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Bioremediation and
Bioeconomy
­Bioremediation and
Bioeconomy

Edited by
M.N.V. Prasad
Department of Plant Sciences
University of Hyderabad, Telangana, India

AMSTERDAM • BOSTON • HEIDELBERG • LONDON • NEW YORK • OXFORD

PARIS • SAN DIEGO • SAN FRANCISCO • SINGAPORE • SYDNEY • TOKYO
Elsevier
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ISBN: 978-0-12-802830-8
This book is dedicated to Challapalli Savithri.
Contributors
S.Z. Ahammad
Indian Institute of Technology, Delhi, India
K. Amulya
Bioengineering & Environmental Sciences; CSIR-Indian Institute of Chemical Technology
(CSIR-IICT), Hyderabad, India
D. Annapurna
University of Hyderabad, Hyderabad, Telangana, India
B. Barbosa
Universidade Nova de Lisboa, Caparica, Portugal
N.S. Bolan
University of Newcastle, Newcastle, NSW, Australia
A. Brandys
Institute of Natural Fibres and Medicinal Plants, Poznan, Poland
E.N.V.M. Carrilho
Universidade Federal de São Carlos, Araras, SP, Brazil
R. Chaturvedi
St. John’s College, Agra, India
T.S. Chibrik
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg, Russia
J. Costa
Universidade Nova de Lisboa, Caparica, Portugal
Shikha Dahiya
Bioengineering & Environmental Sciences; CSIR-Indian Institute of Chemical Technology
(CSIR-IICT), Hyderabad, India
K.C. Das
University of Georgia, Athens, GA, USA
V. Escande
FRE 3673 – Bioinspired Chemistry and Ecological Innovations, – CNRS, University of Montpellier
2, Stratoz – Cap Alpha, Avenue de l’Europe, 34830 Clapiers, France
P.J.C. Favas
University of Trás-os-Montes e Alto Douro, Vila Real, Portugal
A.L. Fernando
Universidade Nova de Lisboa, Caparica, Portugal
E.I. Filimonova
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg, Russia

xxi
xxii Contributors

Edward Gatliff
Applied Natural Sciences, Inc., Hamilton, Ohio, USA
M.A. Glazyrina
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg, Russia
S. Gopalakrishnan
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru,
Telangana, India
C. Grison
FRE 3673 – Bioinspired Chemistry and Ecological Innovations – CNRS, University of Montpellier
2, Stratoz – Cap Alpha, Avenue de l’Europe, 34830 Clapiers, France
U. Jena
Desert Research Institute, Reno, NV, USA
V. Kanaganahalli
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru,
Telangana, India
Srujana Kathi
Pondicherry University, Kalapet, Puducherry, India
J. Koelmel
University of Hyderabad, Hyderabad, Telangana, India
A. Kumar
Indian School of Mines, Dhanbad, Jharkhand, India
G. Labuto
Universidade Federal de São Paulo, São Paulo, SP, Brazil
G. Lemoine
Établissement Public Foncier Nord-Pas de Calais, Euralille, France
P. James Linton
Geosyntec, Clearwater, Florida, USA
N.V. Lukina
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg,
Russia
S.K. Maiti
Indian School of Mines, Dhanbad, Jharkhand, India
M.G. Maleva
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg,
Russia
O. Meesungnoen
Mahasarakham University, Maha Sarakham, Thailand
G. Mohanakrishna
VITO—Flemish Institute for Technological Research, Mol, Belgium
 Contributors xxiii

R. Naidu
University of Newcastle, Newcastle, NSW, Australia
W. Nakbanpote
Mahasarakham University, Maha Sarakham, Thailand
T.K. Olszewski
Organic Chemistry, Wroclaw University of Technology, Wroclaw
D. Pant
VITO—Flemish Institute for Technological Research, Mol, Belgium
E.G. Papazoglou
Agricultural University of Athens, Athens, Greece
M.S. Paul
St. John’s College, Agra, India
C. Phadermrod
Padaeng Industry Public Co. Ltd. (Mae Sot Office), Tak, Thailand
M.N.V. Prasad
University of Hyderabad, Hyderabad, Telangana, India, and Ural Federal University named after
First President of Russia B.N. Yeltsin, Ekaterinburg, Russia
J. Pratas
University of Coimbra, Coimbra, Portugal
M. Rajkumar
Central University of Tamil Nadu, Thiruvarur, Tamil Nadu, India
E.A. Rakov
Ural Federal University named after First President of Russia B.N. Yeltsin, Ekaterinburg,
Russia
P. Srinivas Rao
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru,
Telangana, India
G.A. Ravishankar
Dayananda Sagar Institutions, Bengaluru, Karnataka, India
Douglas J. Riddle
RELLC, Mountain Center, California, USA
D. Rose
University of Hyderabad, Hyderabad, Telangana, India
H. Sarma
N N Saikia College, Titabar, Assam, India
A. Sathya
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru,
Telangana, India
xxiv Contributors

K. Schmidt-Przewoźna
Institute of Natural Fibres and Medicinal Plants, Poznan, Poland
A. Sebastian
University of Hyderabad, Hyderabad, Telangana, India
B. Seshadri
University of Newcastle, Newcastle, NSW, Australia
V. Sivasubramanian
Phycospectrum Environmental Research Centre (PERC), Chennai, Tamil Nadu, India
T.R. Sreekrishnan
Indian Institute of Technology, Delhi, India
S. Srikanth
VITO—Flemish Institute for Technological Research, Mol, Belgium
S. Suthari
University of Hyderabad, Hyderabad, Telangana, India
O. Sytar
Kyiv National University of Taras Shevchenko, Kyiv, Ukraine
J.C. Tewari
Central Arid Zone Research Institute, Jodhpur, Rajasthan, India
R. Thangarajan
University of South Australia, Adelaide, SA, Australia
Paul R. Thomas
Thomas Consultants, Inc., Cincinnati, Ohio, USA
S. Venkata Mohan
Bioengineering & Environmental Sciences; CSIR-Indian Institute of Chemical Technology
(CSIR-IICT), Hyderabad, India
S. Vidyashankar
M/s. DaZiran Health Products, Coimbatore, Tamil Nadu, India
H. Wang
Zhejiang A & F University, Hangzhou, Zhejiang, China
A.A. Warra
Kebbi State University of Science and Technology, Aliero, Nigeria
Foreword

