Bioremediation of olive-mill wastewaters by composting

wmr p054 30-10-95 13:50:51 W aste M anagement & R esearch (1995) 13, 509–518 BIOR EM ED IATION OF OLIVE-M ILL WASTEWATER S BY COM POSTIN G U. Tomati, E. Galli, L. Pasetti and E. Volterra I.B.E.V. N ational Council of R esearch, A rea della R icerca di R oma, Via S alaria Km 29.300, 00016 M onterotondo S calo ( R oma) Italy (R eceived 8 June 1994, accepted in revised form 11 October 1994) Olive-mill wastewaters (O.M .W.) containing about 7% solids were composted with wheat straw in a forced aeration static pile. Two percent urea was added to ensure a C/N ratio of about 35. To avoid overdosing with water, a fraction of the O.M .W. equal to the weight of the straw was added initially. When composting had reached the thermophilic phase, additional doses of O.M .W. were added every 3 days as water evaporated. The ratio of O.M .W. solids:straw approached 1:1 and the thermophilic phase was extended to 35 days. Temperature, oxygen consumption, pH , C/N , total organic carbon, total extractable carbon, humic and fulvic acids and lignin degradation were followed during the process. The humification was assayed following the degree of humification, the humification rate and the humification index which respectively reached the values of 78%, 37.8% and 0.28 after 2 months. H umic acids were characterized by their elemental composition and molecular weight. A lignin degradation of about 70% was assayed at the end of the thermophilic phase. N o phytotoxicity was recorded on the end product, the chemical and physical properties of which suggest its possible use as fertilizer.  1995 ISWA K ey Words—Olive-mill wastewaters, wheat straw, compost, lignin, phytotoxicity, C/N , water removal, index, rate and degree of humification. 1. Introduction In the M editerranean areas, olive mills generate a stream of liquid waste which is roughly estimated at about 1.2 million tonnes year −1. The waste from olive-mill pressing (O.M .W.) is usually considered to be highly polluting, as it is produced in a very large amount during a brief rainy season, it rapidly undergoes biodegradation and it is toxic to plant and soil micro-organisms when applied to agricultural land in significant quantities (Zucconi & Bukovac 1969; Jelmini et al. 1976; Perez et al. 1986; Paredes et al. 1987). Therefore, in the European U nion countries, it is illegal to dispose of O.M .W. directly into the environment, especially into surface waters, and there are also concerns that the use of unremediated O.M .W. as fertilizer will lead to contamination of soil and groundwaters. Looking at O.M .W. from the point of view of their organic matter content and nutrients for plants, they constitute a valuable resource as a fertilizer and a source of organic compounds (F iestas R os de U rsinos 1959; Lanzani & F edeli 1986; Catalano & D e F elice 1989; Tomati et al. 1990). The two aspects mentioned above lead to cleaning or recycling techniques. R ecycling, rather than purification, is the most suitable solution because cleaning waters produces sludge which must then be disposed of. R esearch aimed to evaluate the possibility of using O.M .W. as a source for biomass production has mainly been performed in Spain, Italy and G reece. 0734–242X/95/060509+10 $12.00/0  1995 ISWA 510 U. Tomati et al. TABLE 1 Chemical and physical characteristics of olive-mill wastewaters pH Water Organic matter K jeldahl N itrogen Polyphenols, tannins, gums, pectins M ineral salts (550°C) D ry matter (105°C) 5.4 93.2% 6.0% 0.3% 0.2% 0.4% 6.8% Olive-mill wastewaters were proposed as a source to produce enzymes for food industries (Petruccioli et al. 1986), yeast for feed (M ontedoro et al. 1988), hormonelike substances for agriculture (H ernàndez & M endoza 1976; Tomati et al. 1990) and edible mushroom mycelia (G alli et al. 1988; Satta et al. 1989). H owever, these technologies are of theoretical rather than of practical interest. To recover organic matter, composting is a technique of ever-increasing interest, and a very suitable way to solve the problem posed by O.M .W., also considering the enormous quantity of wastewaters to transform. Composting experiments of O.M .W. mixed with husks or alone have been performed by Calvet et al. (1985), Estaùn & Calvet (1985), Amirante & D i R ienzo (1991) and Amirante et al. (1991), who obtained a safe end-product with a high fertilizing value. H owever, not enough data are available to evaluate the process performance and product stabilization. This study aims to elucidate the evolution of some parameters dealing with the stabilization and humification of O.M .W. mixed with wheat straw during the composting process, and to provide evidence on the possibility of employing a composting process to obtain high quality fertilizers. 