Bioenerg. Res.

(2013) 6:851–863
DOI 10.1007/s12155-013-9341-4

Environmental Advances Due to the Integration of Food

Industries and Anaerobic Digestion for Biogas Production:
Perspectives of the Italian Milk and Dairy Product Sector
Deborah Traversi & Silvia Bonetta & Raffaella Degan &
Silvia Villa & Arianna Porfido & Monica Bellero &
Elisabetta Carraro & Giorgio Gilli

Published online: 6 June 2013

# Springer Science+Business Media New York 2013

Abstract Anaerobic digestion (AD) is a biotechnology that is
able to produce an energy vector from waste organic mate-
rials. Its inclusion in the food industries, especially if farming
activities are present, represents an interesting perspective.
This work reviews the main aspects of the inclusion of AD
in milk and dairy production. It includes an analysis of the
sector characteristics in terms of diffusion, a discussion
of waste production and disposal concerning milk and
dairy production, a description of the biological and
technological aspects of AD, a discussion regarding
the application of AD for milk by-products, a descrip-
tion of environmental and legislative aspects, and the
suggestion of an integrated model that includes the
anaerobic co-digestion of whey and cattle slurry in the
milk and dairy production industries. Both environmen-
tal and economic impacts of such integration promise to
Highlights Anaerobic digestion is a biotechnology that is able to
produce biogas. Renewable energy production can be a strategic
advantage for the dairy industries. Food producers in milk and dairy
sector generate a large amount of organic refuses. Whey and cattle
slurry can be coupling substrates for biogas digestion. The benefits
coming from the integration promise to pay for the investment.
D. Traversi (*) : R. Degan : S. Villa : A. Porfido : G. Gilli
Dipartimento di Scienze della Sanità Pubblica e Pediatriche,
Università di Torino, via Santena 5 bis,
10126 Turin, Italy
e-mail: deborah.traversi@unito.it
S. Bonetta : M. Bellero : E. Carraro
Dipartimento di Scienze ed Innovazione Tecnologica, Università
del Piemonte Orientale A. Avogadro, Viale T. Michel 11,
15121 Alessandria, Italy
pay for the preliminary investment in AD plant instal-
lation and the development of additional expertise.
Keywords Anaerobic digestion . Whey . Dairy wastes .
Cattle slurry . Milk . Biogas . Renewable energy
Introduction
Milk and dairy products are essential parts of a healthy diet.
Their consumption is encouraged by World Health Organi-
zation (WHO) guidelines with consideration to limitations
for fat and salt intake [1]. Milk is a nutrient concentrate for
humans, and, during the last several decades, various im-
provements to its health characteristics have been made.
Some examples are the development of lactose-free milk,
milk with oligosaccharide content, calcium-enriched milk,
milk with fatty concentrations, milk with n-3 polyunsaturat-
ed fatty acid content, milk with micro-nutrient concentra-
tions, and the addition of probiotics and prebiotics [2]. Milk
and milk-derived products comprise 10 % of the food pro-
duction sector in Europe. Italy is one of the top five main
European producers, with production comparable to the
Netherlands [3]. Italian production and direct utilisation at
the farm level have shown a 3 % increase in the last decade
(1999–2009). This evidence is also consistent for the last
few years [4] despite the economic crisis, which gives us an
idea of the strategic role of this food production sector.
Despite the popularity of milk and milk products, the
notifications recorded by the European Rapid Alert System
for Food and Feed (RASSF) for milk and milk products affect
at least 2.3 % of the total. The most frequent hazard categories
852 Bioenerg. Res. (2013) 6:851–863

are the presence of potentially pathogenic microorgan- The Dairy Sector in Europe
isms and the presence of various contaminants, includ-
ing food additives, residues from veterinary products, The vast majority of milk collected in the EU in 2006 came
and foreign bodies [5]. In the last decade, the introduc- from cows, although in a number of the southern European
tion of standards [6] and hazard assessment regulations Member States, significant quantities of milk were also pro-
[7] for the full production process has led to a marked trans- duced from sheep and goats. There were 2.8 million holdings
formation of the organisation, with the adoption of effective in the EU-27 with dairy cows. However, 1.9 million holdings
food safety management systems [8]. In Italy, particularly in had only one or two cows, with Romania and Poland account-
some areas, this approach has been rapidly developed, taking ing for 1.6 million of these small dairy units. Among member
advantage of traditional hygiene, veterinary, and food- states, the number of farms with dairy cows declined sharply
production capabilities. In 2010, Italy was the top European between 1995 and 2005; in Italy, the number of holdings with
country for typical food production. More than 18 % of typical dairy cows was halved, and the number fell by almost two-
products, classified as Protected Designation of Origin (PDO), thirds in Spain [4]. Although the number of cows also de-
protected geographical indication (PGI), or traditional speci- clined in the same period, the average number of cows per
ality guaranteed (TSG) by the European Union, are dairy holding increased (European mean from 32 to 49), sometimes
products [9]. sharply (e.g., in Denmark, Greece, and Portugal). In contrast
These data confirm not only the skilled history resulting to the small herd sizes in Romania and Poland, the average
from centuries of work, but also the improvements to tech- size of a dairy herd in the United Kingdom was just under 80
nologies provided with modern tools to control hazards and head in 2005, had risen to 85 head in Denmark, and was just
improve the efficiency of production systems. Nevertheless, over 100 head on average in Cyprus.