Recent years have seen significant worldwide investment in the bioeconomy, in both political and eco-
nomic terms. The bioeconomy encompasses the production of renewable biological resources and their
conversion into food, feed, bio-based products and bioenergy via innovative and efficient technologies
provided by industrial biotechnology. It is already a reality offering great opportunities and solutions to
major societal, environmental and economic challenges, including climate change mitigation, energy
and food security and resource efficiency.
Bioremediation, on the other hand, is a waste management technique that involves the use of or-
ganisms to remove or neutralize pollutants from a contaminated site. Bioremediation provides a tech-
nique for cleaning up pollution by enhancing the natural attenuation, a process that occurs in nature.
Depending on the site and its contaminants, bioremediation may be safer and less expensive than alter-
native solutions such as incineration or landfilling of the contaminated materials.
This book on “Bioremediation and Bioeconomy” has the ambition to address the research gap at
the interface between pollution mitigation and boosting the bio-based economy aiming at sustainable
development. There is pressing need for pollution abatement and there is an even more pressing need to
create viable value chains from contaminated substrates especially in developing economies.
This book will tackle a number of key questions: How can scientific innovations be more efficiently
exploited to achieve bioremediation and bioeconomy? What are the needs of cutting edge environmen-
tal industries for scale-up and large-scale market penetration? What are important research priorities
for industry and policymakers? and How can environmental sustainability be maintained and simulta-
neously economic sustainability be achieved.
The chapters of this book on bioproducts, biomass energy and biofuels, ornamentals and crops from
contaminated substrates, brownfield development, algal bioproducts, bioprocesses and bioengineering,
and on new biology specifically address the interface and synergies between bioremediation and the
bioeconomy and will therefore contribute to turn into reality the promise to create viable value chains
from contaminated substrates in the context of the emerging global bioeconomy.
Dr. Rainer Janssen
WIP Renewable Energies
Head of Unit Biomass Department
München
Germany

xxv
Preface

Environmental decontamination is an integral part of bioeconomy and sustainable development.

Biodiversity is being used as raw material for environmental decontamination, and this field has grown
phenomenally in recent years, having emerged less than 3 decades ago. On the other hand, the volume
of contaminated substrates (water, soil, and air) is increasing due to anthropogenic and technogenic
sources of organic and inorganic contaminants. Metals are the most prevalent inorganic pollutants/
contaminants and are widely used for a wide variety of needs from building materials to information
technology. Metal contamination is a global problem.
Today with a growing economy extensive industrialization and extraction of natural resources have
resulted in environmental contamination and pollution. Large amounts of toxic waste have been dis-
persed in thousands of contaminated sites spread all over the globe. These pollutants belong to two
main classes: inorganic and organic. The challenge is to develop innovative and cost-effective solutions
to decontaminate polluted environments. In this direction, bioremediation is emerging as an invaluable
tool for environmental cleanup.
Various strategies are being applied to reduce the levels of contamination. Cultivation of industrial
and environmental crops in contaminated soils is one such option. If the contaminant concentration
exceeds the permissible level in edible parts, it poses serious health concerns. Therefore, in such cases
non-edible crop production and valorization to value chain and value additions is a feasible proposition.
The advancement in this field is toward production of diverse biofuels (solid, liquid, and gaseous).
Essentially, bioremediation is centered on bioenergy and use of bioresources harvested from environ-
mentally perturbed/stressed agro-ecosystems. Although bioremediation contractors must profit from the
activity, the primary driver is regulatory compliance rather than manufacturing profit. It is an attractive
technology in the context of bioeconomy. This book will address the bottlenecks and solutions to the ex-
isting limitations in field scale and the relevant techniques. Crucial aspects of biorefinery are also covered.
Globally, land and water resources are under immense pressure due to land degradation, pollu-
tion, population explosion, urbanization, and global economic development. Large amounts of toxic
waste have been dispersed in thousands of contaminated sites, and bioremediation is emerging as an
invaluable tool for environmental cleanup. Bioremediation and Bioeconomy addresses this challenge
by presenting innovative and cost-effective solutions to decontaminate polluted environments, includ-
ing usage of contaminated land and wastewater for bioproducts such as natural fibers, biocomposites,
and fuels to boost the economy.
Bioremediation and Bioeconomy provides a common platform for scientists from various back-
grounds to find sustainable solutions to these environmental issues. This book will also address all the
topical issues crucial for understanding the ecosystem approaches for a sustainable development. It
provides an overview of ecosystem approaches, conservation of natural resources, pollution abatement,
and mitigation.
This book is a collective effort of 65 contributors from 14 countries: Australia, Belgium, Brazil,
China, France, Greece, India, Nigeria, Poland, Portugal, Russia, Thailand, Ukraine and USA.
The book includes 26 chapters grouped in 8 sections:
1. Bioproducts from Contaminated Substrates (Soil and Water) (Chapters 1–4)
2. Biomass Energy, Biodiesel and Biofuel from Contaminated Substrates (Chapters 5–8)

xxvii
xxviii Preface

3. Ornamentals and Crops for Contaminated Substrates (Chapters 9–12)

4. Brownfield Development for Smart Bioeconomy (Chapters 13–16)
5. Algal Bioproducts, Biofuels, Biorefinery for Business Opportunities (Chapters 17 and 18)
6. Bioprocesses, Bioengineering for Boosting Bio-Based Economy (Chapters 19–21)
7. Case Studies (Chapters 22 and 24)
8. New Biology (Chapters 25 and 26)
The book provides a comprehensive review of new information on bio-phyto-rhizoremediation.
Moreover, I feel that this is the first attempt to link bio-phyto-rhizoremediation to bio-based economy
citing several examples. The unique features of this books include (a) numerous color figures, flow
diagrams, tables, and updated literature; (b) strategies to utilize contaminated susbtrates for producing
bioresources and cogeneration of value chain and value addition products boosting bioeconomy; and
(c) several success stories.
M.N.V. Prasad
Department of Plant Sciences
University of Hyderabad
Telangana, India
5 May 2015
Acknowledgments

I would like to thank all the authors of this volume for their cogent and comprehensive contributions.
I am thankful to Candice Janco, Senior Acquisitions Editor, Environmental Science; Laura Kelleher;
Rowena Prasad; and Marisa LaFleur for excellent coordination of this fascinating project, suggestions;
thanks are due to Mr Paul Prasad Chandramohan, and his team for excellent technical help in many
ways that resulted in record time of publication of this edition.
I am grateful to Dr. Rainer Janssen, WIP Renewable Energies, Head of Unit Biomass Department,
Sylvensteinstr. 2, D-81369 Muenchen, Germany, for preparing the Foreword at short notice.
My profuse thanks to Prof. Dr. Irina Kiseleva, Head, Department of Plant Physiology and
Biochemistry, and all faculty members of the department; Ural Federal University, Ekaterinburg,
Russia, for extending help in several ways.
Thanks are due to Dr James Philp (France), Organisation for Economic Co-operation and
Development (OECD), Directorate for Science, Technology and Industry, Paris, and Anonymous re-
viewers from Germany and the United States for constructive comments that enhanced the outlook of
this work.
Last but not least I wish to acknowledge the University of Hyderabad for granting sabbatical leave
that enabled me to concentrate on this work, and the numerous colleagues from overseas, my students,
and research associates for sharing knowledge, ideas, and assistance that helped in developing and
shaping this book.

