Journal of the American Oil Chemists' Society

, Volume 89, Issue 3, pp 529–540 | Cite as

Odorous Compounds in Bioplastics Derived from Bloodmeal

  • Casparus J. R. Verbeek
  • Talia Hicks
  • Alan Langdon
Original Paper

Abstract

During the processing of bloodmeal-based thermoplastics (BMT), volatile compounds are released which are odorous and unpleasant. Literature searches revealed that blood components are thermally unstable under oxidising conditions and the presence of iron in bloodmeal (BM) may catalyse the formation of various odorous compounds. The objective of this work was to establish an odour profile for BM as well as BMT prior to extrusion and to determine the effect of oxidative treatment on the resulting odour profile. A comparison of the volatile compounds arising from using BM with those arising from using red blood cells (RB) was carried out using headspace-solid phase micro-extraction–gas chromatography–mass spectrometry (HS-SPME–GC/MS). A total of 23 compounds were identified, 4 of which were products of putrefaction also identified in RB subjected to putrefactive degradation; these were phenol, 4-methyl phenol, indole and methyl indole. In addition, several aldehydes and ketones were identified in BM and RB and may result from thermal, microbiological or auto-oxidative deterioration of the lipids and proteins present in blood during BM manufacture. Oxidative treatment of BM removed some of the compounds that were generated by putrefaction and thermal degradation. Treatment led to an improvement in the perceived odour type and intensity of BM, however, upon conversion to BMT the perceived odour became significantly worse. This malodour was able to be mitigated by the addition of 10–20 wt% activated carbon or greater than 20 wt% natural zeolite, however some malodour reoccurred after storage.

Keywords

Protein Thermoplastic Degradation Odour 

Introduction

Bioplastics have been of interest since the early 1900s, but production figures have remained low. A wide range of proteins have been trialled as possible bioplastics, such as casein, corn gluten meal, sorghum meal, wheat, zein, egg albumin, feather meal and fish meal [1]. Bloodmeal (BM) can also be used to form thermoplastic materials, representing a significant value added proposition [2]. Large quantities of raw bovine blood are collected annually and contain about 80% water and 18% protein with the balance fats and minerals. For economic and environmental reasons, blood is converted into BM and typically sold as fertilizer.

Fresh whole blood is almost odourless and the same is true if it is dried immediately. Due to the chemical composition of blood and how it is handled, BM has a characteristic odour. There are several stages during which odorous volatile organic compounds could be formed; during raw blood storage, drying, BM storage or as a result of side reactions during bioplastic production. Therefore the use of an odorous raw material results in an odorous product which could negatively impact marketing.

The objective of this paper was to determine the origin of malodour in BM and how thermoplastic conversion affects odour. In addition, strategies for reducing odour were evaluated and included oxidative treatment and physical adsorption. Materials were tested before extrusion (or thermoplastic processing) in order to exclude from the results presented here, thermal degradation effects occurring during processing.

Processes Leading to Odorous Compounds

Degradation of organic compounds is known to occur in both aerobic and anaerobic conditions via biotic and abiotic pathways [3, 4]. The diversity of chemical compounds in organic substances combined with these degradation pathways will lead to many different degradation products. Some of the compounds formed during putrefaction and aerobic degradation are volatile and can be odorous. Some common compounds observed in agricultural operations and their characteristic odours are listed in Table 1.
Table 1

Typical odorous compounds found in meat processing and related processes

Name/formula

Characteristic odour

Odour threshold (ppm)

Present in

Sulphur compounds

 Dimethyl disulphide (CH3)2S2

Decaying vegetables [3]

0.0123 [3]

Meat rendering, meat meal [3]

Swine operation [3]

 Mercaptan CH3SH

Decaying cabbage, potato-like [3]

0.0011[3]

Meat rendering [3], swine operation [3]

Alcohols

 1-Octen-3-ol CH2CH–CH(OH)(CH2)4CH3

Freshly cut grass-like, perfumy, sweet. Mushroom [4]

Processed foods [3], cooked meat [5], mushrooms [4]

 Phenol C6H5OH

Phenolic [3]

0.110 [3]

Swine operation [3]

 4-Methyl phenol CH3C6H5OH

Phenolic

0.00186 [3]

Swine operation [3]

Aldehydes

 Hexanal CH3(CH2)4CHO

Grassy, fruity, sour, sharp, pungent, almond [3, 6]

0.0073–0.0129 [6]

Meat rendering [3], swine operation [3]

 3-Methyl butanal CHOCH2CH2(CH3)–CH3

Fatty

0.00015–0.0023 [7]

Meat rendering, meat meal [3]

Swine operation [3]

 Benzaldehyde C6H5CHO

Almond [3]

0.0417 [3]

Swine operation [3]