the global trade system has garnered various criticisms. The annual milk production in various European coun-
Among these criticisms are business competition from tries (EU-27) is shown in Table 1: the production of cows’
producers in other parts of the world where production milk remained fairly steady. Average milk yields across the
costs are lower, changes in transportation overheads and in EU-27 in 2010 were about 13.5 × 107 tonnes; Germany is
particular major expenditures due to energy consumption, and the country with the highest milk production (21 %)
the increasing attention to environmental sustainability. Euro- followed by France (17.2 %). Approximately 90 % of milk
pean regulations, especially those promulgated in the last few produced in the EU-27 was collected by dairies for processing
years, have had the objective of reducing the environmental into drinking milk and a variety of milk products [3]. The 133
impact of farming and food production activities through million tonnes of milk delivered to dairies across the EU-27
the integration of waste disposal and energy production. were principally transformed into 31 million tonnes of drink-
Some examples are directives concerning the protection ing milk, 9 million tonnes of cheese, 7 million tonnes of
of waters against pollution caused by nitrates and more acidified milk, 2 million tonnes of butter and 1 million tonnes
recently [10] on the Common Agricultural Policy (CAP) of both skimmed milk powder and other milk powder. In
towards 2020, where it is established that “…the overarching
objective for the future CAP should be sustainable competi- Table 1 Cows’ milk collection in EU-27 countries in 2005 and 2011
tiveness to achieve an economically viable food production [4]
sector, in tandem with sustainable management of the EU’s Delivery of cow milk (tonnes×1000) EU-27
natural land-based resources.” Milk and milk by-products fall
under this last point as a great prospective field. Country 2005 2011
AD is one of the renewable technologies for the pro-
Germany 27,311 29,339
duction of green energy; from a plant diffusion point of
France 23,325 24,607
view, this technology is most likely the first. In this
United Kingdom 14,038 13,804
review, we analyse and discuss the environmental impli-
Netherlands 10,407 11,627
cations of integration between milk and dairy production
and AD technologies from an environmental and an eco- Italy 10,127 10,260
efficiency point of view. The main argument is the pro- Spain 5,914 5,950
duction of biogas as an energy vector, first to cover the Ireland 5,061 5,536
energy necessities of production processes and second to Portugal 1,911 1,837
even generate an energy surplus that is re-investable into Othersa 34,306 34,762
the production system for the improvement and the mod- TOTAL EU-27 132,400 137,722
ernization of the production chain. Then, the environmen- a
Austria, Belgium, Denmark, Finland, Greece, Luxembourg, Sweden,
tal impact in terms of polluted waste production is also Cyprus, Czech Republic, Estonia, Hungary, Latvia, Lithuania, Malta,
discussed. Poland, Slovakia, Slovenia, Romania, Bulgaria
Bioenerg. Res. (2013) 6:851–863
Fig. 1 Percentage of milk used in dairy products respect to the total of
raw milk produced in the EU (estimate) [4]
terms of milk equivalent quantities, however, the European
Commission has estimated that more milk was designated for
the production of cheese (Fig. 1) than any other dairy product
in 2006. There were approximately 5,400 dairies across the
EU-27 (excluding Bulgaria, Cyprus and Luxembourg), and
approximately 30 % of these were in Italy, while a further
30 % were in Greece, Spain and the United Kingdom com-
bined. There were more than 13,000 dairy product manufactur-
ing enterprises (including dairies) in the EU-27 in 2005,
employing approximately 400,000 people. One-third (33.1 %)
of the dairy product manufacturing enterprises in the EU-27
were located in Italy, with a further fifth in Spain (11.5 %) and
France (11.2 %) combined. A little over half (53.6 %) of the
turnover generated by the EU-27’s dairy product manufacturing
enterprises came from enterprises in France (20.0 %), Germany
(19.2 %) and Italy (14.3 %). Further down the chain, there were
approximately 15,000 wholesalers specialising in dairy pro-
duce, eggs, edible oils and fats in the EU-27, a majority of
which combined were based in Italy (24.0 %), Spain (18.8 %)
and Greece (12.5 %). These specialist wholesalers employed a
further 119,000 persons, representing 1.2 % of the wholesale
and commission trade workforce within the EU-27.
The observed transformation in favour of large plants for
farming and milk-based production resulted in the ability to
collect a wider amount of waste and produce refuse, and
moreover resulted in a large range of floating investment
assets. The economic resources allocated as developing funds
for a large enterprise could more easily distinguish medium-
Table 2 Milk production in Italy—details for geographical areas in 2010 (quantity in metric tonnes), elaborated from ISTAT [9]
Geographical areas Cow milk Sheep milk
North Italy 8,866,909 3,500
Central Italy 644,490 116,809
South Italy 1,061,783 311,913
Total 10,573,182 432,222
853
and long-term growth instead of immediate feedback. There
were approximately 13,000 enterprises across the EU-27
whose main activity was the manufacture of dairy products
(NACE Group 15.5) in 2006. These enterprises employed an
estimated 400,000 persons, representing 8.5 % of the work-
force of the food, beverage, and tobacco manufacturing sector.
The overwhelming majority of these workers (83.9 %) were
engaged in the operation of dairies and cheese-making
(NACE Class 15.51). The dairy product manufacturing sector
generated € 17.7 billion of value added in 2005, equivalent to
8.9 % of the value added generated by food, beverage, and
tobacco manufacturing activities in 2005 [11].