xxix
 CHAPTER

PRODUCTION OF BIODIESEL
FEEDSTOCK FROM
THE TRACE ELEMENT
CONTAMINATED LANDS
1
IN UKRAINE
O. Sytar1, M.N.V. Prasad2
Kyiv National University of Taras Shevchenko, Kyiv, Ukraine1
University of Hyderabad, Hyderabad, Telangana, India2

1 ­INTRODUCTION
The total area of Ukraine is about 60.37 million ha of which agricultural land is 41.76 million ha.
Ukraine has several rivers and lakes. The prominent crops cultivated for grain production are winter rye,
oats, corn, barley, and buckwheat (in recent years about 15% of the world’s production has come from
Ukraine). Flax, hops, sugar beets, and potatoes are other important crops in Ukraine. Soybean, rape-
seed, and sunflower production has increased in recent years, boosting agro-based economy (Figure 1).
After the Chernobyl accident in 1986, the trace element pollution and contamination in Ukraine
received considerable attention. In total, air pollution-related mortality represents about 6% of total
mortality in Ukraine (Mnatsakanian, 1992; Strukova et al., 2006). In the Donets Basin (48.3°N, 38.0°E)
of eastern Ukraine, there are about 1200 mine-waste dumpsites, mostly from coal mining. These have
accumulated over more than 200 years of mining (Panov et al., 1999). Ore processing and the use of
metal-enriched coal in coal-fired electric generation and home heating have led to widespread con-
tamination of the environment (Panov et al., 1999; Kolker et al., 2009). Of the 26 elements considered
to be pollutants in the Donets Basin, arsenic (As) and mercury (Hg) are of primary concern; most of
the As-polluted soils in this region are near burning-coal waste heaps (Panov et al., 1999; Conko et al.,
2013). For more than 50 years, the largest Hg production facility of the former Soviet Union operated in
Horlivka (Gorlovka, Russian spelling), a city of approximately 300,000 residents in the Donets Basin.
During this period, about 30,000 metric tons of Hg were produced from ore extracted from the adjacent
Mykytivka (Nikitovka) mines.
The contaminated land that is unsuitable for food production and of little or no economic return
could be used for the production of bioenergy. The problem of competition for fertile land can be al-
leviated by using contaminated land for the production of bioenergy plants.
Using biomass for fuel production is an attractive option for three main reasons. First, it is a renew-
able resource and thus its development could be sustained in the future. Second, because there is no net
release of carbon dioxide and sulfur contents are low, it is environmentally attractive. Thirdly, given the
rise in fossil fuel prices, it is also economically advantageous (Demirbas, 2009).
Bioremediation and Bioeconomy. http://dx.doi.org/10.1016/B978-0-12-802830-8.00001-0
Copyright © 2016 Elsevier Inc. All rights reserved.
3
4 CHAPTER 1 TRACE ELEMENT CONTAMINATED LANDS IN UKRAINE

a b

Primary feed-stock for

a = Brassica napus (rape-seed)
b = Helianthus annuus (sunflower)
c = Glycine max (soybean)
FIGURE 1
Primary feed-stock for production of biodiesel in Ukraine.
production of biodiesel in
Ukraine. Brassica and
Helianthus are acknowledged
for their phytoremediation
potential to polish contaminated
soils. The harvested produce is
ideal for bioenergy.

This review chapter presents the nowadays situation of heavy metals pollution on Ukraine territory
and the possibility of using oilseed crops (rapeseed, soybean, and sunflower) for 50% cleanup of the
areas polluted by heavy metals.

2 ­MONITORING OF HEAVY METALS POLLUTION IN UKRAINE

The report of EEA-UNEP (2000) regarding the status of soil contamination in Europe (Figure 2) has
shown that in Eastern Europe problems of diffuse soil contamination is greatest compared to other
European countries. The high contamination of heavy metals is shown in Ukraine, especially in the
Chernobyl area. Unfortunately, the international literature is still missing detailed analysis of heavy
metals contamination in Ukraine.
The Ministry of Nature Protection of Ukraine reported that lead concentrations in the capital city of
Kyiv were 4.6 times higher than the permissible limit. It was reported that the main source of lead in
the air and soil is from automobile activities. Major sources of lead include gasoline combustion, also
nonferrous smelting, and mining (Chiras, 2009).
About 90% of toxic metals accumulate from atmosphere to soil, where they migrate in groundwater,
become absorbed by plants, and get into the trophic chains (Lozanovskaya et al., 1998; Anonymous,
1998; Trahtenberg, 1998; Alekseeva, 1987). According to coefficient of load factor, it has been es-
tablished that in some regions of Ukraine the level of metal pollution of soil, including mobile forms,
exceeds the permissible level by 2-14 times (Gaevsky and Pelypets, 1999; Nikolaychuk and Hrabovsky,
2000). In particular, the content of lead in soils of northern agricultural regions (Zhytomyr, Sumy,
Rivne, Chernihiv, and Kyiv) exceeds the permissible level by 3 to 9 times (Shestapalov et al., 1996).
A similar tendency is observed for soil in the region of Kyiv Polesye, where concentration of metal
exceeds the natural level by 3 times (Brooks, 1982; Shestapalov et al., 1996) (Figure 3).
 2 ­
MONITORING OF HEAVY METALS POLLUTION IN UKRAINE 5

FIGURE 2
Probable problem areas of diffuse contamination in Europe (EEA-UNEP, 2000) * - agricultural areas which can
use chemical during plant cultivation.

Second place among heavy metals highly contaminating Ukraine soil is nickel. It’s known
that the main anthropogenic source of nickel emission is combustion of fossil fuels and traffic
(Krstić et al., 2007). Nickel is widely used in silver refineries, alloy, pigments electroplating,
zinc-based casting, and storage batteries too, which also can have the effect of increasing nickel
pollution.
 6
CHAPTER 1 TRACE ELEMENT CONTAMINATED LANDS IN UKRAINE
150 km

100 mi
Chemihiv
Volyn
Sumy
Rivne
Zhytomyr
Kyiv

Lviv

Poltava Kharkiv
Ternopil Khmelnytskyi

Cherkasy Luhansk
Ivano-Frankivsk
Vinnytsia
Zakarpattia
Kirovohrad Dnipropetrovsk
Chernivtsi
Donetsk

Mykolaiv Zaporizhia
Odessa

Kherson

SEA OF AZOV

Autonomous Republic
Content of lead on territory of Ukraine of Crimea

High level (more than 20 mg/kg) Sevastopol

Average level (more than 10 mg/kg)

BLACK SEA
Low level (4-10 mg/kg)

The map of lead pollution in Ukraine

FIGURE 3
The map of lead pollution on territory of Ukraine.
 3 ­
ECONOMICAL BACKGROUND OF BIODIESEL PRODUCTION IN UKRAINE 7

