Composting a Mixture of Cactus Pear Pruning Waste and Spent Coffee Grounds: The Chemical Evaluation of Organic Fertilizer in Response to Basil Quality and Growth

Response to Reviewer 1 Comments

1. Summary

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files.

Referee 1

Line

Comments

Responses

3

‘high quality’ and horticultural applications

We revised the title of the manuscript in:

Composting a mixture of cactus pear pruning waste and spent coffee grounds: chemical evaluation of organic fertilizer in response to basil quality and growth.

42

The reference4 is not correct

We included: 4. Ochoa, M.J.; Barbera, G. History and economic and agro-ecological importance. In: Crop ecology, cultivation and uses of cactus pear. FAO and ICARDA, Coquimbo, Chile. 2017, 1-11 https://openknowledge.fao.org/server/api/core/bitstreams/c7920e28-b849-40ae-b091-4d40d295a8e9/content

51

The reference 8 is not correct

We included : 8. Rocchetti, G.; Pellizzoni, M.; Montesano, D.; Lucini, L. Italian opuntia ficus-indica cladodes as rich source of bioactive compounds with health-promoting properties. Foods. 2018, 7(2). doi: 10.3390/foods7020024.

57

The reference 11 is not correct

We included : 9. Martins, M.; Ribeiro, M.H.; Almeida. C.M.M. Physicochemical, Nutritional, and Medicinal Properties of Opuntia ficus-indica (L.) Mill. and Its Main Agro-Industrial Use: A Review’. Plants (Basel). 2023, 12 (7), 1512-1557. doi: 10.3390/plants12071512. PMID: 37050137; PMCID: PMC10096643.

the numbering has changed

57

The reference 12 is not correct

we have checked and corrected all the references

63

The reference 16 is not correct

we have checked and corrected all the references

71

The reference 20 is not correct

we have checked and corrected all the references

88

is a reference needed here?

Table reference has been added into the text

89

Inserire nel testo?

On what basis did you determine these ratios? did you take into account the pH contributed by the coffee waste? On the other hand, how would you ensure that the pH would remain stable if the source of coffee waste varied?

The main substrate of the mixture had to remain the opuntia, which has been highlighted in the previous work to be a feedstock to produce a compost with interesting characteristics (e.g. high moisture content for the whole duration of the process, buffer capacity during the active phase of the process and high potassium content). The addition of coffee in the mixture in smaller quantities, up to a maximum of 50% by weight of the opuntia (OFI:SCG - 2:1), was hypothesized to provide nitrogen to the final soil improver, since the content of this element was low in the compost obtained from only opuntia.

The buffering capacity of opuntia is known in literature, therefore the amount of coffee added was not considered important. In fact, during the active phase of the process a constant sub-alkaline pH was observed in all tests with SCG added.

91

In a nutshell, you can present the results of your previous study with OFI without having to refer the reader to another article. This helps the reader to better understand your study.

We integrate the sentence:

‘An additional feedstock of pure OFI (C0) was composted as a reference for comparison for further details

of the results for this compost the reader can refer to previous work carried out in [12]’ as follow:

‘An additional feedstock of pure OFI (C0) was composted as a reference, in our previous work [11], we demonstrated that raw cladodes are an excellent substrate for aerobic fermentation providing a final product of good quality, with characteristics of buffer capacity during the active phase of the process, high moisture content at the end of the curing and high potassium content’.

110

The total quantities loaded for each composter are shown in Error! Reference source

not found..

We inserted the missing information

131

Do you have references of the methodologies used?

We added [27] and [28]:

Total metals in samples were determined according to the procedures described in “Metodi di analisi del compost” - Manuali e linee guida 3/2001 ANPA, ISBN: 88-448-0258-9; the analysis of Cr (VI), performed on the water-leachable chromium fraction, was conducted as described in Bartlett R. J. and James B. R., 1996. Chromium. In: Methods of Soil Analysis. Part 3 – chemical methods (D.L. Sparks. Eds) SSSA Book Series n. SSSA e ASA Inc., Madison, Wi, USA

147

kg / mc; total porosity = 82%) was utilized to prepare five soil samples as reported in Error!

Reference source not found.

Reference has been added.

170

It is more appropriately expressed in “x g”.

Ok we changed the information in 16.500 g

254

Error! Reference source not found. and Er- 254

 ror! Reference source not found.

Reference has been added.

274

In none of the figures and tables did I observe statistical analysis, nor literals showing or not significant differences in the observed means

This is not correct.