The Dairy Sector in Italy
According to data from the 6th Census of Agriculture in Octo-
ber 2010, there were 1,630,420 active farms and livestock in
Italy. Puglia is the region with the largest number of farms (over
275,000), followed by Sicilia (219,000), Calabria (138,000),
Campania (137,000) and Veneto (121,000). In these five re-
gions are found 54.6 % of Italian farms [9]. Of the total utilised
agricultural area, 46 % is concentrated in Sicilia (1,384,043 ha),
Puglia (1,280,876 ha), Sardegna (1,152,756 ha), Emilia-
Romagna (1,066,773 ha) and Piemonte (1,048,350 ha). The
regions with the greatest numbers of bovine livestock are
Lombardia, with 15,000 holdings and 1.5 million animals,
Veneto with 13,000 holdings and 826,000 cattle, and Piemonte
with 13,000 holdings and 816,000 cattle. Taken together, these
three regions contain approximately 55 % of Italian cattle. In
the last decade, there has been strong growth in the buffalo
farming sector, with a complex of 358,000 buffaloes concen-
trated mainly in Campania (261,000 buffaloes, 1,406 holdings)
and Lazio (63,000 buffaloes, 590 holdings). These two regions
contain 90.4 % of the total buffaloes in Italy. The overall
number of companies operating in the dairy sector is 2,149, of
which 411 are located in Emilia-Romagna, 369 in Campania,
and 259 in Lombardia. In 2010, 11,207,796 tonnes of milk
were produced overall: 10,573,181 tonnes of cow’s milk,
432,222 tonnes of sheep’s milk, 177,457 tonnes of buffalo milk
and 24,935 tonnes of goat’s milk. The regions with the highest
milk production are mainly located in North Italy (Table 2);
Buffalo milk Goat milk Total
4,482 10,152 8,885,043
27,234 1,736 790,269
145,741 13,046 1,532,483
177,457 24,935 11,207,796
854
these are Lombardia (36 % of the total), Emilia-Romagna
(19 %), Veneto (9 %) and Piemonte (8 %) [9].
Dairy Sector By-products
The cheese manufacturing industry generates large amounts
of high-strength wastewaters with different pollution char-
acteristics such as high organic matter; moreover, the dis-
posal of effluents may cause serious environmental pollu-
tion, depending on the plant and production type [12]. The
dairy industry generates a relevant environmental impact,
for example in terms of the volume of effluent generated.
This industry generates approximately 0.2–10 litres of ef-
fluent per litre of processed milk [13], with an average
generation of approximately 2.5 litres of wastewater per litre
of milk processed [14].
A typical European dairy generates approximately 500 m3
of waste effluent daily [15]. Wastewaters from the dairy
industry are usually generated in an intermittent way, such
that the flow rates of these effluents change significantly. High
seasonal variations are also frequently encountered, and cor-
relate with the volume of milk received for processing, which
is typically high in summer and low in the winter months.
Moreover, because the dairy industry produces different prod-
ucts, such as milk, butter, yoghurt, ice cream, various types of
desserts, and cheese, the characteristics of these effluents also
vary greatly, depending on the type of system and the methods
of operation used [16]. The use of acid and alkaline cleaners
and sanitizers in the dairy industry additionally influences
wastewater characteristics, and typically results in a highly
variable pH [12]. Dairy wastewaters are characterised by high
biological-oxygen demand (BOD) and chemical-oxygen de-
mand (COD), but the main contributors to the organic load of
these wastes are carbohydrates (mainly lactose), proteins, fats
(mainly as triglycerides), and nutrients, as well as detergents
and sanitising agents [17].
In addition to the products of industrial processing, the
dairy sector produces the following by-products: (1) by-
products such as whey and buttermilk, (2) waste processing
(e.g., cheese rinds and pulp, scrap, residues of curd, residues
from cleaning), (3) milk and cheese with the presence of
Fig. 2 Dairy products
produced in Italy (tonnes×
1000) [9]
Bioenerg. Res. (2013) 6:851–863
contaminants (e.g., aflatoxins, residues of inhibitory sub-
stances), (4) waste arising from brushing the powdery form
of cheese during ripening, and (5) effluent and process waste-
water washing. In particular, whey, a greenish-yellow and
turbid liquid that remains in the boiler after separation of the
curd (a coagulation process required for all dairy products), is
a by-product of the dairy industry; its principal components
are lactose, proteins and mineral salts [18]. Approximately
47 % of the 108 tonnes of whey produced world-wide every
year are disposed of in the environment [19, 20]. This repre-
sents a significant loss of resources, and causes serious pollu-
tion problems because whey is a high-strength organic pollut-
ant with high BOD5 and COD values of 40,000–60,000 mg/l
and 50,000–80,000 mg/l respectively [21]. Figure 2 shows the
products created by the dairy industries in Italy and the desti-
nation of the main by-product, i.e., whey. Approximately
47 % of the whey produced should be disposed of (Table 3).
Depending on the mode of coagulation for the milk, whey
can be sweet (pH=6–7) or sour (pH<5). Acidic whey has a
higher ash content, especially calcium, lower serum protein
content, and lower lactose content. Sweet whey is obtained if
milk is coagulated by proteolytic enzymes such as chymosine
and pepsin or microbial enzymes produced from the moulds
Mucor miehei and Mucor pusillus. Sweet whey comes from
the production of cheddar, Swiss, and Italian varieties of
cheese. Acidic whey (pH<5) results from processes using
fermentation or the addition of organic or mineral acids to
coagulate the casein, as in the manufacture of fresh cheese or
most industrial casein [22]. It also comes from soft cheese
production, of which cottage cheese is a predominant product.
A summary of data obtained from the literature for the general
properties of whey as an effluent from full-scale operations is
shown in Table 4, where a synthesis of the literature data is
presented [23–25].