2. Materials and methods Composting was performed in a forced aereation static pile. The air required for the process was provided by a blower (0.5 m 3 min −1) connected to a series of perforated pipes laid on a slightly sloping concrete slab. About 200 kg of chopped straw were placed over the pipes to form a pile (2.3×2.0×1.2 m) and 200 l of O.M .W. (Table 1) were then supplied. To ensure a C/N ratio between 35–40, 2% urea was added. Temperature was monitored using a system of thermistors placed in the different horizons of the pile, connected to a temperature controller set with an adjustable temperature set point. When the temperature rose over the set point (50°C), the controller activated the blower until the pile temperature decreased. The thermophilic phase was prolonged for 35 days adding 200 l of O.M .W. every 3 days. A ratio of about 10 l of O.M .W. to 1 kg of straw was reached after 35 days. At this stage, the mass was turned and submitted to the maturation phase. Air injection was stopped as temperature decreased below 40°C. Analyses were performed during the process and on the end-product. Total organic carbon (TOC), total extractable carbon (TEC), nitrogen, pH , conductivity, maximum water capacity, extraction of humic and fulvic acids and humin were performed according to H esse (1971). Phytotoxicity was assayed by the L epidium sativum test Bioremediation of olive-mill wastewaters 511 according to Zucconi et al. (1981). Oxygen consumption was assayed at 28°C (oxygen monitor YSI mod.240/B) on a 3 ml sample of a suspension obtained by stirring for 30 min 5 g fresh weight of compost in 50 ml of 0.9% N aCl solution. Elements were determined, after mineralization, by atomic absorption spectroscopy with a Perkin Elmer mod.380 Spectrophotometer. Lignin was determined by a modified K lason procedure (M oore & Johnson 1967). Elemental composition of humic acids was assayed by a C/H /N Analyser Perkin Elmer mod.240/B; −COOH and −OH groups were determined according to H esse (1971). The characterization of molecular weight of humic acids was carried out by gel filtration on Sephadex G -150 (fine grade Pharmacia, column: 50×1.4 cm; flow rate 8.0 ml h −1). Samples of about 1 mg of organic carbon were applied and eluted with N a phosphate buffer (0.1 M , pH 7.5). F ractions of 1 ml were collected and their absorbances were measured at 280 nm. M olecular weights were compared on a calibration curve performed using Blue D extran, R ibonuclease A, Ovalbumin, Albumin, Aldolase and Catalase as reference substances. 3. Results and discussion Olive-mill wastewaters (Table 1) were composted with chopped dry wheat straw, the disposal of which poses environmental problems in many countries. Straw recycling has been studied to devise low-cost technologies for upgrading the value of such waste. D ue to the absence of polluting compounds and its high absorbing capacity, straw can be favourably considered a “clean” solid substrate for O.M .W. composting to obtain a good fertilizer and a soil conditioner. Olive-mill wastewaters consist in a great volume of water in which about 6–10% of organic matter and salts are suspended or dissolved. When applied to such waste, rational composting has to face the necessity of eliminating most of the liquid component. This latter point has been solved by prolonging the thermophilic phase. The addition of O.M .W. every 3 days maintained adequate moisture in the mixture. This treatment enables the process to reach important goals such as elimination of phytotoxicity, high degradation of lignin and great evaporation of water. In fact, about 70% of lignin was metabolized during the thermophilic phase, whereas only a negligible degradation occurred during the compost maturation (Table 2). Water evaporation was also rather efficient. On average, about 20 l 100 kg−1 of straw (dry weight) were eliminated every day. The phytotoxicity, measured on the end-product, was absent, as indicated by the germination index of 75% (Table 3) obtained diluting to 30% of the liquid fraction extracted by compost (Zucconi et al. 1981). Phytotoxins are usually produced during the decomposition of non-stabilized organic matter. H owever, they are destroyed during the thermophilic phase (Zucconi & D e Bertoldi 1986). F igure 1 shows the temperature and oxygen consumption rate profiles during the first 60 days of the composting process. As the O.M .W.