Ketones

 Heptanone CH3COCH2CH2CH2 CH2CH3

Dairy, cheese, mushroom [3]

0.14-0.28 [6]

Meat rendering [3], swine operation [3]

 Acetophenone C6H5COCH3

Orange blossom [3]

0.363 [3]

Swine operation [3]

Carboxylic acids

 Propanoic acid CH3CH2COOH

Dairy, sour, rancid [3]

0.03–0.16 [6]

Meat rendering [3], swine operation [3]

 Pentanoic acid CH3(CH2)3COOH

Putrid, sour, sweaty, rancid

0.0048 [6]

Meat rendering [3], swine operation [3]

 3-Methyl butanoic acid CH3CH2(CH3)CH2–COOH

Body odour sour, sweaty feet [6]

0.00036–0.0025 [6]

Putrefaction [8], swine operation [3]

Nitrogen compounds

 Putrescine NH2(CH2)4NH2

Putrid [3]

0.0037 [3]

Putrefaction [8]

 Indole C8H6NH

Faecal [3]

0.000032 [3]

Putrefaction [8], swine operation [3]

 2-Methyl indole C9H8NH

Faecal [3]

0.00056 [3]

Putrefaction [8], swine operation [3]

Furans

 2-Pentyl furan C9H14O

Sweet, bitter, almond like. Green, earthy, bean-like [3]

0.016–0.048 [6]

Meat meal [3], swine operation [3]

Protein Degradation

The decomposition of proteins can occur in both aerobic (decay) and anaerobic (putrefaction) environments [3]. Decay is the process of aerobic decomposition of proteins, where amino acids that were formed via hydrolysis undergo oxidative deamination to form the corresponding carboxylic acids with one less carbon. Putrefaction is the process of anaerobic decomposition of proteins. Amino acids formed via protein hydrolysis are able to undergo reactions within bacterial cells, mediated by intercellular or respiratory enzymes which result in rancid smelling compounds [3].

The rancid and putrid odours associated with degrading blood can be classified into three main groups: amines and nitrogen containing compounds, sulphurous compounds and organic acids.

The products of deamination and decarboxylation of amino acids include amines, mercaptans, sulphides, indole, 2-methyl indole, phenol, 4-methyl phenol, carboxylic acids, carbon dioxide and ammonia and are often found in swine and poultry farms, rendering plants and other agricultural operations dealing with proteinous materials [5, 6, 7, 8]. Amines have been identified in many animal by-product meals, including BM, formed in the raw animal by-product prior to drying by the action of micro-organisms that contain peptidase and decarboxylase enzymes [9, 10]. Amines are infamous for the odour they impart, often described as fishy or putrid.

Sulphur containing compounds such as methyl mercaptan, are also known for their distinct odour, typically described as rotten and are often formed during putrefactive degradation of proteins [7]. Both amines and sulphurous compounds have relatively low odour thresholds and can be sensed at levels that are difficult to detect analytically without pre-concentration prior to analysis.

Lipid Oxidation

Fatty acids, saturated or unsaturated, present in bovine blood are attached to triglycerides, cholesterol esters or phospholipids and comprise part of the cell membrane of blood cells [11]. The two most abundant unsaturated fatty acids found in plasma and red blood cells are oleic and linoleic acid.

Free fatty acids may be formed via deamination and desaturation at the α-β-linkage of peptides linked to a fatty acid. Alternatively, hydrolytic cleavage of the ester bonds between a fatty acid chain and the glycerol or cholesterol to which it is attached in whole blood can also lead to free fatty acids [12]. Hydrolytic cleavage may occur during enzymatic activity, thermally induced hydrolysis or oxidation during drying and storage of whole blood.

Auto-oxidation of unsaturated free fatty acids such as oleic, linoleic and linolenic acid results in the formation of malodorous volatile organic compounds such as C5–C9 ketones [13], benzaldehyde, 2-pentyl furan and C1–C9 aldehydes [14]. In addition, hydroperoxide lyase enzymes are known to produce 1-octen-3-ol from the oxidative cleavage of linoleic acid [15]. Auto-oxidation is initiated by the loss of a hydrogen radical in the presence of trace metals such as iron(III), light or heat [16]. Lipid hydro-peroxides, the primary products of lipid auto-oxidation, are very reactive and undergo a variety of oxidation reactions yielding secondary products such as aliphatic alcohols, aldehydes, ketones and hydrocarbons which are responsible for flavour deterioration in meats [16]. The hydroperoxides are also known to cause oxidative damage to saturated fatty acids at temperatures greater than 60 °C, leading to the formation of methyl ketones such as 2-heptanone, 2-nonanone and 2-decanone [17].