The greatest Ni content found in soils of the regions of Zhytomyr, Kyiv, Zaporizhia Donetsk, and
Lugansk (in the soil 25-50 mg/kg when maximal permissible level 50 mg/kg) Figure 2. At these regions
it is possible to get relatively safe crops with tolerance to Ni but in limited quantities. The agricultural
land Volyn, Poltava, Vinnytsia, and partly other regions have pollution levels 5-10 and 10-15 mg/kg of
soil areas for ecological agriculture nowadays. In these areas it is possible to grow vegetable raw mate-
rials suitable for the production of baby food. Soils with the average pollution level of Ni (15-25 mg/kg)
(Figure 2) are mainly distributed in Chernihiv, Sumy, Zaporizhya, Ivano-Frankivsk, Rivne, and
Transcarpathian region (Anonymous, 2004). High level of copper found in Kiev, Chernihiv, Zgytomyr,
Volyn, Zakarpatiya regions. Chernivtsi, Kharkiv, Donetsk and Zaporizhya resgins has been character-
ized high level of chromium (Figure 3). In the Ivano-Frankivsk region, Pb concentrations exceed toxic
levels in the areas close to industrial sectors (10% of the region territory). Local background levels of
heavy metals are greatly exceeded in snow close to industrial regions. Cd and Mo accumulate in forest
soils in the Ukrainian Carpathians region (Shparyk and Parpan, 2004) (Figure 3). A similar situation
has been observed with pollution of soil with Zn and Co. The maximal permissible level of Zn and Co
in air is 300 and 50 Clarke, respectively (Clarke is unit of each element and in soil it is 50 and 8 mg/
kg, respectively). In general, for these three heavy metals (Ni, Zn, and Co) in many areas of Ukrainian
territory their content in the soil is at less than maximal permissible level (Figures 4 and 5).
It’s known that the maximum permissible level of Pb in soil is 30 mg/kg of dry soil (Samokhvalova
et al., 2001). The limits of maximum permissible level depend on the complex of soil and ecological
conditions, among which the most important are soil pH and content of hummus (Glazovskaya,
1994; Kabata-Pendias and Pendias, 1989; Obuhov et al., 1980). It has been proved that the main
quantity of heavy metals is concentrated in the upper layer of arable soil (0-10 cm) (Glazovskaya,
1994; Trahtenberg, 1998). The period of heavy metal removal from soil is near 740 to 5900 years (for
example, cadmium and lead) (Dobrovolsky, 1997). Most inorganic compounds of Pb and Cd are insol-
uble or poorly soluble in water. However, their dissolubility depends on soil pH. Dissolubility reaches
a 4.0, but in the range of 6.5-8.0, dissolubility declines distinctly, but after further increasing of pH,
slightly increases (Glazovskaya, 1994). The redox system, level of gross content, and the granulomet-
ric composition of soil have an influence on solubility of heavy metal compounds in the soil (Lysenko
et al., 2001). Soils serve as a comprehensive geochemical barrier that keeps solid compounds and water-
soluble compounds of heavy metals in the form of organo-mineral, mineral complexes, and compounds
(Glazovskaya, 1994). It is known that soil has a buffer capacity as to toxic effect of pollutants, which
provides for transformation of pollutants at not-soluble forms for plants. Granulometric structure of soil
rich in organic matter and carbonate usually has the highest buffer capacity (Grakovsky et al., 1994).

3 ­ECONOMICAL BACKGROUND OF BIODIESEL PRODUCTION IN UKRAINE

AND IN THE WORLD
It is known that humus-rich black soil of Ukraine is world famous. Since the dissolution of the Soviet
Union with ongoing land reform and decreasing activity in the agriculture sector, Ukraine has not been
making full use of its agricultural potential (Schaffartzik et al., 2014).
Ukraine is often singled out for its potential as a supplier of agricultural goods to the EU region. 42 of
60 Mha of Ukraine can be used for crop production (Elbersen et al., 2009). At 0.7 hectare per person (ha/
cap), Ukraine has more land available than any other European country, except for Russia (0.9 ha/cap),
 8
CHAPTER 1 TRACE ELEMENT CONTAMINATED LANDS IN UKRAINE
150 km

100 mi

Chemihiv
Volyn
Sumy
Rivne
Zhytomyr

Kyiv
Lviv
Kharkiv
Poltava
Ternopil Khmelnytskyi
Cherkasy Luhansk

Ivano-Frankivsk Vinnytsia
Zakarpattia
Kirovohrad Dnipropetrovsk
Chemivlsi Donetsk

Zaporizhia
Mykolaiv
Odessa
Kherson

SEA OF AZOV

Autonomous Republic
Content of nickel on territiry of Ukraine of Crimea
High level (25-30 mg/kg)
Sevastopol
Average level (15-25 mg/kg) BLACK SEA
Low level (lass than 10 mg/kg)

The map of nickel pollution in Ukraine

FIGURE 4
The map of nickel pollution on territory of Ukraine.
 3 ­
ECONOMICAL BACKGROUND OF BIODIESEL PRODUCTION IN UKRAINE
150 km

100 mi
Volyn
Chernihiv
Rivne
Zhytomyr Sumy
Kyiv

Lviv Poltava Kharkiv

Ternopil Luhansk
Khmelnytskyl Cherkasy

Zakarpatiya Ivano-Frankivsk Vinnytsia

Kirovohrad Dnipropetrovsk
Chernivtsi Donetsk

Mykolaiv Zaporizhia
Odesa

Kherson

SEA OF AZOV

Autonomous Republic
of Crimea

Area polluted by copper (65-250 mg/kg) Sevastopol

Area polluted by chromium (25-80 mg/kg) BLACK SEA

The map of copper pollution in Ukraine

FIGURE 5
The map of copper and chromium pollution on territory of Ukraine.