-wheat straw mixture was prepared, mesophilic microflora started to multiply within the composting mass; the respiration rate rapidly increased in the first 10 days, reaching a maximum of about 110 mmol O 2 h −1 kg−1 (dry weight) and then slowly decreased to about 48 mmol O 2 h −1 kg−1 (dry weight). As a consequence of hydrolitic activities, rapid warming occurred and the temperature rose to 50°C in 5 days. The temperature was then lowered by injecting air, using the control unit. The temperature remained at that value for several days. As a consequence, microbial populations were selected favouring the 512 U. Tomati et al. TABLE 2 Organic matter evolution during the composting process TOC TEC HA FA NH D ays % d.w. % d.w. % d.w. % d.w. % d.w. C/N DH % HR % HI Lignin degradation % 0 6 12 23 35 63 90 110 140 35.4 — — — 28.2 12.6 — 11.8 11.5 — — — — 75.5 78.7 79.4 81.6 78.2 — — — — 25.9 37.8 42.5 42.7 41.6 — — — — 0.32 0.27 0.25 0.22 0.28 0 — — 70 70 — 70 — — 45.2 44.6 42.8 41.2 40.4 35.2 36.2 36.3 35.3 n.d. — — — 13.9 16.9 19.4 19.0 18.8 n.d. — — — 6.6 8.9 10.0 10.0 10.0 n.d. — — — 3.9 4.4 5.4 5.5 4.7 n.d. — — — 3.4 3.6 4.0 3.5 4.1 TOC, total organic carbon; TEC, total extractable carbon; H A, H umic acid; FA, F ulvic acid; N H , humin; D H , degree of humification; H R , humification rate; H I, humification index; d.w., dry weight; n.d., not detectable. The end of the thermophilic phase occurred after 35 days, when the addition of olive-mill wastewaters was stopped. TABLE 3 Chemical composition and physical properties of olive-mill wastewaters–wheat straw mixture after 140 days (end-product). D ata are the average of 4 different composting cycles H umidity K jeldhal N itrogen P 2O 5 K 2O M gO CaO Fe Zn Mn B Al Cu Cd Pb Hg Specific weight M aximum water capacity Electrical conductivity G ermination index 37.3% 3.1% d.w. 1.4% d.w. 2.1% d.w. 1.3% d.w. 1.9% d.w. 0.5% d.w. 0.02% d.w. 0.02% d.w. 1.50% d.w. 0.20% d.w. < 1 ppm < 1 ppm < 1 ppm < 1 ppm 0.345 kg dm −3 195% 9.56 mS cm −1 75% d.w., dry weight. thermophilic microflora; besides, temperature acted positively in suppressing phytotoxicity (Zucconi & D e Bertoldi 1986). Transition from the thermophilic to the maturation phase was characterized by a decrease of moisture in the mixture (F ig. 2). At the beginning of the mesophilic stage, pH rapidly increased from 6.6 to 9.0 (F ig. 2). After 5 days, a drop was observed, probably due to breakdown of the carboneous 513 60 120 50 100 40 80 30 60 20 40 10 20 0 10 20 30 40 50 mmol O2 h–1 kg–1 dry weight Temperature (°C) Bioremediation of olive-mill wastewaters 0 60 Composting (days) F ig. 1. Temperature and oxygen consumption rate profiles during the first 60 days of the composting process of olive-mill wastewaters–wheat straw mixture. The arrow indicates the last addition of O.M .W. Χ—Χ Windrow temperature at 50 cm depth from the top; Χ---Χ Air temperature; Β—Β Oxygen consumption. substrates into acid intermediates (Zucconi & D e Bertoldi 1986). Then pH slowly increased reaching a value of 8.8 after 20 days. The alkaline hydrolysis of K and N a salts, continuously added through O.M .W., can explain this and could be responsible for the loss of nitrogen (about 20%). Salt concentration is also responsible for the increase in electrical conductivity assayed on the end-product (Table 3). At the beginning of the experiment, the water content of the mixture was about 80%. This high moisture was maintained throughout the process by the continuous addition of O.M .W. which balanced the evaporation. After 35 days, when addition of O.M .W. was stopped, moisture rapidly decreased and the end of the thermophilic phase occurred (F ig. 2). A rapid consumption of the organic matter, probably caused by respiration during the thermophilic phase, is apparent from the decrease of total organic carbon (TOC), shown in Table 2. Contemporarily, humification processes occurred, as shown by the data of extractable carboneous fractions (TEC) (Table 2), the production of