Auto-oxidation is observed during the aging of raw and cooked meat and is initiated by inorganic iron, which is released from haem when haemoglobin is denatured during cooking or through enzymatic activity. Igene et al. [18] found that chelating the free iron with EDTA significantly reduced lipid oxidation.

Sulphite Side-Reactions

Thermoplastic processing of BM requires the addition of excess sodium sulphite [19]. Sulphite ions are known to form sulphite and sulphate radical anions, where the sulphate radicals are capable of interacting with methionine in the presence of transition metal ions and oxygen to form methyl mercaptan [20, 21]. Methyl mercaptan formation is thought to be initiated by sulphite radicals produced from the interaction of sulphite ions with free transition metal ions and oxygen. The sulphite radicals then react with oxygen and other transition metal ions to form the peroxysulphate radical (SO5.) and sulphate radical (SO4.) anions. The sulphate radical anion has an oxidising power similar to that of the hydroxyl radical and may induce the formation of methyl mercaptan in a similar manner to the hydroxyl radical [21]. Sulphite ions and methionine alone do not react to form methyl mercaptan, but in the presence of transition metal ions the formation of methyl mercaptan occurs spontaneously [21].

Odour Mitigation

Many strategies exist for the mitigation of malodours including addition of pleasant smelling compounds or removal of the odorous compounds [22]. The latter can be achieved by adsorption or chemical alteration, such as oxidation.

Oxidation

Oxidation of organic compounds is a known method to reduce malodour. Several examples include the oxidation of odorous compounds in air from rendering, swine and poultry operations using electrically activated oxygen, ozone treatment, incineration and bio-filters [5, 6].

Some of the compounds known to be oxidised to form odourless products include sulphides, disulphides, mercaptans, aldehydes, esters, ammonia, phenols, alcohols, indoles and skatoles [6]. Ozone treatment is an effective means of removing odorous compounds, however, in high humidity conditions it reacts with water to form hydrogen and oxygen, eliminating the ozone before it can react with the odorants [6].

Hydrogen peroxide has been used to oxidise odorous sulphides in wastewater facilities [23], and other common bleaching agents such as sodium chlorite have also been used to manage odour associated with various waste streams [24, 25].

Physical Adsorption

Adsorption is a common technique which has been used to minimise malodour in the home, workplace, industrial waste treatment facilities and for treating other waste emissions. For adsorption to be effective in odour removal, the sorbent must have a high specific surface and a high degree of porosity to maximise transport to adsorption sites.

Odour removal can be achieved using activated alumina, bauxite, activated carbon or forms of zeolite [26]. Zeolites, are aluminosilicate materials which have pores of differing sizes depending on their crystal structure [27]. Natural zeolites are polar sorbents [28] and have been used as heavy metal scavengers, ion exchange media, soil conditioners, ammonium ion scavengers and biofilter odour removers [5, 29, 30, 31, 32]. Activated carbon is a non-polar sorbent and has a specific surface area of 800–1,200 m2 g−1 and 35–40% porosity [28]. Due to its ability to adsorb up to 40% of its own mass, it is often used to remove odorous compounds, particularly hydrogen sulphide, during sewage treatment [33].

Experimental

Materials

Bloodmeal, (agricultural grade) and bovine red blood cells (food grade) were obtained from Taranaki By-Products New Zealand. Sodium dodecyl sulphate, SDS (technical grade) was obtained from Sigma-Aldrich, sodium sulphite, SS (analytical grade) from BDH Lab Supplies, and urea (agricultural grade) from Agrinutrients-Ballance. 37% hydrogen peroxide (technical grade) was sourced from Asia Pacific Specialty Chemicals Ltd, and sodium chlorite (technical grade) from Ajax Finechem. Natural Zeolite (technical grade) was obtained from Blue Pacific Minerals Ltd New Zealand and activated carbon (technical grade) was purchased from Ajax Laboratory Chemicals. Chemicals were used as received, without any purification.

Methods

Thermoplastic Protein Preparation

The patented bloodmeal thermoplastic [19], was prepared by blending 100 parts BM sieved to 710 μm with 3 parts per hundred parts BM (pphBM) SDS, 3 pphBM SS and 10 pphBM urea dissolved in 60 pphBM water heated to 50 °C in a high speed mixer. This mixture was blended for 10 min to ensure a homogeneous powder. The mixture is known as pre-extruded bloodmeal (PBM).