Anaerobic Digestion of Whey
Dairy wastewaters are treated using physico-chemical and bio-
logical treatment methods. However, because reagent costs are
high and soluble, COD removal is poor in physico–chemical
treatment processes, biological processes are usually preferred
Bioenerg. Res. (2013) 6:851–863 855

Table 3 Estimate of the primary waste of the dairy sector COD concentration, and tendency to become acidified very
Type Amount (tonnes/year) rapidly, leading to acidification and the inhibition of
methanogenic activity. Whey is initially hydrolysed and
Whey disposal 2,022,000 converted to organic acids by acidogenic microorgan-
Sweet 345,000 isms, and degradation is then followed by methanation
Buttermilk 331,000 by Methanosarcina and other methanogenic bacteria
Sludge 171,400 [28]. During the AD of whey, Methanosarcina barkeri
Wastes 5,700 and Methanothrix soehngenii have been identified as the domi-
Total effluent 19,270,000 nant acetate-utilising methanogens, and Methanobacterium
formicicum as the prevalent hydrogen-utilising methanogen.
The predominant shift to Methanosaetaceae can be observed in
[16]. Among biological treatment processes, treatment in environments with acetate concentrations below 1 mM, indi-
ponds, activated sludge plants, and anaerobic treatment are cating that, under these conditions, the specific growth rate of
commonly employed for dairy wastewater treatment. A lack Methanosaeta spp. is higher than that of Methanosarcina spp
of a requirement for aeration, a low amount of excess sludge [29].. Whey addiction is also used as a bio-stimulation method
production and low area demand are additional advantages of during the AD process to analyse microbial population varia-
anaerobic treatment processes in comparison to aerobic pro- tions [30]. A number of studies have been reported in literature
cesses [12]. for the treatment of dairy wastewater with anaerobic methods.
A particular ecosystem is present in an anaerobic reactor, Demirel et al. summarised the typical reported operating con-
where several groups of microorganisms work interactively ditions for anaerobic digesters in the treatment of dairy waste-
to convert complex organic matter into biogas. In the diges- waters [23]. High-rate reactors, such as upflow anaerobic
tion process, four stages take place: hydrolysis, acidogenesis, sludge bed reactors (UASBR) [31], hybrid UASBRs, expanded
acetogenesis, and methanogenesis [26]. granular sludge bed reactors (EGSBR) [32], and anaerobic
In particular, for medium-size cheese factories that have filters [33], have been used in the treatment of dairy wastewa-
growing disposal problems and cannot afford the high in- ters. Moreover, a recently published review describes the main
vestment costs for whey valorisation technologies, physico- reactors used. After the comparison between the different treat-
chemical and/or biological treatment of effluent is impera- ment alternatives, it suggests a configuration including ultra-
tive. When disposing of whey as wastewater into a sewage filtration as pre-treatment [34].
treatment plant, it has been estimated that 50 kg of whey are
equivalent to the waste produced by 22 people every day. In
other words, a cheese plant producing 50,000 kg of whey Anaerobic Whey Digestion: Inhibition Factors
per day requires a treatment plant of about the same size as a and Co-digestion Opportunities
city with a population of 22,000 inhabitants [27].
Raw whey is known to be quite problematic for anaero- Because single-stage anaerobic reactors have experienced
bic treatment because of its low bicarbonate alkalinity, high instability or failure during the treatment of complex waste-
waters, two-stage anaerobic reactors have been operated,
Table 4 Main summarised chemical characteristics of whey as dairy with better performance in the case of complex wastewaters
waste as reported in the literature [23–25]
and sludges such as cheese whey [35]..
Parameter Value Lipid degradation and inhibition in single-phase anaero-
bic systems is frequently discussed in the literature because
COD (g/l) 68.6±3.3 lipids are potential inhibitors in anaerobic systems, a dilem-
BOD5 (g/l) 37.71±2.84 ma that can often be encountered by environmental engineers
COD/BOD5 1.83±0.05 and wastewater treatment plant operators. Lipids are slowly
TSS (g/l) 1.35±0.06 degraded because of their limited availability as a function of
Lactose (g/l) 45.9±0.88 their low solubility, so the most frequent problems reported in
Proteins (g/l) 2.71±0.05 the literature are biomass flotation in reactors, unavailability
TS (%) 5.93±0.38 of the substrate for micro-organisms within biofilms, and
VS (%) 5.61±0.36 inhibition of methanogenesis due to the presence of interme-
Fat (g/L) 9.439±1.14 diaries from the degradation of lipids [36]. During anaerobic
pH 4.9±0.27 degradation, lipids are first hydrolysed to glycerol and long
TKN (g/l) 1.12±0.01 chain fatty acids (LCFAs), which are inhibitors of
TP (g/l) 0.5±1.8×10−3 methanogenesis during digestion [37], followed by oxidation,
which produces acetate and hydrogen. Unsaturated LCFAs
856
appear to have a greater inhibitory effect than saturated
LCFAs. They strongly inhibit methane production from ace-
tate, and moderately inhibit beta-oxidation, so they should
thus be saturated to prevent lipid inhibition in anaerobic
processes [38]. Pereira et al. quantified LCFAs adsorbed to
anaerobic biomass [39]; their conclusion was that these sub-
stances inhibited acidogenesis and acetoclastic and
hydrogenotrophic methanogenesis.
High concentrations of suspended solids in dairy waste
streams can also adversely affect the performance of conven-
tional anaerobic treatment processes, particularly the most
commonly used upflow anaerobic filters. Thus, two-phase
AD processes should be considered more often to overcome
these problems that may be experienced with conventional
single-phase design applications.
Despite its advantages, however, AD is not extensively
used in the dairy industry, largely due to its slow reaction
problem, requiring longer hydraulic retention times, and its
rapid acidification [40]. This slow reaction has been overcome
by novel hybrid systems such as upflow anaerobic sludge
fixed-film bioreactors and upflow packed bed biofilters.