which continued during the maturation phase. Assessing the extent of the stabilization is a difficult task, considering the incomplete understanding of the parameters which may indicate the degree of humification. N o univocal parameter is yet available to assess acceptable levels of compost stability and humification. N owadays, the most reliable tests are those based on the separation of labile organic matter (non-humified) from stabilized organic matter (humic and fulvic acids) (Sequi et al. 1986; D e N obili & Petrussi 1988; Saviozzi et al. 1988). The first 514 U. Tomati et al. 90 10 9 70 pH Moisture content (%) 80 60 8 50 7 40 30 20 40 60 80 100 120 140 6 Composting (days) F ig. 2. M oisture content and pH of the windrow during the composting process. Χ—Χ M oisture content; Β—Β pH . parameter proposed was the humification index (H I), i.e. the ratio between non-humified (N H ) and humified fractions (H A, humic acids; FA, fulvic acids): H I= NH H A+FA [1] It reaches values less than 0.5 for humified substrates (Sequi et al. 1986). M ore recently, two new parameters to evaluate the humification level of composts have been proposed (Ciavatta et al. 1988)—the degree of humification (D H ) and the humification rate (H R ): H A+FA ×100 TEC [2] H A+FA ×100 TOC [3] D H (%)= H R (%)= which tend to increase as humification proceeds. The efficiency of the humification process is clearly shown in Table 2 where humification degree, humification rate and humification index are reported. Values of 75.5%, 25.9% and 0.32 are respectively reached at the end of the thermophilic phase. A further evolution of the humification process is indicated by the values reached during the maturation. The evolution of the C/N ratio is also reported (Table 2). Starting from 515 Bioremediation of olive-mill wastewaters TABLE 4 Characteristics of humic acids (dry ash-free basis) % D ry weight D ays 35 63 90 110 meq g−1 Atomic ratio C N H O S H /C O/C C/N 53.87 51.16 53.53 52.26 3.93 4.08 4.15 4.10 6.14 5.27 5.58 5.35 34.66 38.62 36.04 37.68 1.40 0.87 0.70 0.61 1.37 1.24 1.25 1.23 0.48 0.57 0.50 0.54 15.99 14.63 15.05 14.87 − COOH 3.42 4.06 4.02 4.55 − OH 1.40 1.45 1.69 0.57 an initial C/N value of about 35.6, a value of 28.2 was obtained at the end of the thermophilic phase. At the end of the maturation phase, the C/N ratio reached a value of 11.5. The chemical characteristics of humic acids (Table 4) confirm the evolution of humification, and permit qualitative estimation of the degree of compost maturity. The increase of the ratio between oxygen and carbon shows that the process evolved towards oxidation. The high level of humification reached during the process was clear from the comparison between the various molecular weights of humic acids performed at different composting times (0, 35, 63, 90 and 140 days). Elution profiles (Sephadex G -150) of the humic fractions showed evolution towards high molecular weights as the process proceeded (F ig. 3). The humification of lignocellulosic wastes depends on lignin hydrolisis which mainly occurs during the thermophilic phase. Afterwards, humic substances are produced by the condensation and polymerization of aromatic units and cellular debris such as sugars, amino acids etc. As a consequence of the microbial synthesis, polymerization occurred during the maturation phase, as shown by the elution profile at 140 days, which indicated that about 60% of polymers had a molecular weight greater than 200 kD a. To gather information on the quality and agronomic value of the thus obtained endproduct, analyses on the nutrient content, heavy metals and physical properties were performed (Table 3). R esults revealed considerable amounts of macronutrients, particularly nitrogen, which reached a concentration of about 3%. The absence of heavy metals confirms the “safety” of the starting materials, which can be taken into consideration for composting in an environmental protection policy. Besides, on the basis of specific weight and maximum water capacity, use as a turf substitute could be proposed. Although the biological properties (i.e. microbial population, microbial metabolites) were not investigated, the high level of humification leads us to suppose that compost can benefit soil and plant nutrition through the humus properties. In general, an increase in the maximum water capacity and the ion-exchange capacity, together with an improvement in soil fertility were recorded. Besides, N 2-fixers, ammonia producing bacteria, nitrifying bacteria, cellulosic and lignolitic micro-organisms increased when compost was supplied to landfill. Similar results have also been obtained when O.M .W. undergo biological oxidation directly in the soil. Improvement in chemical, physical and biological properties clearly appear when soil activities eliminate the harmful effects linked to O.M .W. characteristics and doses (Paredes et al. 1987; F louri et al. 1990). 