9
10 CHAPTER 1 TRACE ELEMENT CONTAMINATED LANDS IN UKRAINE

significantly surpassing France (0.3 ha/cap) and Germany (0.1 ha/cap) (FAOSTAT, 2012), for example.
The country’s geographical location as neighbor to the countries of the European Union and its access to
the Black Sea is additionally advantageous to its role as a global supplier of biomass resources. Ukraine is
a major exporter of agricultural primary products: In 2010 and in physical units, it ranked second among
the international top exporters of barley, third among the exporters of rapeseed, seventh for exports of
sunflower seed, and ninth among the exporters of wheat. Rapeseed production and exports drastically
increased from 2004 onwards and grew by a factor of almost 20 until 2008 (FAOSTAT, 2012).
In Ukraine the sunflower, rapeseed, and soybean are the most widespread oilseed crops that can be
used for phytoremediation process and production of biodiesel and biogas. In Eastern Europe, prob-
lems of diffuse soil contamination are greatest in Azerbaijan, Belarus, Moldova, Russia, and Ukraine
(Denisov et al., 1997). In the total cultivation area of these crops in Ukraine, the area of heavy metals-
contaminated soils is approximately 15-20%.
At the same time biofuels are currently not produced or consumed in relevant amounts within Ukraine.
Yet in Ukraine potential is mainly as an alternative to liquid fossil fuel use and not to the use of natural gas
and coal in industrial applications and electricity generation. The production of biofuel requires energy
inputs. But the biggest advantage that biodiesel has over gasoline and petroleum diesel is its environmen-
tal friendliness. Biodiesel burns similar to petroleum diesel as it concerns regulated pollutants. On the
other hand, biodiesel probably has better efficiency than gasoline. One such fuel for compression ignition
engines that exhibits great potential is biodiesel. Diesel fuel can also be replaced by biodiesel made from
vegetable oils. Biodiesel is now mainly being produced from soybean, rapeseed, and palm oils.
The higher heating values (HHVs) of biodiesels are relatively high. The HHVs of biodiesels
(39-41 MJ/kg) are slightly lower than that of gasoline (46 MJ/kg), petrol diesel (43 MJ/kg), or petro-
leum (42 MJ/kg), but higher than coal (32-37 MJ/kg). Biodiesel has over double the price of petrol die-
sel (Demirbas, 2007). The major economic factor to consider for input costs of biodiesel production is
the feedstock, which is about 80% of the total operating cost. The high price of biodiesel is in large part
due to the high price of the feedstock. Economic benefits of a biodiesel industry would include value
added to the feedstock, an increased number of rural manufacturing jobs, increased income taxes, and
investments in plant and equipment.
The production and utilization of biodiesel is facilitated first through the agricultural policy of sub-
sidizing the cultivation of nonfood crops (rapeseed, for example). Second, biodiesel is exempt from the
oil tax. The European Union in 2005 got nearly 89% of all biodiesel production worldwide. Germany
produced 1.9 billion liters or more than half the world total. In Germany biodiesel is also sold at a lower
price than fossil diesel fuel. Biodiesel is treated like any other vehicle fuel in the UK. Other coun-
tries with significant biodiesel markets included France, the United States, Italy, and Brazil. All other
countries combined accounted for only 11% of world biodiesel consumption. By 2004, the United
States is expected to become the world's largest single biodiesel market, accounting for roughly 18%
of world biodiesel consumption, followed by Germany (EBB, 2004). For Ukraine with its very low
share of renewables in TPES, the promise of reduced environmental burdens is especially appealing
(Schaffartzik et al., 2014).
Rapeseed, as an important feedstock for biofuel, contributed only about 2% to the agricultural
Ukrainian harvest in 2008 but wheat, maize, sugar beets, and sunflower seeds (all also potential feedstock
for biofuel production) figured much more prominently. Yet, while overall agricultural production de-
creased, rapeseed production grew by factor 65 between 1993 and 2008. Almost all of this production is
designated for export. In 2008, 83% of Ukraine's direct material input of rapeseed was exported. In 2009,
this share reached 99% (calculations by the authors based on data extracted from FAOSTAT (2012)).
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No. 158, September 18, 1915: The blue veil; or, Nick
Carter's torn trail
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Title: Nick Carter Stories No. 158, September 18, 1915: The blue veil;
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Author: Nicholas Carter

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CARTER STORIES NO. 158, SEPTEMBER 18, 1915: THE BLUE
VEIL; OR, NICK CARTER'S TORN TRAIL ***
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 THE BLUE VEIL;

Or, NICK CARTER’S TORN TRAIL.

Edited by CHICKERING CARTER.

 CHAPTER I.

REMARKABLE TRICKERY.

Nick Carter listened without interrupting.