Origin of Odorous Compounds

To determine the source of odorous compounds, BM and BMT prior to extrusion (PBM) were analysed and compared to freshly dried, thermally degraded and biologically degraded red blood cells as well as PBM prepared from dry red blood cells (DRB) (Table 2).
Table 2

Sample preparation for determining the origin of odorous compounds in BM

Sample

Preparation

Bloodmeal (BM)

Used as received

PBM

Thermoplastic protein prior to extrusion

Red blood cells (RB)

Fresh red blood cells

Dry red blood cells (DRB)

500 mL fresh red blood cells oven dried in an open air container at 100 °C for 24 h

PBM prepared from red blood cells (RBPBM)

Oven dried fresh red blood cells were prepared and used instead of bloodmeal according to the PBM process

Thermally degraded red blood cells (RBT)

2.0 g fresh red blood cells were placed inside a sealed 10-mL headspace vial and heated to 120 °C for 1 h

Biologically degraded red blood cells (RBB)

50 mL of fresh red blood cells were placed inside an air-free sealed container without light for 1 week at room temperature

Odour Removal

In order to remove malodours from BM, two oxidative treatment methods were used, as well as physical adsorption. Treated bloodmeal was used to replace the untreated bloodmeal in PBM.

Oxidation

A 100-mL sample of either a 1.0 mol L−1 H2O2 or 0.63 mol L−1 acidified H+/NaClO2 solution was mixed with 50 g BM and stirred for 1 h. After the reaction, the solution was decanted, filtered and washed with 200 mL water followed with oven drying at 80 °C for 24 h.

Adsorption

PBM was prepared as described earlier, after which 5, 10 and 25 pphBM of adsorbent material (natural zeolite or activated carbon) was added and dispersed homogeneously by blending for a further 3 min. The adsorbent was left in the BM to become part of the PBM.

Analysis

Headspace Solid Phase Micro-extraction

Solid phase micro-extraction (SPME) is a rapid, solvent-free extraction technique developed by Pawliszyn and co-workers in 1989. A fused silica fibre coated with a solid sorbent is mounted in a modified syringe to adsorb organic compounds [34]. SPME is an equilibrium method which depends on analyte partitioning between the sample and the SPME fibre [35].

Clear glass 10-mL SPME headspace vials and septa were purchased from Supelco (Sigma Aldrich, Bellefonte, PA, USA). Two grams of powdered sample was thermally equilibrated at 75 °C inside headspace vials for 1 h.

SPME holder and SPME fibres in “Fibre Kit 1 and 4” were purchased from Supelco (Sigma Aldrich, Bellefonte, PA, USA). A 65-μm PDMS/DVB fibre was used for extraction of the volatile compounds from the sample headspace. An extraction time of 5 min was used for the concentration of the volatile compounds from the headspace onto the fibre. All analyses were done in duplicate.

This paper focussed on other volatile organics known to be odorous (Table 1) although the SPME fibre chosen has the ability to extract amines, the amines were not derivatised and would therefore not be detected on the low polarity GC column.

Gas Chromatography–Mass Spectroscopy

GC/MS analysis was performed using an HP 6890 series GC system (Agilent Technologies, Little Falls, DE, USA) on a low polarity Phenomenex™ 30 m × 0.25 mm I.D., 0.25 μm df ZB-5 capillary column (Agilent Technologies, Little Falls, DE, USA). Desorption of the SPME fibre was accomplished by thermal desorption in the back inlet at 250 °C for 2 min, with the fibre set to 3.8 on the vernier gauge on the holder during desorption. The GC/MS run was commenced using a pulsed splitless injection at 15 psi as soon as the fibre was injected to ensure all analytes enter the column simultaneously. A glass inlet liner of 1 mm I.D. was used in the injection port. The oven was held at 40 °C for 5 min and ramped at 8 °C/min to 200 °C and this was held for 15 min. Helium was used as the carrier gas. The HP 5973 mass spectrometric detector was operated in the scan mode (m/z 40–300), the MS Quad operated at 150 °C and the MS source at 230 °C.

Prior to sampling and analysis, a blank run was performed using a sealed empty headspace vial to determine the cumulative effect of volatile compounds arising from the SPME fibre, vial septa, vial, GC inlet septa and GC column during the GC/MS run using the same sampling conditions as described above.

The NIST spectral library was used to aid identification of compounds in the total ion chromatogram. Compounds were identified based on their mass spectrum by visually matching the compound suggested by the NIST library. The compounds were deemed identified when their mass spectrum matched the NIST library spectrum with a greater than 45% probability and they were visually similar. When the probability of a match was less than 45%, the retention time and parent ions were noted, and used to aid detection in other total ion chromatograms. In these cases detection of the parent ion at the expected retention time was used to determine the compound’s presence in the sample.

Results and Discussion

Origin of Odorous Compounds

Bloodmeal Degradation

Three types of degradation reactions leading to formation of odorous compounds have been identified in the literature. These were auto-oxidation of lipids during storage, thermal degradation during drying and bacterial putrefaction. A summary of malodorous compounds identified in BM is presented in Table 3, along with those in:
  • red blood cells (RB) and dried RB (DRB) which are not malodorous

  • RB exposed to high temperature (RBT) to identify compounds that form as a result of thermal degradation and lipid oxidation

  • putrefied RB to identify microbial breakdown products (RBB).