516 U. Tomati et al. Molecular weight (kDa) 1.2 10 3 2 10 10 Molecular weight (kDa) 3 1 1.2 10 2 10 10 35 days O.D. at 280 nm O.D. at 280 nm 0 days 0.8 0.4 0 40 80 120 0.8 0.4 0 40 ml 80 1.2 90 days O.D. at 280 nm O.D. at 280 nm 63 days 0.8 0.4 40 80 120 0.8 0.4 0 ml 40 80 120 ml 140 days 1.2 O.D. at 280 nm 120 ml 1.2 0 1 0.8 0.4 0 40 80 120 ml F ig. 3. Elution profiles of humic acids extracted from olive-mill wastewaters–wheat straw mixture after 0, 35, 63, 90 and 140 days from the beginning of the composting process (G el filtration on Sephadex G -150; column: 50×1.4 cm; flow rate: 8 ml h −1). 4. Conclusions Olive-mill wastewaters can be usefully composted with other agricultural wastes which have a high absorbing capacity. To eliminate most of the liquid component of O.M .W., Bioremediation of olive-mill wastewaters 517 it has been necessary to prolong the thermophilic phase. This treatment also permits consistent degradation of lignin compounds present in the wheat straw and suppression of phytotoxicity. The values of C/N , H I, D H and H R , together with the evolution of humic acids towards high molecular weights, show the high level of humification reached at the end of the process. The characteristics of the end-product suggest that the thus obtained composted material can be suitably used as fertilizer. Composting O.M .W. with lignocellulose agricultural wastes is proposed as an effective way to solve the problem posed by olive-mill wastes. Acknowledgments The authors wish to thank M r. G razio Palma for his assistance. R esearch was carried out in the framework of EC contract N o. EVWA-CT 92-0006. References Amirante, P. & D i R ienzo, G . C. (1991) Tecnologias e instalaciones para la depuracion de alpechines por medio de procesos fisicos de concentracion y osmosis. (Physico-chemical treatment for O.M .W. cleaning.) In R eunion International S obre: Tratamiento de A lpechines (Junta de Andalucia, ed). Sevilla, Spain: PAO Suministros graficos, S.A., pp. 47–62. Amirante, P., Brunetti, G ., Senesi, N ., D i R ienzo, G . C. & M iano, T. M . (1991) Il compostaggio di concentrati di reflui oleari. Prove sperimentali in un impianto pilota statico ad asse verticale. (Olive-mill wastewater composting. Experiments in a pilot static vertical plant.) In R iciclo di Biomasse di R ifiuto e di S carto e Fertilizzazione Organica del S uolo (N . Senesi & T. M . M iano, eds). Bologna, Italy: Patron Editore, pp. 231–234. Calvet, C., Pagés, M . & Estaùn, V. (1985) Composting of olive marc. In Composts as H orticultural S ubstrates (O. Verdonck, ed). Belgium: G hent/M elle, 172, 255–259. Catalano, M . & D e F elice, M . (1989) U tilizzazione delle acque reflue come fertilizzante. (U tilization of waste water as fertilizer.) In S eminario Internazionale sul Trattamento Delle A cque R eflue Degli Oleifici. Lecce (Italia). Ciavatta, C., Vittori Antisari, L. & Sequi, P. (1988) Caratteristiche dei principali concimi a base di cuoio torrefatto disponibili in Italia. (Characteristics of the main fertilizers by torrefied leather in Italy.) A gricoltura M editerranea, 119, 67–73. D e N obili, M . & Petrussi, F. (1988) H umification index (H I) as evaluation of the stabilization degree during composting. Journal Fermentation Technology, 66, 577–582. Estaùn, V. & Calvet, C. (1985) Chemical determination of fatty acids, organic acids and phenols during olive marc composting processes. In T he use of composts as horticultural substrates (O. Verdonck, ed). Belgium: G hent/M elle, 172, 263–270. F iestas R os de U rsinos, J. A. (1959) Estudio del alpechin para su aprovechamiento industrial. Separaciòn de algunos de sus componentes y identificaciòn de los acidos organicos para cromatografia de particiòn. (Study on olive-mill wastewater for its industrial use. Separation of some components and organic acid determination by chromatography.) Grasa y A