Lactose is the main carbohydrate in dairy wastewater,
and is a readily available substrate for anaerobic bacteria.
Anaerobic methanation of lactose needs cooperative biolog-
ical activity from acidogens, acetogens and methanogens.
Anaerobic fermentation of lactose yields organic acids, spe-
cifically acetate, propionate, iso- and normal butyrate, iso- and
normal valerate, caproate, lactate, formate, and ethanol. Two
possible carbon flow schemes have been proposed for the
acidogenic fermentation of lactose: carbon flow from pyruvate
to butyrate and lactate, both occurring in parallel [23].
The presence of high carbohydrate concentrations in
synthetic dairy wastewater has been found to reduce the
amount of proteolytic enzymes synthesised, resulting in
low levels of protein degradation [41]. It was previously
reported that carbohydrates could suppress the synthesis of exo-
peptidases, a group of enzymes facilitating protein hydrolysis.
Anaerobic degradation of proteins, and the effects of
ammonia on this mechanism, were recently investigated in
detail. Casein is the major protein in milk and in dairy
effluents. When fed to acclimated anaerobic reactors, the
degradation of casein is very fast, and the degradation
products are non-inhibitory [42].
It has been stated that raw cheese whey is a difficult
substrate to treat anaerobically due to its lack of alkalinity,
COD concentration, and tendency to acidify very rapidly.
Due to the low bicarbonate alkalinity, supplemental alkalin-
ity is required to avoid anaerobic process failure. This alka-
linity supplementation can be minimised using operational
conditions directed at obtaining better treatment efficiency,
such as higher hydraulic residence times or dilution of the
influent. Moreover, when undiluted cheese whey is directly
treated in anaerobic reactors, stability problems arise. It has
Bioenerg. Res. (2013) 6:851–863
been reported that the co-digestion of whey with manure is
possible without any need for chemical additions for up to
50 % participation of whey (by volume) in the daily feed
mixture. The combined treatment of different waste types
such as manure and cheese whey gives rise to the possibility
of treating waste, which cannot be successfully treated sep-
arately. Whey was quantitatively degraded to biogas when
co-digested with diluted manure in the absence of any added
chemicals. Manure has a high content of lipids, while whey
has a high content of easily biodegradable carbohydrates.
The co-digestion of these two wastes is more advantageous
than processing each one separately. The AD of whey can be
applied to existing facilities that are already used for manure
digestion alone. As a result, the co-digestion of cheese whey
together with local agricultural residues, such as manure, is
a sustainable and environmentally attractive method.
Energy Recovery from Cheese Whey by Anaerobic
Digestion
Due to its high biodegradability (aerobic degradabili-
ty≈99 %, anaerobic degradability 94–99 %), the biological
treatment of whey is the most appropriate method of
stabilisation, and due to the high organic content, the basic
biological treatment process used can only be anaerobic
digestion; regular treatment processes such as the activated
sludge process are completely inappropriate [43]. During
whey fermentation, most of the lactose is transformed into
lactic acid, acetic acid, and other VFAs. The majority of
lactose (62 %) is converted into VFA (5 g/l) and lactic acid
(18 g/l), and it is then transformed into methane. Addition-
ally, serum proteins are readily degraded by acclimated
sludge, and the lipid content is not sufficient to cause inhi-
bition. Biogas yields from whey range between 76 and 99 %
[44]. In batch-trials carried out to assess the biogas produc-
tivity potential of some agro-industrial biomasses, it was
found that the most productive, in terms of specific yields,
was whey. Its specific yields were 953 lN biogas kg volatile
solids−1 and 501 lN CH4 kg VS−1 [45]. Due to its high
biodegradability, high organic load, and very low bicarbon-
ate alkalinity, whey tends to acidify very rapidly, thus
impairing maintenance of the process. Therefore, to main-
tain process stability, a system to control pH is necessary,
requiring in most cases the addition of some external source
of alkalinity, such as bicarbonate, carbonate, or hydroxide
[46]. As previously discussed, a possible solution to this
problem is the two-phase plant configuration, which pro-
vides for the separation of the acidogenic and methanogenic
steps [43], while another possible alternative is the co-
fermentation of whey together with substrates with suffi-
cient buffering capacities such as cattle manure. By mixing
whey and manure, one can obtain a substrate that is easier to
Bioenerg. Res. (2013) 6:851–863
manage. Indeed, manure can balance the low nitrogen and
alkalinity content of whey and the high content of rapidly
hydrolysable substances, making the degradative chain mo-
re stable. Gelegenis et al. [47] reported a methane yield of
2.2 vm·vr−1 ·d−1 (vm: volume of methane, vr: volume of
reactor) in a continuously stirred pilot reactor containing a
diluted poultry manure and whey mixture. Mixtures in
which whey accounts for 20–50 % of the total volatile solids
were found to be optimal [47]. According to Kavacik et al.