The man addressing the famous detective was not one to be wisely
interrupted. His strong face, his broad, thin-lipped mouth and square jaw, the
glint of his steel-blue eyes, his portly and imposing figure—all denoted that
he was the type of man that insists upon having his way, his inning at the bat,
as it were, but who then would graciously accord the same privilege to
another.
“The danger, Mr. Carter, cannot be overestimated,” he was forcibly
saying. “It really is very terrible. We are living in constant peril. That man is
a perpetual menace. Unless he can be wiped out of existence, or put behind
prison bars, there is no telling what he might accomplish, no possible way to
anticipate it and guard against it. I cannot for the life of me understand how
he got by a detective as marvelously keen and discerning as you. I cannot,
Carter, on my word.”
Nick smiled and knocked the ashes from his cigar.
“It is not very difficult to understand,” he replied, with patience unruffled.
“There were two reasons for it, Mr. Langham.”
“Two reasons?”
“Yes. One, because the likeness between Chester Clayton and David
Margate, or Doctor David Guelpa, in which character this exceedingly clever
rascal then was posing, is a most extraordinary one. I doubt that two other
persons could be found, not excluding the most perfect of twins, who look so
precisely alike.”
“But you already knew of that extraordinary resemblance, Mr. Carter,
when Margate eluded you and made his escape.”
“Very true,” Nick admitted. “But there were other facts which I did not
know, and which I had had no way of learning. That is why there was a
second reason for Margate’s escape. Any detective, even one as ‘keen and
discerning’ as myself, if I may quote you, would be deceived by a seeming
impossibility.”
 “Impossibility?”
“Seeming impossibility,” corrected Nick.
“What do you mean?”
“Bear in mind, Mr. Langham, that Margate rushed from the house in
which we secured his confederates and ran to his suite in the Hotel Westgate,
of which Clayton still is manager.”
“I know about that.”
“I then did not know that a secret electric communication existed between
the very room in which we made the arrest and the apartments to which
Margate had gone, nor that a signal informing him of the arrest and warning
him to flee could be communicated to him by stepping on a concealed button
under the carpet. I since have learned all about that. That was done by
Scoville, one of the arrested crooks, unknown to me and my assistants.”
“But, Mr. Carter——”
“One moment, please,” Nick now interrupted. “I want you to see how
impossible Margate’s exploit must have appeared.”
“Go on, then.”
“Only ten minutes elapsed from the time Margate left his confederates,
until I entered the Westgate in pursuit of him. The first person I saw in the
hotel office was, I supposed, Manager Clayton.”
“Well?”
“How could I believe anything else?” Nick went on more earnestly. “He
was in the office inclosure and wearing an entirely different suit from what
Margate was wearing ten minutes before. Ten minutes is an incredibly short
time in which to have covered the distance between the two houses, to have
gone to his suite and changed his outside garments and got down to the hotel
office.”
“I admit that, Carter, of course.”
“I called to the supposed Clayton, therefore, and we went up to Margate’s
suite, in company with my junior assistant, Patsy Garvan,” continued Nick.
“We found the supposed Margate unconscious on his bed, clad in the same
suit in which I had seen him, as I have said, only ten minutes before. Who on
earth would have suspected, despite the extraordinary resemblance and all
that previously had occurred, that such a lightninglike change of character
could be accomplished; that the man on the bed was Clayton, and the man at
my elbow was the crook himself? It would have seemed incredible, utterly
impossible. That is why I did not give it a thought.”
“How was it accomplished, Mr. Carter?”
“I since have learned, of course,” said Nick. “Margate received the
warning signal the moment he entered his suite. He instantly telephoned
down to the hotel office and requested Clayton to come up there
immediately on important business.”
“He did so?”
“Certainly. Clayton had no occasion to suspect Margate, whom he knew
only as Doctor Guelpa. He complied, of course, and Margate invited him to
his suite. Then, passing back of him, he threw one arm around his head and
over his mouth, at the same time injecting into his neck a quantity of the
same swiftly acting drug with which he had overcome Patsy Garvan earlier
in the evening.”
“Clayton has told me about that.”
“It was done in a couple of minutes,” Nick went on. “Margate then
stripped Clayton of his outside garments, exchanging them for his own, and
placed his senseless form on the bed.”
“But what motive had he?” questioned Langham. “Why did he not flee at
once after receiving the warning?”
Nick laughed a bit derisively.
“You don’t know this rascal, Mr. Langham,” he replied. “I now know
more about him than I then did. He turned that trick only because he was
short of funds. He then went down to the hotel office, a human counterfeit of
Clayton, with the intention of stealing the money from the hotel vault.”
“Ah, I see,” Mr. Langham nodded. “A rascal, Carter, indeed.”
“My timely arrival with Patsy at just that moment prevented his design,”
said the detective. “He had no sane alternative, when I called to him, but to
accompany us to the suite. My assistant then made a hurried examination of
the man on the bed, and he at once inferred that Margate had committed
suicide.”
“I suppose it appeared so,” Mr. Langham allowed.
“In the meantime,” Nick added; “the supposed Clayton cried that he must
telephone the good news to his mother and to Mademoiselle Falloni, whose
stolen jewels we had just recovered. He hurried from the room, as if to do so.
We now know that he hurried from the house, and that is the last we saw of
him. But the whole business from beginning to end occurred in less than
fifteen minutes, Mr. Langham, and no detective on earth, unless gifted with
clairvoyance, would have suspected the trick.”
“I admit, of course, that it would have seemed impossible,” bowed
Langham.
“Now, sir, let me tell you what I since have learned about this crook,”
said Nick. “I have looked up his record abroad. He twice had been convicted
and sent to prison. He at one time was associated in Paris with the notorious
Doctor Leon Deverge, who was executed two years ago for wholesale
murder by means of drugs and poisons, of which he had made so profound a
study that he knew much more of their subtle and deadly qualities than has
been learned by any of his contemporaries.”
“I remember having read of the man.”
“This notorious physician and chemist imparted to David Margate much
of his dangerous knowledge, and the career of the latter has always been one
of vice and crime. It has been accomplished with such exceeding craft and
cunning, moreover, that he most of the time has completely baffled the
police. I admit that Margate is a terrible menace to society and to——”
“To us, Mr. Carter, in particular,” said Mr. Langham, interrupting. “For he
threatened Clayton by letter many months ago that he would wreak
vengeance upon him for having put you on his track, and that your life
would be the price for having foiled him and imprisoned his confederates. In
view of all this, Carter, and particularly his extraordinary likeness to
Clayton, his very existence is a constant menace.”
“Those are the only reasons, Mr. Langham, why I consented to drive up
here into the Berkshire Hills with my assistants to attend these festivities,”
Nick again interposed.
“That was very good of you, Mr. Carter, to be sure,” bowed the other.
“I was pleased, of course, to be present at the marriage of Clayton and
your daughter, and both assured me that they would feel easier if I was
here,” Nick added. “Clayton apprehended that Margate, despite that he has
not been seen or heard from save once since his jewel robbery, might attempt
knavery at this time. I attribute that, however, to Clayton’s somewhat
nervous temperament. I don’t take very much stock in the threats of crooks,
you know, for I long have been accustomed to them. Very few of them ever
make good. I doubt that David Margate ever will.”
“Well, I hope not, I’m sure.”
“It is nearly time, I think, for Clayton and his bride to depart,” Nick now
said, glancing at his watch. “You will wish to see them leave, I suppose.”
It then was ten o’clock in the evening, that of a bright day in June—a fit
day, indeed, for the marriage of as beautiful a girl as charming Clara
Langham, the only daughter of the multimillionaire president of the Century
Trust Company, with whom Nick Carter had been talking.
More than six months had passed since the extraordinary case they had
been discussing, that involving the theft and recovery of the world-famous
jewels of Mademoiselle Falloni, the celebrated prima donna, a case resulting
also in the arrest and conviction of all of the crooks save their ringleader,
whose unparalleled elusion of Nick Carter at the last moment they had been
reviewing.
Nick never had confided, not even to his trusty assistants, the terrible
secret intrusted to his keeping by Clayton’s cultured and attractive mother;
that his extraordinary personal resemblance to the notorious crook was due
to his twin relationship; that he bore his mother’s maiden name, and David
Margate that of the criminal father of both, who had deserted his wife in
England while the children were infants, taking with him this son, who
afterward fell naturally into the evil footsteps of his vicious father, who since
had died under sentence in a German prison.
Nick would not have thought of betraying such a secret, of which Clayton
was entirely ignorant, and the disclosure of which would serve only to mar
his happiness and in a measure wreck his subsequent life.
The secret then was known, in fact, only by Nick and the sad-hearted
mother, Mrs. Julia Clayton, who had confided it to him only in order that the
detective might prove Clayton innocent of the great jewel robbery
mentioned. It was a secret that could be safely trusted to a man of Nick
Carter’s sterling integrity.
The room in which he then was seated was the private library of Mr.
Gustavus Langham, in the money magnate’s great stone mansion, occupied
only as a summer residence. It had been built several years before at an
enormous expense, before the death of his gay and fashionable wife.
 It was like an old feudal castle, with its massive walls and parapets, its
broad halls and winding stairways, its stately rooms and attractive
surroundings, covering a vast wooded estate in one of the most picturesque
and secluded sections of the beautiful Berkshire Hills.
From the room in which Nick was seated could be heard, though the door
was closed, the strains of the orchestral music, also the vivacious
conversation and gay laughter of a multitude of guests, gathered at the
wedding reception by a special train from New York, or with motor cars
from select summer colonies from a radius of fifty miles.
The driveways and roads through the vast estate of nearly a square mile
were alive with moving conveyances of one kind or another, some of the
guests residing at a distance already having made their departure.
For the wedding ceremony had been performed two hours before, the
reception was nearing its end, and the bride and groom were making final
preparations for a precipitous departure to avoid the customary good-luck
shower on such occasions.
Mr. Langham also drew out his watch and glanced at it.
“Nearly ten,” he remarked, replying to the detective. “Why, yes, I
certainly wish to see them leave. I also want a last word in private with
Clara. I will go and see her before she leaves her room. I told her I would do
so about this time. She is expecting me, no doubt, and——”
But Mr. Langham, who had arisen while speaking, got no further with his
remarks.
He was interrupted by the unceremonious opening of the door and by the
hurried entrance of Clayton’s best man, George Vandyke, a New York
lawyer with whom Nick Carter was very well acquainted.
One glance at the young man’s white face and dilated eyes was enough to
convince the detective that something both alarming and extraordinary had
occurred.
“Out with it, Vandyke,” he exclaimed, starting up and dropping his cigar
into the cuspidor. “What’s the matter with you? What has happened?”
 CHAPTER II.