Table 3

Volatile compounds identified from blood products (x, compound identified)

Compound

BM

RB

DRB

RBT

RBB

Compounds found in most samples

 Dimethyl disulphide

x

x

x

x

 

 Hexanal

x

x

x

x

x

 2-Pentyl furan

x

x

x

x

 

Compounds formed as a result of thermal degradation

 2-Heptanone

x

 

x

  

 6-Methyl-2-heptanone

x

 

x

 

 2-Nonanone

x

 

x

x

 

 Nonanal

x

 

x

x

 

 2-Decanone

x

 

x

x

 

Compounds formed as a result of bacterial and/or thermal degradation

 3-Methyl butanoic acid

x

 

x

x

x

 Dimethyl trisulphide

   

x

x

 Dimethyl tetrasulphide

   

x

 

Compounds formed as a result of lipid auto-oxidation

 Heptanal

x

 

x

  

 Benzaldehyde

x

    

 Acetophenone

x

    

 Decanal

x

    

Compounds from the action of putrefactive bacteria

 2-Methyl butanoic acid

    

x

 Phenol

x

   

x

 4-Methyl phenol

x

   

x

 Indole

x

 

x

 

x

 Methyl indole

x

   

x

Unclassified compounds

 1-Octen-3-ol

x

 

x

  

 Tributylamine

  

x

  

 4-Methyl pentanoic acid

x

    

A total of 23 volatile compounds were identified, listed in Table 3. Identification was based on the sample mass spectra matching those found in the NIST spectral library with a minimum of 45% certainty. Due to the nature of the samples being investigated, it cannot be presumed that all possible volatile compounds have been identified. This is a result of investigating a complex sample matrix and additionally, using SPME as a sampling method, the fibre adsorbent produces siloxane artefacts that elute throughout the GC/MS trace and can lead to peak over-lap [36].

Three compounds were identified that appear in most samples; dimethyl disulphide, hexanal and 2-pentyl furan, which may have formed from low temperature (<75 °C) thermal deterioration of lipids and proteins. Compounds which appear to have formed as a direct result of thermal conditions include 2-heptanone, 6-methyl-2-heptanone, 2-nonanone, nonanal and 2-decanone as they are only found in dried blood product samples. These are likely a result of lipid oxidation or thermal degradation at elevated temperatures.

Dimethyl tri- and tetra-sulphide were identified in thermally degraded red blood cells (RBT) but were not found in BM. This suggests that the formation of these two compounds requires thermal conditions which are not reached during the manufacture of BM. 3-Methyl butanoic acid is found in blood samples subjected to thermal or biological deterioration, and may be formed during both of these processes during the manufacture of bloodmeal.

It appears that benzaldehyde, acetophenone and decanal are formed by auto-oxidative degradation of lipids during storage, as they are present only in BM. Heptanal may also be formed via the oxidation of lipids when exposed to elevated temperatures for longer periods of time as it is also observed in dry red blood cells.

Putrefactive breakdown products, such as 2-methyl-butanoic acid, phenol, 4-methyl phenol and methyl indole are found only in bloodmeal and putrefied red blood cells. Indole was additionally found in dried red blood cells and may have been a result of microbial activity prior to drying. Indole and methyl indole (possibly skatole) were tentatively identified at only 25% probability due to the mass spectra of these compounds closely resembling other indole derivatives. However, indole derivatives are a known by-product of putrefactive degradation of proteins and it can be concluded that some putrefaction must have occurred for the compounds to also be detected in BM.

The source of 1-octen-3-ol identified in BM is thought to be formed from the auto-oxidation of unsaturated fatty acids such as linoleic acid. As it was not detected in thermally degraded red blood cells, it can be inferred that 1-octen-3-ol is formed possibly by enzymatic oxidation of linoleic acid in the presence of bacteria or through auto-oxidation during long-term storage of BM. Due to the age of the RB used for this analysis, indole is detected, and possibly 1-octen-3-ol for the same reason, however indole is typically not found in fresh RB [37].

From these results we can conclude that the initial odour of BM may be a result of protein and lipid degradation caused by biological and thermal deterioration of the blood throughout the manufacturing process.