[25], the codigestion of cheese whey and dairy manure
under continuous fermentation conditions with an HRT
(hydraulic retention time) of 5 days and 8 % of total solids
resulted in 0.969 m3·biogas for each m3 of feeding daily
added. Comino et al. [48] investigated the biogas potential
of a cow manure and whey biomass mix, and achieved
211.4 l CH4·kgVS−1 and very high rates of BOD5 (78 %)
and COD (74 %) removal. The Centre of Theoretical and
Applied Ecology of Gorizia tested various mixtures of whey
and cow manure with increasing amounts of whey (20–50 %
and 80 %) with respect to manure. The highest specific
biogas yield per unit of organic matter, equivalent to
486 l/kg of volatile substance, was obtained using a mixture
with 80 % whey and 20 % slurry. The biogas produced,
however, contained a lower percentage of methane (60.7 %)
than for mixtures with lower amounts of whey (64–
65 % methane). This fact can be explained by the
higher concentration of carbohydrates (mainly lactose)
in the whey compared to proteins or lipids, which gives
rise to a biogas that is richer in CO2. Table 5 summa-
rises some literature data on the biogas yield in relation
to the introduced COD during anaerobic digestion and
co-digestion of whey. The slurry maintained the stability
of the anaerobic process, providing adequate alkalinity
to maintain optimal pH values (pH 6.8 to 7.2) for
methanogenic bacteria. Furthermore, animal slurry, especially
cattle slurry, provides microorganisms, macro- and micro-
nutrients that help to establish and maintain a balanced bioce-
nosis in the reactor by favouring optimal conditions for the
process [49]. Considering the various aspects described
previously, we propose a diagram in which milk and dairy
product production is integrated with an AD plant (Fig. 3).
Hypothetically, existing facilities could be integrated with
Table 5 Data on current production of biogas by the anaerobic digestion of whey from the literature
Substrate
Cheese whey
Acid whey
Pre-fermented acid whey
Co-digestion (50 % whey and 50 % cattle manure)
Whey
857
such technology, or eventually it may be possible to achieve
integration among producers in the same district around a
circumscribed territory. The most abundant and also suitable
organic materials in this integrated system would be whey and
cattle slurry; manure can also be used, but there could be
problems in hydrolysing the complex molecules of straw, such
as cellulose and lignin. In such a system, the main methanogen
substrates could be ∼80 % acetate and <15 % hydrogen. The
biogas specific yield could be approximately 650 l/kg VS, with
a mean methane percentage of at least 55 % [48]. This config-
uration would produce a COD removal efficiency greater than
95 % [50].
Obviously, a wide range of variables must be evaluated.
First, the fermentable organic solids in cattle sludge vary
according to different factors, among which are the animal
species and weight production class, the type of animal feed
and the efficiency of the feed transformation, the topology
of the cattle-pound, and the quantity of the washing or
meteoric water collected with the stools and urine. However,
the mean amount of cattle slurry produced per year is
approximately 54 m3/head. Thus, cattle slurry disposal
could be transformed from a problem to a resource and from
a cost to a gain. The gain in terms of biogas production for
each head could be estimated at 500€. This gain does not
include the cost for construction and management of the
anaerobic digester, and does not include the eventual use of
other organic materials such as agricultural refuse or dedi-
cated cultures that are able to increase the biogas yield. The
first point is not insignificant, but in Italy today there are
521 anaerobic digesters used in an agricultural context, and
58 % of these plants use a co-digestion configuration in-
cluding organic materials that are currently suitable [51].
The 78 % of the plants has an installed potency below
1 MW, mainly ranges between 500 and 1,000 kW, showing
a diffusion of the AD technology on farms of no large size
[52]. The set-up configuration proposed in Fig. 3 includes a
digester of approximately 2,500 m3 in volume, and this
could produce 8*105 m3 biogas per year with a value of
approximately 0.30€ per m3, including green incentives and
heat surplus for farm necessities. For the proposed system,
the raw economic return could be approximately 240,000€
per year.
Methane yield (m3/kg COD) Reference
0.23–0.424 Prazeners, 2012 [34]
0.3 Saddoud 2007 [43]
0.236 Goblos 2008 [76]
0.222 Bertin, 2013 [77]
∼0.33 Hassan, 2012 [24]
858
AD in European and Italian Regulations
AD represents a sustainable, natural route for the treat-
ment and recycling of wastes of biological origin, and
of a wide range of useful industrial organic by-products.
In response to steadily increasing biowaste collection,
treatment, and recovery, numerous EC regulations and
guidelines have been issued in this area or are currently
under development. Most of these regulations profound-
ly influence the technological developments and practi-
cal applications of AD. There are many pieces of leg-
islation and regulations that apply to AD depending on
the wastes treated, the types of facilities, and the use of
by-products. As a waste management facility, an AD
site must be run with a license, and some wastes, such
as animal by-products, must be treated with specific
care. The application of digestate on land must respect
limits, and specific regulations also apply for potentially
harmful feedstock, such as sewage sludge. In favour of
AD, the electricity produced with biogas has ensured
that AD is part of the EU policies on new renewable
energy.
The development of AD could be boosted by legislation
as well as good practice and sustainability. The key sets of
EU legislation are:
Fig. 3 Flow diagram for the anaerobic co-digestion of cattle slurry and whey as basic feed products in the dairy production system
Bioenerg. Res. (2013) 6:851–863
1) a Commission Regulation [53], implementing a pre-
vious one of the European Parliament and of the Coun-
cil, which lays down health rules with regard to animal
by-products and derived products that are not intended
for human consumption, and implementing Council
Directive which permits certain samples and items to
be exempt from veterinary checks at the border.
(2) the new European Directive [54] for the promotion of
electricity from RES (renewable energy sources), which
includes the National Renewable Energy Action Plans.