THE STOLEN BRIDE.

Nick Carter evidently was the man George Vandyke was seeking. He
appeared unable to speak for a moment, nevertheless, so great was his
suppressed excitement.
“I’ve been looking for you,” he finally gasped, when Nick seized him by
the arm and shook him. “They told me you were here. I——”
“Out with it!” repeated Nick more sharply. “What’s the trouble?”
“Clayton has disappeared,” choked Vandyke. “He cannot be found. His
bride also is missing. Neither of them are in their rooms, nor——”
“Good God! Has the blow fallen?”
Mr. Langham staggered as if he had, indeed, received a brutal blow.
Nick Carter immediately took the ribbons.
“Don’t create a stir!” he commanded quickly. “Leave me to look into the
matter. Since both are missing, they may have departed together, bent upon
eluding their very zealous friends and a deluge of confetti.”
“That cannot be, Nick,” Vandyke hurriedly protested. “Clayton’s suit case
is still in his room. He would have taken it with him, of course, if he——”
“Leave it to me. Don’t alarm the guests needlessly.”
“But some of them already know——”
Nick did not wait for more. He brushed by the two men, and, outwardly
perfectly calm, hastened through the crowded hall toward the main stairway.
Both Chick Carter and Patsy Garvan then were on the main floor of the
vast house, the former near the open front door, where, both in the hall and
on the granite steps and the broad veranda outside, scores of guests had
gathered to speed the happy couple on their wedding journey.
Chick saw Nick approaching and caught the ominous gleam in his
expressive eyes.
“What’s up?” he asked quietly, hastily meeting him.
Nick now said what he really thought.
“That devil has got in his work again.”
 “Not Margate?”
“I fear so. Both bride and groom are missing.”
“The deuce you say!”
“Nothing could have been pulled off, however, under the eyes of this mob
on the steps and veranda. Slip around to the side door and see what you can
learn,” Nick hurriedly directed. “Keep your eyes open and nail any one
acting suspiciously. Get word to Patsy and send him to the rear door. The
trick may not have been turned yet. They can have been missing only a few
minutes.”
“I’m wise,” Chick nodded, starting for the side hall and the broad exit
under the massive porte-cochère.
Nick hastened to the second floor and toward the two rear rooms used by
the bride and groom that evening, those in front having been needed to
accommodate the throng of guests.
Nick discovered a solitary bridesmaid near the door of Clara’s room, and
somewhat apart from the group of women then near the stairs. She happened
to be one with whom he was acquainted, and he hurriedly approached her.
“What’s this I hear, Miss Arden?” he said quietly. “What do you know
about it?”
“Little enough, Mr. Carter,” she replied, pale and mystified. “I only know
that Clara sent us all from her room after she was dressed for her journey.
She explained that her father wanted to see her privately before she left, and
that she was momentarily expecting him. We left her alone, therefore, and
went downstairs.”
“You mean yourself and the other bridesmaids?”
“Yes, sir.”
“How long ago was that?”
“Not more than ten minutes; hardly as long, I think.”
“Who discovered her absence?”
“I did. I returned to get my handkerchief, which I had left in the room. I
found the room deserted. Clara had gone, but her suit case and hand bag still
are there. I came out, of course, and I at once saw Mr. Vandyke coming up
the side stairs. I told him about it, Mr. Carter, and he said that Clara probably
was with Mr. Clayton in his room. He knocked, but received no answer. He
then went in and found that Mr. Clayton also was missing.”
 “Did you make any inquiries among the guests here in the hall?”
“Yes, immediately. We could find no one who had seen either of them go
out. Strange though it seems, both of them have mysteriously disappeared,
leaving their luggage in their rooms.”
“You say that Miss Langham, or, rather, Mrs. Clayton, was clad in her
outside garments?”
“Yes, sir. She had on her hat, veil, and jacket, and was ready to leave at
any moment.”
“What is her traveling costume?”
“A navy-blue suit with hat and veil to match.”
“Who, now, is in her room?”
“No one, Mr. Carter. She——”
“Wait!” Nick interrupted. “I will look in there.”
He stepped into the room while speaking. It was in considerable disorder
after the change of attire from a wedding gown to a traveling costume. There
was no sign of the missing girl, no written line explaining her sudden
departure, no evidence of when, why, or how she had gone. Both windows
were open, but in each there was a wire screen secured on the inside. Nick
saw plainly that neither of them had been tampered with.
“By Jove, this looks bad enough. It looks, indeed, as if Dave Margate has
again got in his work,” he said to himself while retracing his steps. “Has the
rascal designs upon this girl, disregarding the valuable gifts now in the
house? Those were safely guarded from every side, but who would have
thought it necessary to guard her in such a throng as this?”
“What do you think about it, Mr. Carter?” questioned Miss Arden, awed
by the more serious expression on the detective’s face when he came from
the room.
“I cannot say at present,” Nick replied. “Don’t be alarmed, nor spread the
news too quickly. There still is a possibility that they will return.”
He did not wait for an answer, but hastened into an opposite room, that
occupied by Chester Clayton.
There Nick found, at first, the same negative conditions. A single window
overlooked the rear grounds. It was closed and locked. Clayton’s suit case
stood near the door. His overcoat and hat were missing, however, though a
pair of new kid gloves lay on the dressing stand.
 Nick had only time to note these features of the scene when Vandyke
hurriedly entered, looking even more pale and disturbed.
“Why did you apprehend so quickly that something was wrong?” Nick
asked a bit abruptly, turning to him.
“Only because Clayton appeared to fear some mishap,” Vandyke replied.
“He admitted he had no definite reason for it, but he seemed very nervous.”
“Where were you when he left? You were his best man.”
“True. I came here to tell you about that.”
“About what?”
“One of the caterer’s assistants came in here a short time ago, not more
than twenty minutes, and stated that Mr. Lenaire wanted to see me in the
dining room.”
“Lenaire is the caterer?”
“Yes. It was upon my recommendation that he was given this job. I asked
Clayton if he had any immediate use for me, and he told me to go ahead and
see what Lenaire wanted. I did so and found him in the dining room.”
“What did he want?”
“He wanted to thank me again for having recommended him, and also to
ask me to express his gratitude to Clayton for having seconded my
suggestion, which he feared he would not have an opportunity to do
personally before Clayton departed. He explained at some length, Nick, and
when I returned I found that Clayton was missing. Then, when unable to find
Clara, I feared something was wrong.”
“I see,” Nick nodded. “Did the waiter who came up return to the dining
room with you?”
“No, not with me,” said Vandyke. “I hurried down ahead of him. I did not
see him again.”
“Do you know his name?”
“I think Lenaire called him Toulon.”
“By Jove, I think I scent the rat in the meal,” Nick muttered. “Have you
looked in the closet, Vandyke?”
“Not yet. Who would expect to find Clayton in the closet, or concealed in
any part of the room? It would be absurd to suppose anything of the kind
——”
“Not absurd to me,” Nick suddenly interrupted. “See for yourself.”
 He had, while Vandyke was speaking, looked hurriedly into the wardrobe
closet and under the bed. A broad, old-fashioned couch near one of the walls
then claimed his attention. It was draped with a valance, which he quickly
raised, and then he found what he was seeking.
Flat on his back under the couch lay the senseless form of Chester
Clayton, his eyes closed and his white face upturned, as ghastly as if the
hand of death had been laid upon him.
Vandyke recoiled with a shudder.
“Good heavens!” he cried. “Is he dead? Is he dead, Mr. Carter?”
“Quiet,” Nick cautioned. “No, not dead. The rascal who did this job
doesn’t thrust his knavish neck into a noose. Clayton has been drugged. It’s
the work of the same miscreant who downed him at the time of the jewel
robbery.”
“David Margate?”
“Yes.”
“What shall we——”
“Don’t stop to question,” Nick interrupted. “Lend me a hand and we will
place him on the couch. Slip out and find a physician, if there is one among
the guests. Don’t alarm them, however, by stating what has occurred. A
physician soon can revive him. Send Mr. Langham in here, but not a word
about this to Mrs. Julia Clayton. Leave me to inform her.”
“You think——”
“Never mind what I think,” Nick again cut in while they placed the
senseless man on the couch. “Do what I have directed.”
“But Clara, his wife—what of her?”
“There’s nothing to it, Vandyke,” said the detective. “It’s as plain as twice
two. The bride has been stolen.”
 CHAPTER III.