Regarding Table 3, the values are expressed as mean of 3 replications in mg/kg dry mass ± their associated expanded uncertainty ue which was estimated in accordance with the Guide to the Expression of Uncertainty in Measurement (GUM, ISO/IEC Guide 98-3:2008) with a coverage factor, K = 2. This corresponds to a level of confidence of about 95%. Expanded uncertainty, defining an interval about the result of a measurement that may be expected to encompass a large fraction of the distribution of values that could reasonably be attributed to the measurand, characterizes the reliability of the shown measurements.

277

no statistical difference? Table 3

See the reply above

280

became a statistician on this?

We deleted: ’Seed germinated and grew without significant morphological differences in soil and

added with compost during the four months of the experiment ’and rewrite:

‘Seed germinated and developed seedlings both in commercial soil (Ctrl) and in those mixed with compost (C0, C1, C2, C3).

281

Error! Reference source 281

not found

Reference has been added.

286-287

is the same text, check.

We modified: ‘Ctrl and 10% OFI compost(C0); Ctrl and 10% OFI compost(C1); Ctrl and 10% OFI compost(C2) and Ctrl and 10% OFI compost(C3)’ in:

Ctrl and 10% OFI compost(C0); Ctrl and 10% compost(C1); Ctrl and 10% compost(C2) and Ctrl and 10% compost(C3).

290

307

222

324

330

Error! Reference source not

found.)

Reference has been added.

313

Have any tests shown significant differences?

Figure 6: in this case we observed the trend of plant growth, and we performed only descriptive statistical analysis

316

Apparently, this table shows better results in Ctrl+0 compost!

With reference to basil morphological plant development (roots length fresh and weight shoots) significant differences were found only for the plants grown on soil mixed to 10% of C3 or C2. Although we did not find significant differences, basil grown in C2 showed a double fresh weight of roots and +38% and +23% of shoot fresh weight and length with respect to those of Ctrl

353

based on what? explain

We modified this sentence: ‘it is speculated as explained in our previous work [13] that the ability of the cladodes to absorb gases limited these emissions.

With this:

“It is speculated, as explained in our previous work [11], that the physical and chemical properties of cladodes increased the metabolic activity of the microbiota involved in composting process, therefore limiting gas emissions. A hypothesis to understand this behaviour may be that the temperature peak remained below 50 °C and the content of proteins of the cladodes is low; both conditions could lead to less ammonia formation and volatilization.”

358

However, you do not explain the pH change by the interaction of the other components. For example, coffee bagasse normally has an acidic pH, as does OFI as you present it. These conditions, plus the temperature, favored the gradual decomposition with the help of the microbiota, until a stable reaction was achieved, resulting in a neutral pH. Under these conditions, it could be discussed which compost will reach neutrality in less time.

Mucilage, which is the main constituent of cladodes, is a gelatinous high-molecular-weight substance. It was demonstrated by different authors  that the OFI mucilage extracts in acidic or basic water showed a reversed bell trend, due to pH-buffering capacity of Opuntia ficus-indica tissues rich in numerous small organic molecules typical of CAM plants, such as organic acids, which bear multiple acid–base groups able to act as buffer components in the physiological pH range (see: Trachtenberg SH, Mayer AM (1981) Phytochemistry 20(12):2665–2668; Medina-Torres L et al, 2000. Food Hydr 14:417–424; Bacchetta et al.2019. J Food Sci Technol 56, 3627–3634 (2019). https://doi.org/10.1007/s13197-019-03794-7).

Furthermore, recent articles (Bernard P et al, 2019  https://doi.org/10.2136/vzj2018.12.0211; Nazari et al. 2021 https://doi.org/10.3389/fpls.2021.798992) suggest that mucilage enables high rhizo-microbial abundance and activity by functioning as carbon and nutrient source offering a protected moist habitat optimized for nutrient mobilization. Moreover, composting microbiota operate best under neutral to acidic conditions. During the initial stages of decomposition, organic acids are mainly formed. The acidic conditions are favourable for microorganisms’ growth and for the breakdown of lignin and cellulose. As composting process proceeds, the organic acids are neutralized contributing to the increment of pH (6 -8) in the mature end-product. The ratio of carbon to nitrogen in the initial material significantly affects pH changes. A high C/N ratio (as in bagasse) can lead to initial acidity as microbes prioritize carbon decomposition.

377

but without statistical differences... or yes?