Thermoplastic Bloodmeal Preparation

The effect of bloodmeal production was further explored by a repeat analysis of BM, RB and the corresponding bioplastic formulations. The compounds from BM and PBM, identified between duplicate analysis, are shown in Table 4. A representative total ion chromatogram obtained for BM and PBM is shown in Fig. 1.
Table 4

Volatile compounds identified from blood products (x, compound identified)

Compound

BM

PBM

DRB

RBPBM

Compounds found in most samples

 Dimethyl disulphide

x

x

x

x

 Hexanal

x

x

x

x

 2-Pentyl furan

x

x

x

x

Compounds formed as a result of thermal degradation

 2-Heptanone

x

x

x

x

 6-Methyl-2-heptanone

x

x

x

x

 2-Nonanone

x

x

x

x

 Nonanal

x

x

x

 

 2-Decanone

x

x

x

 

Compounds formed as a result of bacterial and/or thermal degradation

 3-Methyl butanoic acid

x

 

x

x

Compounds formed as a result of lipid auto-oxidation

 Heptanal

x

x

x

 

 Benzaldehyde

x

x

 

x

 Acetophenone

x

x

  

 Decanal

x

x

  

Compounds from the action of putrefactive bacteria

 Phenol

x

x

  

 4-Methyl phenol

x

x

  

 Indole

x

x

x

x

 Methyl indole

x

x

  

Unclassified compounds

 1-Octen-3-ol

x

 

x

 

 Tributylamine

  

x

 

 4-Methyl pentanoic acid

x

   
Fig. 1

Total ion chromatograms obtained for the HS–GC/MS analysis of a blank vial, b bloodmeal (BM), c pre-extruded bloodmeal (PBM), d acidified sodium chlorite treated bloodmeal and e pre-extruded bloodmeal treated with acidified sodium chlorite. NB Peaks have been truncated for clarity. For b and d the baseline has been offset by 50 and 100 × 105 units of abundance respectively. Some of the compounds identified have been numbered on the chromatograms; 1 hexanal, 2 2-heptanone, 3 heptanal, 4 6-methyl-2-heptanone, 5 benzaldehyde, 6 1-octen-3-ol, 7 phenol, 8 2-pentyl furan, 9 acetophenone, 10 4-methyl phenol, 11 2-nonanone, 12 nonanal, 13 2-decanone, 14 decanal, 15 indole and 16 2-methyl indole

No compounds of those identified in BM were detected in the PBM mixture, however, 1-octen-3-ol, 3-methyl butanoic acid and 4-methyl butanoic acid were no longer detected.

The aldehydes and ketones detected are known to be caused by auto-oxidation of free unsaturated fatty acids and are often found in meat industries. Acetophenone, is known to be present in aging meat, and is thought to be formed during thermal treatment via oxidation of fatty acids [38, 39]. Similarly, 2-heptanone and 2-decanone are formed by thermal oxidation of fatty acids.

Three alcohols were identified from BM and RB: 1-octen-3-ol, phenol and 4-methyl phenol. 1-octen-3-ol could be caused by enzyme mediated oxidative cleavage of linoleic acid [33], however, its origin is unknown as it is observed in red blood cells which should have had little enzyme activity while they were stored at low temperature. Phenol and 4-methyl phenol are found only in bloodmeal and are known to occur in the meat industry, caused by putrefaction of the amino acid tyrosine [15].

Two carboxylic acids were identified in BM; 3-methyl butanoic acid and 4-methyl pentanoic acid have been found in swine farms and may result from putrefaction of proteins [39, 40]. However, 3-methyl butanoic acid is also observed in thermally degraded red blood cells, possibly suggesting it may be formed through thermal oxidative processes.

BM was found to contain putrefaction products of tryptophan, namely indole and methyl indole. These putrefactive compounds are also known to occur in meat industries. Methyl indole was absent in RB suggesting that less putrefaction occurred.

Finally, 2-pentyl furan, another compound caused by lipid oxidation and known to occur in meat industries, was found in both bloodmeal and red blood cells which confirms it is caused by lipid oxidation prior to drying.

Due to the large variation in the peak areas for the TIC for BM and PBM, it is difficult to determine what occurs when the bioplastic mixture is prepared [38]. It appears that upon formation of the PBM a significant increase in the overall quantity of volatile odorous compounds (VOCs) occurs, probably due to the denaturing of the proteins and subsequent plasticisation of the random coils causing VOCs trapped in the protein chains to be released.

A significant change in odour type and intensity was observed after the bioplastic mixtures were prepared. Although this is a subjective observation, it can be concluded that the addition of the additive cocktail required for bioplastic conversion plays a major role in the development of the intense unpleasant odour.

Odour Removal

The influence of oxidation on the odour profile of bloodmeal was determined by comparing the HS-SPME–GC/MS results obtained for bloodmeal and PBM with those obtained after treatment.

