XIV
h
International Conference on Molecular Spectroscopy, Białka Tatrzańska 2017
216
T2: P–1
2D ATR FTIR correlation analysis adaxial and abaxial surfaces of
Urtica dioica
leaves
Paulina Moskal
1
, Aleksandra Wesełucha-Birczyńska
1
,
Maria Łabanowska
1
, and Maria Filek
2
1
Faculty of Chemistry, Jagiellonian University, Krakow, Poland, e-mail: paulina.moskal@uj.edu.pl
2
Institute of Plant Physiology, Polish Academy of Sciences, Krakow, Poland
Urtica dioica
occurs widely all around the world. Although it is often referred to as a weed,
the content of valuable biological substances making that nettle can be considered as an
important source of medicinal and cosmetic ingredients [1].
Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) is a very
useful non destructive method in the study of plant material. The aim of this study was to
compare adaxial and abaxial surfaces of leaf during their growth from two areas of different
environmental pollution. Leaves were collected every month between March and May, from
natural habitats of Bieszczady Mountain and from the south-eastern part of city of Kraków,
district Piaski Wielkie. Lyophilized nettle leaves were investigated using ATR-FTIR technique.
The measured ATR-FTIR spectra for the upper and lower leaf surfaces, from both areas,
were relatively similar. Therefore, 2D correlation method was applied to find changes and
differences in their spectra. ATR-FTIR spectra were used as an imput data while the time of
leaves collection was regarded as an external perturbation [2, 3]. The 2D ATR-FTIR
synchronous and asynchronous correlation maps were analyzed in the 1800–900 cm
−1
range. 2D
synchronous correlation maps for samples coming from the area of the Bieszczady Mountains
and Krakow are different in pattern. For the samples from Bieszczady changes occur mainly in
the cell wall polysaccharides, especially of pectin at 1100 cm
−1
[4] and cellulose at 987, 1000
and 1067 cm
−1
[4–6]. Meanwhile, for samples from Kraków the significant changes take place in
the bands characteristic for Amide I, Amide II and also polysaccharides at 1365 cm
−1
and pectin
at 1106 cm
−1
[4–6]. In both samples, clearer changes were observed in abaxial surfaces of leaf.
Asynchronous correlation maps for samples from both areas are very similar to each other.
Cross-peaks appear roughly in the same positions. However, some of cross-peaks demonstrate a
different order of changes, what indicates the influence of the environment. Additionally the
upper and lower leaf surfaces, for both areas, are differed mainly the intensity of the cross peaks.
Keywords: Urtica dioica, adaxial and abaxial leaf surface; attenuated total reflectance FT-IR spectroscopy
(ATR-FTIR); 2D Correlation
References
[1] R. Upton, 3 (2013) 9.
[2] I. Noda, 47 (1993) 1329.
[3] 2Dshige (c) Shigeaki Morita, Kwansei-Gakuin University, 2004-2005.
[4] J.K.C. Rose, The Plant Cell Wall, Blackwell Publishing Ltd, Oxford, 2003
[5] H. Schulz, M. Baranska, 43 (2007) 13–25
[6] S.T. Gorgulu, M. Dogan, F. Severcan, 61(3) (2007) 300.
XIV
h
International Conference on Molecular Spectroscopy, Białka Tatrzańska 2017
217
T2: P–2
Human leukocytes in malaria investigated by Raman spectroscopy
Aleksandra Wesełucha-Birczyńska
1
, Anna Kołodziej
1
, Jacek Czepiel
2,3
,
Paulina Moskal
1
, Malwina Birczyńska
3
, Grażyna Biesiada
2,3
,
and Aleksander Garlicki
2,3
1
Faculty of Chemistry, Jagiellonian University, Kraków, Poland; e-mail: birczyns@chemia.uj.edu.pl
2
Department of Infectious Diseases, Jagiellonian University, Medical College, Kraków, Poland
3
Department of Infectious Diseases, The University Hospital in Kraków, Kraków, Poland
Along with tuberculosis and AIDS, malaria remains the biggest public health problem in the world.