Across the EU, there are many different strategies for
supporting the development and implementation of renew-
able energy, as well as for solving agricultural and environ-
mental problems. These have advanced at different paces in
each country. The Italian body of legislation for the man-
agement of biomass for energy recovery in AD plants is
quite extensive, and consists of several laws. The Italian
Regulation D. Lgs. 387 [55] transposes the European Direc-
tive [56] for the promotion of electricity produced from re-
newable energy sources. It establishes steps for the construc-
tion and management of plants, and in particular, it introduces
a “unique authorisation” for the construction and management
of plants powered from renewable sources to be released by
the region (or the institution delegated for this purpose). In
Bioenerg. Res. (2013) 6:851–863 859

compliance with the provisions of art. 12, paragraph 10 of the
Italian Regulation D. Lgs. 28 [57], “Guidelines for the autho-
risation of plants powered by renewable sources” have been
issued. National guidelines establish the documentation to be
submitted for the construction of plants powered by renewable
sources, depending on the size of the plant. Authorisation
procedures are required for the different types of plants [57].
In 2011, the Italian Regulation [58], which implements
Directive [54] for the promotion of energy from renewable
sources, was issued. It includes and implements the goals set
by Europe, and translates them into concrete action strategies
outlined in the National Action Plan. The decree maintained the
current incentive system for the biogas industry until the end of
2012. The decree makes it clear that SCIA (a certified report
signalling the start of an activity) does not apply to renewable
energy, and introduces changes to the authorisation system
established by the National Guidelines, providing three new
and different procedural processes: (A) a simple communica-
tion to the City Hall, (B) simplified procedure for authorisa-
tion (PAS), and (C) unique authorisation. To present the PAS
with the threshold power that is required, its power is the same
as that specified for the SCIA, even if the regions can extend
this limit up to 1 MW. The decree promotes the efficient use of
biogas from animal slurries and from by-products of agricul-
tural, agrifood, and farming activities, as well as “short chain”
biogas. It also promotes the construction of plants operating in
cogeneration and production by farmers who have biogas
plants to serve their agricultural activities. The definition of
“short chain” is established for the traceability of biomass to
produce electricity. “Short chain” biomass comprises biomass
and biogas produced within 70 km of the production of
electricity (i.e., the distance as the crow flies between the plant
and the administrative boundaries of the municipality contain-
ing the location of biomass production). This definition per-
mits increased incentives.
Italy, with an annual production of 2,891 TWh of elec-
tricity from biogas, ranks third in Europe after Germany and
the United Kingdom. In 2011, there were more than 500
biogas plants nationwide, which was an increase of 13 %
over the previous year and was almost entirely due to
agricultural and livestock biogas.
Destination of Digestate in Relation to Hygienic-Sanitary
Characteristics, Agronomic Quality and Legislation
Digestate, that is, the residual matrix from AD treatment,
can act as an organic amendment and nutrient fertiliser;
however, its application to land represents a health concern
because it may lead to the transmission of pathogens to

Table 6 Legislation standards for digestate

Parameter Reference value Legislation

Escherichia coli or Enterococcaceae ma =1,000 CFU, Mb =5,000 CFU in 1 g, cc =1, nd =5 Reg. EC 142/2011
Salmonella ma =0, Mb =0 in 25 g, cc =0, nd =5
Escherichia coli ma =1,000 CFU, Mb =5,000 CFU in 1 g, cc =1; nd =5 D.Lgs. 75/2010
Salmonella ma =0, Mb = 0 in 25 g, cc =0, nd =5
Lead 140 mg/kg dry matter
Cadmium 1.5 mg/kg dry matter
Nickel 100 mg/kg dry matter
Zinc 500 mg/kg dry matter
Copper 230 mg/kg dry matter
Mercury 1.5 mg/kg dry matter
Hexavalent chromium 0.5 mg/kg dry matter
Salmonella 103 MPN/g dry matter D.Lgs. 99/1992
Cadmium 20 mg/kg dry matter
Mercury 10 mg/kg dry matter
Nickel 300 mg/kg dry matter
Lead 750 mg/kg dry matter
Copper 1,000 mg/kg dry matter
Zinc 2,500 mg/kg dry matter
a
Threshold value for the number of bacteria; the result is considered satisfactory if the number of bacteria in all samples does not exceed m
b
Maximum value for the number of bacteria; the result is considered unsatisfactory if the number of bacteria in one or more samples is M or more
c
Number of samples the bacterial count of which may be between m and M, the samples still being considered acceptable if the bacterial count of
the other samples is m or less
d
Number of samples to be tested
860
humans or animals and introduce them into the environment
[59]. Hygienic risks partly depend on the biowastes that are
treated in a plant [60]. Whey is not well-characterised micro-
biologically, so this substrate can potentially contain pathogens
that may occur in milk, such as Escherichia coli O157:H7,
Listeria monocytogenes and Salmonella spp. Among the
biowastes used in codigestion with whey, animal faeces and
sewage sludge may contain a wide variety of bacteria, parasites
(protozoan and helminths), and viruses [61]. Some pathogens
(e.g., Salmonella and Ascaris eggs) can also be present in mu-
nicipal solid wastes [62]. In crops and silage, pathogenic
enterobacteria such as Salmonella, toxin-producing E. coli, and
L. monocytogenes are considered hazardous [60, 63]. AD is able
to reduce the number of microorganisms that are present in
biowastes, improving safety; however, some pathogens may
survive treatment and thus end up in the residue that is spread
on soil [64]. In particular, the inactivation rates for pathogens
depend on temperature, treatment time, pH, the presence of
volatile fatty acids or ammonia, and nutrient availability in the
reactor [60]. In addition to pathogens, digestate applied as
fertiliser may transport dissolved heavy metals and organic com-
pounds to agricultural fields. Heavy metals are of particular
concern for health risks, eco-toxicity, and environmental accu-
mulation [65]. Milk whey should not contain heavy metals, but,
for example, they can be present in manure because they are
associated with livestock diets and excreted in faeces, and in
sewage sludges with industrial or domestic sources [66].