Hydrogen Peroxide

A summary of the compounds identified after oxidative treatment are given in Table 5. Hydrogen peroxide is a strong oxidizing agent, and is often used to remove odorous compounds. Due to the high abundance of ions in the first 10 min of the GC analysis for H2O2 derived PBM only 2-heptanone could be identified in the early portion of the TIC.
Table 5

Compounds identified from bloodmeal and bloodmeal treatments

Compound

Untreated

H2O2 treated

H+/NaClO2 treated

BM

PBM

BM

PBM

BM

PBM

Alcohols

 1-Octen-3-ol

x

x

    

 Phenol

x

x

    

 Octanol

    

x

x

 4-Methyl phenol

x

x

 

x

x

x

Aldehydes

 Hexanal

x

x

  

x

x

 Heptanal

x

x

 

x

x

x

 Benzaldehyde

x

x

x

x

x

x

 Octanal

    

x

 

 Nonanal

x

x

x

x

x

x

 Decanal

x

x

x

x

x

x

Ketones

 2-Heptanone

 

x

x

x

x

x

 6-Methyl-2-heptanone

x

x

x

x

x

x

 Acetophenone

x

x

x

x

x

x

 2-Nonanone

x

x

x

x

x

x

 2-Decanone

x

x

 

x

  

Carboxylic acids

 Butyl acetate

    

x

x

Nitrogen compounds

 Tetrazole 5-amine

  

x

   

 Indole

x

x

 

x

  

 2-Methyl indole

x

x

 

x

  

Furans

 2-Pentyl furan

x

x

x

x

x

x

 2-n-Octyl furan

   

x

  

Esters

 Butyl butanoate

    

x

x

 Hexyl acetate

    

x

 

 4-Octen-1-yl acetate

    

x

 

 3-Methyl butyl butanoate

    

x

x

 2,3-Dimethyl butyl propanoate

    

x

x

 4-Hex-en-1-yl butanoate

    

x

x

 Hexanoic acid diethylmethyl ester

  

x

   

Treatment with H2O2 lead to the detection of 2-n-octyl furan, presumably by oxidation of fatty acids. After treatment, 1-octen-3-ol, phenol, 4-methyl phenol, indole and methyl indole were no longer observed. However, 4-methyl phenol, indole and methyl indole were present in the resulting PBM indicating that they were oxidised initially, but significant quantities were also trapped in the protein chains, and when plasticised, become volatile.

Hexanal is also removed after H2O2 treatment (its parent ion was no longer detected at the expected retention time), however no organic acids were identified in the sample and heptanal was not removed suggesting that the oxidising agent was consumed before the aldehydes could be oxidised and removal of hexanal has occurred due to its slight miscibility in aqueous solution.

The odour of the treated bloodmeal was found to be much less offensive than untreated bloodmeal due to the removal of putrefaction compounds 4-methyl phenol, indole and methyl indole. However, no major changes in odour characteristics were observed for the PBM prepared from H2O2 treated bloodmeal. These results suggest that the treatment of bloodmeal with H2O2 is effective for oxidising the compounds responsible for bloodmeal’s odour, but has minimal affect on the resulting PBM.

Sodium Chlorite

Oxidation using acidified sodium chlorite yielded the most information about the impact of volatile compounds on the odour profile. A summary of the compounds identified is given in Table 5.

A variety of esters were identified, however, their source was not confirmed. Esters can be formed in the presence of alkyl chlorides derived from alcohols or via the condensation reaction of carboxylic acids with alcohols present in the BM. No organic acids were identified in untreated BM suggesting that carboxylic acids would have to be formed via oxidation of the aldehydes present, in addition to this, alkyl chlorides could have been produced catalysed by the acidic conditions employed from the reaction between alcohols and free chloride ions [40] leading to ester formation.

The only putrefaction product identified from the TIC after treatment was 4-methyl phenol. Other putrefaction products were probably oxidised and removed from bloodmeal. Although the sodium chlorite treated BM appears to have contained larger amounts of VOCs, its odour was found to be less offensive but upon conversion to the corresponding PBM, the odour was similar to that of PBM produced from untreated bloodmeal. Sodium chlorite was the weakest oxidising agent employed for removal of odorous compounds. It was found to cause the formation of significant quantities of aldehydes and even esters, and any carboxylic acids formed may have been consumed during the formation of esters or have been extracted during the removal of the aqueous solution. Despite large increases in aldehydes and esters in the treated BM there appears to be an improvement to the overall odour. Treated BM was virtually odourless so it can be concluded that the aldehydes and esters have little impact on the odour of the treated BM, and consequently that aldehydes probably have minimal contribution to the objectionable odour of BM.

The major contributors to the characteristic odour of BM appear to be the compounds produced via putrefaction of the blood proteins when whole blood is stored prior to drying.

An overall observation regarding chemical treatment is that alcohols and aldehydes can be minimised or removed from bloodmeal by oxidative treatment. In addition, various new compounds were observed, typically esters. All bloodmeal samples regardless of treatment employed were found to contain benzaldehyde, nonanal, decanal, 6-methyl 2-heptanone, acetophenone, 2-nonanone and 2-pentyl furan as shown in Table 5. These compounds are all known to occur from lipid oxidation and may or may not be thermally induced.

