrolysis of acetylcholine and are therefore investigated as therapeutic
targets in the treatment of neurodegenerative diseases associated with a change in the
concentration of that neurotransmitter, especially Alzheimer's disease. In this study, eight
carbamates with piperidine and pyrrolidine substituents on the nitrogen of the carbamate group
were synthesized. The structure of the compounds was characterized by 1H and 13C NMR
spectra. For four carbamates whose purity was higher than 85% the ability to inhibit AChE and
BChE was determined, and ability to pass the blood-brain barrier evaluated in silico. All
compounds proved to be fast or very fast inhibitors of both cholinesterases with inhibition rate
constants in the range of 103 -106 mol-1 dm3 min-1. The fastest AChE inhibitors were compounds
8 and 5, piperidine carbamates with a phenylethyl or piperidine substituent at 1- hydroxy-2-
aminoethyl chain, respectively, while the fastest BChE inhibitors were compounds 3
(pyrrolidine piperidine monocarbamate with a cyclic pyrrolidine substituent on the 1-hydroxy-
2-aminoethyl chain nitrogen of the carbamate group) and 5. All of the tested carbamates were
selective inhibitors of BChE compared to AChE. For the tested carbamates, it was estimated
that they can pass through the blood-brain barrier
Karst forms on the coastal area of the peninsula Ražnjić on Korčula island
No full textPriobalna zona poluotoka Ražnjića na Korčuli obiluje krškim oblicima koji su posebno dobro vidljivi zbog izostanka tla i vegetacije. Glavni cilj ovog rad bio je utvrditi osnovna obilježja krške geomorfologije ove zone. Za 40 mjernih točaka izvršena su mjerenja osnovnih parametara (širine, visine i nagiba obalne zone) in-situ i na digitalnom modelu reljefa rezolucije 1x1 m. Na četiri odabrane lokacije izvršeno je detaljno kartiranje krških oblika duž profila orijentiranih poprečno na pružanja obalne zone. Rezultati su pokazali značajnu raznolikost između krškog reljefa južnih i istočnih obala s jedna te sjevernih i zapadnih obala s druge strane. Južne i istočne obala su uglavnom više, šire i strmije od onih na sjevernom i zapadnom dijelu istraživanog područja. Utvrđeno je da se priobalni krški oblici uglavnom pojavljuju i grupiraju sukladno ekološkoj zonaciji priobalne zone. Kartiranjem su zabilježene različite udubine, škrape, pozitivne rezidualne forme, erozijski ostaci sedimenata te denudirani podzemni krški oblici.The coastal zone of the Ražnjić peninsula on Korčula is rich in karst landforms that are particularly well visible due to the absence of soil and vegetation. The main objective of this work was to determine the basic characteristics of the karst geomorphology of this zone. For 40 measurement points, measurements of basic parameters (width, height and slope of the coastal zone) were carried out in-situ and on a 1x1 m digital relief model. At four selected locations, detailed mapping of karst landforms was carried out along profiles oriented transversely to the length of the coastal zone. The results showed significant diversity between the karst relief of the southern and eastern coasts on the one hand and the northern and western coasts on the other. The southern and eastern coasts are generally higher, wider and steeper than those in the northern and western parts of the research area. It was found that coastal karst landforms mainly occur and are grouped according to the ecological zonation of the coastal zone. The mapping recorded various depressions, karren, positive residual forms, erosional remains of sediments and denuded underground karst landforms
Influence of the final layer and aging time on the hydrophobicity of the chitosan/carboxymethylcellulose polyelectrolyte multilayer ; Intermolecular interactions - student's misconceptions
No full textU istraživačkom dijelu ovog rada naizmjeničnom adsorpcijom pozitivnih i negativnih polielektrolita na čvrstoj podlozi pripravljen je polielektrolitni višesloj kitozan/karboksimetilceluloza (CHI/CMC). Istražen je utjecaj završnog sloja i vremena starenja na površinsku močivost i rast višesloja. Površinska močivost analizirana je tenziometrijskim mjerenjima kontaktnog kuta pri različitim uvjetima relativne vlažnosti. Također, istražena je ovisnost kontaktnog kuta silicijskih pločica o načinu čišćenja supstrata. Rast višesloja praćen je elipsometrijskim mjerenjem debljine u odabranim eksperimentalnim uvjetima. Rezultati su pokazali da završni sloj i vrijeme starenja značajno utječu na površinsku močivost i rast višesloja CHI/CMC.
U metodičkom dijelu rada izrađena je metodičko-didaktička priprema za učenike prvog razreda gimnazije, namijenjena obradi nastavne jedinice Međumolekulske interakcije. Priprema je osmišljena u skladu sa strategijom učenja otkrivanjem i temelji se na odgojno-obrazovnim ishodima i najčešćim učeničkim pogrešnim shvaćanjima vezanim uz koncept. U nastavnom satu povezana je građa molekula i međumolekulske interakcije s fizikalnim svojstvima tvari, temeljenim na makroskopskim opažanjima učenika.In the research section, chitosan/carboxymethylcellulose (CHI/CMC) polyelectrolyte multilayers were prepared by alternating adsorption of polycations and polyanions on a solid surface. The influence of the terminating layer and aging on surface wettability and multilayer growth were investigated. Wettability was analyzed using tensiometric contact angle measurements under different humidity conditions, while multilayer growth was monitored by ellipsometry. The impact of substrate cleaning methods on the contact angle of silicon wafers was also examined. Results showed that the terminating layer and aging significantly affected the hydrophobicity and growth of the CHI/CMC multilayer. In the methodological part, a lesson plan on Intermolecular interactions was created for first- year high school students. The lesson was based on educational outcomes and the most common student misconceptions, designed for strategy of learning by discovery. The lesson connected molecular structure and intermolecular interactions to the physical properties of substances through students' macroscopic observations