ceites, 10, 30–34. F luori, F., Chatjipavdlidis, I., Balis, K ., Servis, D . & Jerakis, C. (1990) Effect of olive oil mills liquid wastes on soil fertility. In R eunion International sobre: Tratamiento de alpechines (Junta de Andalucia, ed). Sevilla, Spain: PAO Suministros graficos, S.A., pp. 85–101. G alli, E., Tomati, U ., G rappelli, A. & Buffone, R . (1988) R ecycle of olive oil waste waters for Pleurotus mycelium production in submerged culture. A grochimica 32, 451–456. H ernàndez, E. & M endoza, D . (1976) Producciòn de àcido giberélico por Gibberella fujikuroi en substratos que contienen pulpa de aceituna, aceite de oliva o subproductos de la extracciòn de este ultimo. (G ibberellic acid production by Gibberella fujikuroi grown on olive coproducts.) R evista de A groquimica y Tecnologia de A limientos 16, 357–366. H esse, P. R . (ed) (1971) A Tex tbook of S oil Chemical A nalysis. London, U .K .: W. Clowes and Sons Ltd. Jelmini, M ., Sanna, M . & Pelosi, N . (1976) Indagine sulle acque di rifiuto degli stabilimenti di 518 U. Tomati et al. produzione olearia in provincia di R oma: possibilità di depurazione. (Olive-mill wastewaters in R ome M unicipality: cleaning possibilities.) Industrie A limentari 15, 11 (133), 123–131. Lanzani, A. & F edeli, E. (1986) Composizione e utilizzazione delle acque di vegetazione. (Composition and utilization of olive-mill wastewaters.) In Tavola R otonda: L o S maltimento Delle A cque R eflue dei Frantoi (Accademia N azionale dell’olivo, ed). Spoleto, Italy: Arti grafiche Panetto & Petrelli, pp. 15–28. M ontedoro, G . F., F ederici, F., Servili, M . & Petruccioli, M . (1988) U tilizzazione delle acque reflue. (Olive-mill wastewater utilization.) In II Tavola R otonda: A cque R eflue dei Frantoi Oleari (Accademia N azionale dell’olivo, ed). Spoleto, Italy: Arti G rafiche Panetto & Petrelli, pp. 23–40. M oore, W. E. & Johnson, D . B. (1967) Procedures for the chemical analysis of wood and wood products. F orest Products Laboratory, U .S. D epartment of Agriculture, M adison. Wisc., U .S.A. Paredes, M . J., M oreno, E., R amos-Cormenzana, A & M artinez, J. (1987) Characteristics of soil after pollution with waste waters from olive oil extraction plants. Chemosphere, 16, 1557–1564. Perez, D ., Esteban, E., G omez, M . & G allardo-Lara, M . (1986) Effects of waste waters from olive processing on seed germination and early plant growth on different vegetable species. Journal Environmental S cience and H ealth, 21, 349–357. Petruccioli, M ., Servili, M ., M ontedoro, G . F. & F ederici, F. (1986) Produzione di pectinasi da parte di Cryptococcus albidus var. albidus sviluppato su substrato costituito da acque di vegetazione dei frantoi tal quali o arricchite di residui della coltivazione del girasole. (Pectinase production by Cryptococcus albidus var. albidus grown on olive-mill wastewater alone or with sunflower residues.) in Tavola R otonda: L o S maltimento Delle A cque R eflue Dei Frantoi (Accademia N azionale dell’olivo). Spoleto, Italy: Arti G rafiche Panetto & Petrelli, pp. 79–85. Satta, G ., Amat di Sanfilippo, P., Sanjust, E., Pompei, R . & R inaldi, A. (1989) Procedimento biologico per la detossificazione delle acque di vegetazione effluenti da frantoi oleari. (Biological process for olive-mill wastewater cleaning.) Brevetto CN R 48230A 89, Italy. Saviozzi, A., Levi-M inzi, R . & R iffaldi, R . (1988) M aturity evaluation of organic waste. BioCycle 198(3), 54–56. Sequi, P., D e N obili, M ., Leita, L. & Cercignani, G . (1986) A new index of humification. A grochimica 30, 175–179. Tomati, U ., D i Lena, G ., G alli, E., G rappelli, A. & Buffone, R . (1990) Indolacetic acid production from olive oil waste water by A rthrobacter sp. A grochimica 34, 223–228. Zucconi, F. & Bukovac, N . J. (1969) Analisi sull’attività biologica delle acque di vegetazione delle olive. (Biological activities of olive-mill wastewaters.) R ivista dell’Ortoflorofrutticoltura Italiana 53, 443–461. Zucconi, F., F ortc, M ., M onaco, A. & D e Bertoldi, M . (1981) Biological evaluation of compost maturity. BioCycle 22(4), 27–29. Zucconi, F. & D e Bertoldi, M . (1986) Compost specifications for the production and characterization of compost from municipal solid waste. In Compost: Production, Quality and Use (M . D e Bertoldi, M . P. F erranti, P. L’H ermite & F. Zucconi, eds). London, U .K .: Elsevier Applied Science, pp. 30–50.