Physical Adsorption of Odorous Compounds

Natural Zeolite

The addition of natural zeolite caused a noticeable decrease in the odour intensity only when present at 25 pphBM but the apparent perceived odour type improved at greater than 10 pphBM. At these levels natural zeolite was not an effective means of minimising odour associated with the PBM.

Activated Carbon

The addition of activated carbon dramatically decreased the odour intensity and an improved odour type observed for quantities of 10 pphBM or more. Addition of 10 pphBM activated carbon to improve odour characteristics is a plausible means of improving the PBM’s odour characteristics.

The high specific surface area and porosity of zeolite and activated carbon makes adsorption a possible remedy to the odour of bloodmeal. Despite noticeable changes in the odour of bloodmeal upon addition of adsorbents, the odour returns during storage as the lower molecular mass volatile compounds are displaced by the higher molecular mass compounds.

Conclusions and Recommendations

It is evident that the initial odour associated with bloodmeal is a direct result of the collection and storage of raw blood prior to drying. All compounds identified as a result of odour profiling have all been identified in meat industries and are known to be caused by deterioration of lipids and proteins. Putrefaction products are known to cause malodour, the identification of 3-methyl butanoic acid, 2-methyl butanoic acid, in addition to phenol, 4-methyl phenol, indole and methyl indole are indicative of putrefaction occurring during the storage of whole blood. The inability to detect known odorous compounds such as amines and mercaptans using HS-SPME–GC/MS, is a shortcoming of this technique.

In addition to the formation of putrefaction products during storage, thermal decomposition of lipids also occurred. The results suggested that oxidation of lipids occurred at elevated temperatures leading to the formation of aldehydes, ketones and furans. These compounds are found in bloodmeal, each having a characteristic odour. Removing these compounds changes the odour, but may not actually improve the malodour associated with bloodmeal. This is evident from the GC/MS analysis of RB which despite containing the same ketones, aldehydes and furans that were found in BM, did not have the same objectionable odour.

All bloodmeal samples regardless of treatment employed were found to contain benzaldehyde, nonanal, decanal, 6-methyl 2-heptanone, acetophenone, 2-nonanone and 2-pentyl furan. These compounds may occur from the thermal degradation of proteins or lipids or may be caused by their dissociation from the protein chain. Their persistence after oxidative treatment of BM suggests they are formed thermally and even if removed by oxidation, more will form upon re-heating. Thus their detection by HS-SPME–GC/MS analysis may be an artifact of sample drying and heating involved in this technique.

Although HS-SPME–GC/MS analysis revealed an increase in VOCs in PBM due to plasticisation of the proteins, it alone was unable to determine whether these compounds cause the dramatic change in the odour type and intensity.

BM odour can be minimised by removing the putrefaction products indole and methyl indole using oxidative treatment. After oxidative treatment using sodium chlorite, there was a significant increase in the number of VOCs identified but no corresponding increase in odour intensity. This is further evidence that putrefactive compounds both identified via HS-SPME–GC/MS and any below the detection limits, are the cause of malodour in BM. However, the reason the odour of PBM is more offensive and intense than that of BM is still unknown. A likely cause is reactions involving the sodium sulphite reducing agent used in the PBM process. The formation of sulphur compounds such as mercaptans is a possibility. Incorporation of sorbents appear to be an effective means of minimising the odour. A minimum of 10 pphBM activated carbon appears to be effective.

The major limitation of the investigation of volatile compounds arising from PBM throughout processing includes the inability to detect amines or mercaptans. To determine the causes of the odour from PBM the HS-SPME–GC/MS method should be amended to detect amines and sulphurous compounds. This can be achieved by derivitisation techniques or by applying cryogenic cooling to the GC capillary column and possibly the employment of a polar column. In addition, the VOCs identified should be quantified by use of an internal standard. Quantification can be aided by cryogenic cooling and the use of a narrow bore inlet liner allowing improved chromatographic resolution, as the eluted peaks would be sharp and symmetrical, easing integration.

Prior to the processing of PBM the inorganic iron present upon denaturing of the blood proteins should be removed or chelated to minimise catalytic reactions that may occur upon addition of the reducing agent sodium sulphite. Conversely, the method should be adapted to use less sodium sulphite or a different reducing agent to determine the effect on overall odour.

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Copyright information

© AOCS 2011

Authors and Affiliations

  • Casparus J. R. Verbeek
    • 1
  • Talia Hicks
    • 1
  • Alan Langdon
    • 1
  1. 1.School of EngineeringUniversity of WaikatoHamiltonNew Zealand

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