Digestate can contain organic contaminants according to the
origin of the biowaste; for example, agricultural wastes can
contain pesticides, antibiotics, and other medications, while in-
dustrial organic wastes, sewage sludges and household wastes
can present different contaminants (e.g., pesticides, PAH) [65,
67]. Organic compounds of xenobiotic origin represent a hazard
due to their potential for acute toxicity, mutagenesis, carcinogen-
esis, teratogenesis, and estrogenic effects [68]. The AD process
results in the mineralisation of nitrogen contained in biowaste
and in a lowering of the C/N ratio, thus favouring the short-
term nitrogen fertilisation effect [69]. It has been found that
the N in digestate is almost as effective as the nitrogen in
inorganic fertilisers, thus allowing their reduced application.
In addition, the presence of secondary (Ca2+, Mg2+ and SO42
−
) and trace elements makes digestate a complete and balanced
fertiliser. AD residue can be refined, for example, by solid–
liquid separation, obtaining the liquid fraction, a high efficien-
cy coefficient fertiliser due to the high content of N-NH4 and
the high N/P ratio, and the solid fraction, with amendment
properties related to the ability of the contained organic matter
to maintain the soil humus balance [70]. Digestate resulting
from a biogas plant that uses milk whey with other biowastes,
e.g., manure, activated sludges, agricultural by-products, and
the organic fraction of municipal solid wastes, falls within the
European Regulations on animal by-products and derived
products not intended for human consumption (Regulation
Bioenerg. Res. (2013) 6:851–863
implemented with Commission Regulation 142/2011/EU
[53]) and the Italian law for fertiliser [71]. Moreover, Italian
law on the agricultural use of wastewater digestion sludges [72]
and the European Directive on waste [73], acknowledged in
Italian law D.Lgs.205 [74], must be considered. Commission
Regulation 142/2011/EU [53] sets the following standards for
digestion residues: Escherichia coli or Enterococcaceae for
representative samples of digestion residues taken during or
immediately after transformation at the biogas plant to monitor
the process, and Salmonella for representative samples taken
during or upon withdrawal from storage (Table 6). The Italian
law for fertiliser [71] for fertiliser presents Salmonella and
Escherichia coli as standards for amendments (Table 6). Sal-
monella is also reported by Italian law D.Lgs.99 [72], with a
maximum concentration of 103 MPN/g dry matter in wastewa-
ter digestion sludges. Moreover, these laws indicate the maxi-
mum contents of heavy metals (Table 6). Finally, Italian law
D.Lgs.205 [74] defines “quality digestate” as the product
obtained by the AD of separate collected organic wastes, and
respects the standards and rules that are issued by the Minister
of Environment and Land and Sea Protection with the Minister
of Agricultural, Food, and Forest Policies. These quality criteria
for digestate have not yet been fixed.
The presence of Clostridia in the final sludge seems to be
a relevant point, but limited if the digestate mainly consists
of feed with milk by-products and cattle slurry. Therefore,
generally the environmental impact seems to be highly
decreased compared to the absence of the AD.
Conclusion
Milk and dairy production is a strategic field in the Italian
and European economies, but on the other hand the dairy
sector, worldwide, contributes 4 % to the total global an-
thropogenic greenhouse gas emissions. Moreover, the pro-
duction of milk in the dairy farm represents a hot spot in
several categories under life cycle assessment. Main flows
affecting the results are the methane emissions from enteric
fermentation, as well as nitrogen compound emissions to air
and into the wastewater. In addition, on-site emissions de-
rived from dairy factories are important in terms of acidifi-
cation, eutrophication, global warming, and the formation of
photo-oxidants [75]. Differences were identified in the im-
pact evaluation due to several factors such as the allocation
approach, data sources, characterisation factors, farm man-
agement regimes, and enteric fermentation emission factors.
AD is leading to an improvement in the sustainability of the
full production cycle, reducing such life-cycle assessment
impacts and mainly energy demand and waste disposal, which
are crucial points in eco-efficiency evaluations. The energy
produced by AD can be annually in the order of a few
thousands of MWh as electricity power and some hundreds
Bioenerg. Res. (2013) 6:851–863
of MWh as heat power. The first energy resource is useful for
the internal use of an enterprise, or for sales to the electricity
grid; moreover, it provides the economic value of green cer-
tificates. The second energy as heat can be generally used for
closed building heating, hot water production, and so on. As
well as economic reasons that encourage its valorisation, the
proper management of cheese whey is important due to
stricter legislation that does not permit its disposal on land
without prior treatment.
The AD of whey and cattle slurry or manure could be key
for the future development of industrial districts, particularly
where environment pressures are generally elevated due to
the high demographic density. This is particularly true in the
south of Europe: moreover, in the same area energy is very
expensive due to lack of resources.
On the other hand, management of the reactors could be a
crucial point, and an evolution of the skills needed in the
enterprise is necessary, including expertise on the digestion
process and management, both from a biological and an
engineering point of view. Of course, other new technolo-
gies than AD have been proposed for the same wastes, but
they are still intended for application at an industrial level.
Thus, the investment seems to be not only sustainable but
compatible with a good return for business. Finally, the inclu-
sion, in this case, of only waste materials and only those
locally collected for anaerobic digestion optimise the environ-
mental advantages, showing an ideal path for the application
of biotechnology and an example of application for the
bioenergy community.
Acknowledgments This work is funded in the area of Regional
Development (P.O.R. 2007–2013) carried out by Tecnogranga and
Finpiemonte (Piedmont Region). Acknowledgments are due to the
coordinator of the project DWA s.r.l. and the other organisations
involved: LPA s.r.l. and Cooperativa Frabosa Soprana.
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