THE ASSAULTED WAITER.

Chick Carter, hastening to follow Nick’s instructions, found nothing in

the side hall nor out-of-doors that shed any light on the mystery.
Several guests were departing in a limousine from under the porte-
cochère, but Chick knew two of them personally and that none was worthy
of the slightest suspicion.
Returning through the hall, he found Patsy Garvan and quickly told him
what had occurred, while both hastened out of the rear door of the house. As
they were descending the steps, one of the kitchen servants, who was on her
way in, approached them and said somewhat excitedly, addressing Chick:
“Sure, sir, there’s something wrong around here. Would you mind telling
Mr. Langham, sir?”
“Something wrong?” questioned Chick, sharply regarding her. “Where?
What do you mean?”
“Round here, sir,” she replied, leading the way. “I was after taking out
some refuse for the barrels, sir, and I heard moaninglike, as if some one was
hurted.”
“Heard it where?”
“Here, sir, under the cellar door. I was after—there ’tis again, sir!”
The corpulent Irishwoman shrank back affrighted.
A hollow, half-choked moan had issued from under a slanting bulkhead
door abutting the foundation wall on that side of the house.
It was the opposite side from that on which was located the driveway
making around from the front of the vast stone mansion and leading out to
the stable and garage. Aside from the bulkhead door leading down to the
basement there was only another door opening upon an entry and stairway
for the use of the servants.
The adjoining grounds in that locality were deserted, and lighted only by
the stars glittering in the purple sky. A path led across a strip of lawn to
several outbuildings. Beyond this were the trees of the park and woodland
covering the vast estate. Through the gloom beneath them some fifty yards
away could be faintly seen a gray gravel driveway making off to the east.
Patsy caught sight of something white on the ground, just as the hollow
moan interrupted the woman, and he stopped to pick it up.
It was a partly burned cigarette, yet from which only a few puffs had been
taken.
Instinctively Patsy slipped it into his pocket, just as Chick exclaimed:
“By Jove, the woman is right. Lend me a hand, Patsy. This door is not
locked. Here’s a man on the stone steps.”
His words evoked another moan from the prostrate man.
“Wait a bit!” said Patsy. “Here is my searchlight.”
Chick had opened both sections of the slanting door, and Patsy now sent a
beam of light down the several stone steps. In the area below, against an
inner door of the cellar, lay a man in evening dress, bound hand and foot
with stout cords and brutally gagged.
“Gee whiz!” cried Patsy. “Something wrong, Chick, is right.”
“Help me lift him out.”
“Lord save him!” said the woman, crossing herself. “Is he dead, sir?”
“Far from it,” said Chick. “Dead men don’t moan. He’ll be all right when
he can breathe freely. Now, sir, speak for yourself. How came you in this
mess?”
The two detectives had placed him on the greensward outside of the
bulkhead door, and Chick had quickly cut his bonds and removed the gag
from his mouth.
The man choked and gasped convulsively for a moment, then explained
with an effort that he was Pierre Toulon, employed as a waiter by Mr. Jean
Lenaire, the French caterer; that he had stolen out a short time before to
smoke a cigarette, and that he had been suddenly assaulted by three masked
men, who had bound and gagged him, and then confined him under the
bulkhead door.
Chick did not wait to look more deeply into the man’s story, but turned to
Patsy and said hurriedly:
“Go tell the chief. You’ll find him on the second floor, probably in
Clayton’s room. I will help Toulon into the house. Nick will question him
later.”
 Patsy hurried away without replying.
He found Nick, Mr. Langham, and two physicians in Clayton’s room. The
latter had begun to revive from the effects of the drug. He already could talk
intelligently, and in a vague way could recall and state what had occurred.
It appeared, Nick already had learned, that the same waiter who had
called Vandyke from the room, or a man so closely resembling him that
Clayton detected no difference, returned almost immediately after Vandyke
departed, saying that he missed his cuff link and thought it might have
dropped on the floor.
Clayton naturally had bowed to look for it, whereupon the rascal instantly
threw one arm around his head, covering his mouth, and at the same moment
thrust the needle of a hypodermic syringe into his neck, injecting a quantity
of the same potent and quick-acting drug with which, Nick immediately
suspected, Clayton had been overcome by Margate at the time of his escape
after the jewel robbery.
Clayton knew nothing of what had followed, having quickly lost
consciousness, and Nick now left Mr. Langham and the physicians to
enlighten him with the sad information. He withdrew with Patsy and
hastened down to the private library in which he had been talking with
Langham only a few minutes before.
Patsy already had told him about finding the waiter, Toulon, and Nick’s
next move was to send for Mrs. Julia Clayton, whom he briefly informed of
his suspicions, and then cautioned the dismayed woman against
inadvertently betraying the secret she so long had kept from all the world.
The shocking news now was generally known, and the house was in
confusion. Guests were hurriedly departing, leaving sympathetic messages
with the butler and other servants. All keenly felt that they could be of no
assistance in the investigations then in progress, and that they were better out
of the way.
“Gee whiz! there’s nothing to this, chief,” commented Patsy, turning after
closing the door upon Mrs. Clayton. “This is Margate’s doings, all right.”
“Undoubtedly,” said Nick. “He served Clayton the same trick as before.”
“Surest thing you know.”
“We will try later to find out how he got away with the girl. It would be
useless to undertake it at present, and immediate pursuit is out of the
question. A hundred conveyances have left here during the past half hour.”