According to the World Malaria Report created by WHO in 2016 [1], about 212 million cases of malaria
was reported in 2015, and half of million have died. It was also estimated that malaria caused about 70% of
global mortality in children under 5. More than 90% of incidences happen in Africa Region, what means
malaria is a tropical disease. However, the endemic region of malaria is a lot larger than Africa and includes
91 countries, inhabited by 3 billion of people, which makes about 40% of total world population.
Malaria is a parasitic disease causes by one of Plasmodium parasites species, among which P.
falciparum is the most human life-threatening one. The vectors of malaria are female Anopheles mosquitoes
which spread the disease to people by the bites. Firstly, the invasive forms of parasite infect human liver
cells where they mature. After that, they are ready to attack erythrocytes, where they undergo replications
leading to red blood cells’ rupture [2]. The clinical malaria symptoms are non-characteristic, typically
include fever, shivers, vomiting and headaches. Untreated malaria quickly transforms to severe malaria,
which usually results in death [3].
The mechanism of disease remains unknown in lots of areas. It is observed that the leukocyte counts is
low and their functions are blocked during malaria infection, even if erythrocytes are likely to rupture. One
of the reasons causes this unusually occurrence may be disturbance and even inhibition of macrophages
function by phagocytosed hemozoin, which is plasmodium hemoglobin degradation product [4]. In
addition, the excessive immune activation by parasites, mostly neutrofiles and also macrophages, is
believed to lead to the immunopathology [5].
Raman microspectroscopy was used to follow changes in white blood cells (leukocytes) during malaria
infection. Raman spectra of the blood samples were obtained from 5 hospitalized malaria infected patients,
who
were
being
treated
in
the
Department
of
Infectious
Diseases,
Jagiellonian
University
Hospital
in
Krakow.
Leukocytes are quite complex cells, therefore the analysis of obtained spectra was a difficult task.
However, the PCA (Principal Components Analysis) showed that there are separation between the spectra
that were taken in the first and seventh hospitalization day, for big macrophage cells, of about 15 μm
diameter, and also for neutrofiles, of about 10 μm diameter. The most prevalent bands, that come from the
cells in initial stadium of disease and differ from that after treatment appear at about 2970 cm
–1
(in plane
asymmetrical CH stretching vibrations of CH
3
), 2945 cm
–1
(symmetrical CH stretching vibrations of CH
3
),
1490 cm
–1
(CH
2
and CH
3
deformation vibrations), seem to reflect the formation of lipoperoxides within cell
as a result of inhibition of macrophage function [4, 6]. Neutrophils on the first day of the disease
characterizes the 1358 cm
–1
vibration of heme ν
4
mode [7]. It is worth noting that PCA analysis revealed
that leukocytes after treatment, although the morphology indicates full recovery from malaria, still differ
from the healthy ones (obtained from healthy volunteers).
Our study confirmed showed that Raman microspectroscopy is useful technique to follow changes in
white blood cells during malaria.
Keywords: Malaria; Leukocytes; Raman micro-spectroscopy; PCA
References
[1] World Health Organization, “World malaria report 2016”, pp. 39.
[2] “Malaria, Parasite Biology”, DPDx - Laboratory Identification of Parasitic Diseases of Public Health
Concern, CDC Centers for Disease Control and Prevention, 3 May 2016, Web. 24 Apr. 2017
[3] J. Knap, P. Myjak, Malaria in Poland and in the world - yesterday and today, (in Polish), Alfa Medica Press,
Bielsko-Biała, 2009.
[4] B. Urban, D. Roberts, Current Opinion in Immunology 14 (2002) 458.
[5] C. Chua, G. Brown, J. Hamilton, S. Rogerson, P. Boeuf, Trends in Parasitology 29 (2013) 26.
[6] A.T. Tu, Raman Spectroscopy in Biology: Principles and Applications, John Wiley&Sons, New York, 1982.
[7] M.
Kozicki,
J.
Czepiel,
G.
Biesiada,
P.
Nowak,
A.
Garlicki,
A.
Wesełucha-Birczynska,
Analyst
140
(2015)
8007.
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