See reply to statistical note of table 3 (line 274 in this table)

395- 396-397

We modified the sentence:

‘Total N in the soil significantly increases with SCG supplementation while inorganic N (e.g., NO3- and NH4+)

may significantly decrease because of the induced N immobilization [38]? IN:

‘As reported by Hu et al [41], total N in the soil significantly increases with SCG supplementation however we noticed a moderate variation in the C/N ratio by increasing SGC

400-401

Taking into account which results? because not all of them showed statistical differences, or at least they were not indicated.

 We modified in : Our study indicated that the soil supplemented by 10% compost of C2 improved quality basil growth thus sustaining the above-mentioned hypothesis (see line 316 of this table)

473

(This statement is closer to the previous one.) ...that can improve the conditions for better development of a plant like basil.

We modified the following sentence

‘This study demonstrates the potential of using Opuntia Ficus-Indica (OFI) cladodes

and spent coffee grounds (SCG) to produce high-quality compost.’ INTO

This study demonstrates the potential of using Opuntia Ficus Indica (OFI) cladodes and spent coffee grounds (SCG) that can improve the conditions for better development of a plant like basil.

Response to Reviewer 2 Comments

1. Summary

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files.

Referee 2

Line

Comments

Responses

2

Taking into account the biological material that has been used in this experiment, it is better to mention: use a mixture of.......

We revised the title of the manuscript in:

Composting a mixture of cactus pear pruning waste and spent coffee grounds: chemical evaluation of organic fertilizer in response to basil quality and growth.

20

ficus-indica

We checked and modified

31

Spent: lower case

Done

51

delete space after the reference

Done

88

110

1 47

267

281

290

307

322

324

327

330

Error! Reference not found. insert the missing information

We inserted the missing information

90

Why have you not test the SCG as a reference for comparison?

Our previous experience was on Opuntia compost [11] therefore, the aim of our work was to verify if the addition of SCG could ameliorate the quality of compost

Furthermore, from the data obtained from the ultimate analysis, the C/N ratio of spent coffee ground is about 16.5. Values lower than 20 are not suitable for the composting process, therefore a test with only exhausted coffee would not have been possible.

93

Bulking agent: (add abbreviation for the first time - BA)

Done

96

what kind of inoculum? mention the characteristics of the inoculum

Main characteristics of cured compost used as inoculum are reported in Supplementary material (Table S1).

103

It is about quantity of SCG or concentration? Because the proportion of OFI:SCG was 3,3:1 (C2), and 2:1 (C3). Please specify what kind of SCG concentration did you use?

The sentence has been corrected. We are referring to quantities by weight.

109

There are 10 months From November 2021 to August 2022. Maybe is better to mention the number of weeks from April 2022 to August 2022

We modified the date format of the x-axis, but we confirm that the duration of the experimental activities is 9 months from November 2021 to July 2022.

112

Tab 1 Cladodes: lower case

Done

117

this link is not referenced. mention the producer

Done

122

this link is not referenced. mention the producer

Done

125

mention only the type of thermometer

Done

146

H2O: use subscript character

use SI Units. dS m-1

Done

151

-L :without italic character

-mention the varieties of Genovese type. There are at least 4 varieties of Genovese type: Ocimum basilicum ‘Dolly’, O. basilicum ‘Eleonora’, etc.

The Olter Producer did not describe the variety. However, the name ‘Genovese type’ is that reported on DOP regulations.

165

link is not referenced. mention the producer

done

177

use IS units. g∙L-1

done

179

delete space

done

184

TPC: but you use ethanolic extracts

Yes sorry there is a mistake. We corrected in ethanol

done

187

Flavonoids: did you use lyophilized leaves? if so, please  mention

It is reported in paragraph 2.3

194

use IS units. μg∙mL-1

done

253

Fig 1:

-write the month abbreviations in English

- firstly you mentioned 9 months, but appears 10 months in this figure. probably you should mention the number of weeks.

We modified the date format of the x-axis, but we confirm that the duration of the experimental activities is 9 months from November 2021 to July 2022.

254

present the results as shown in figure 2 and figure 3

Figures formats were corrected.

258

FIG 2: firstly you mentioned 9 months, but appears 10 months in this figure. probably you should mention the number of weeks.

We modified the date format of the x-axis, but we confirm that the duration of the experimental activities is 9 months from November 2021 to July 2022.

260

FIG 3: firstly you mentioned 9 months, but appears 10 months in this figure. probably you should mention the number of weeks.

We modified the date format of the x-axis, but we confirm that the duration of the experimental activities is 9 months from November 2021 to July 2022.

273

TAB 3: present these data in the text.

Done

276

(D.lgs. 75/2010) it is not mentioned at the references

We added the reference “Decree, L. Decreto Legislativo n. 75, 29 Aprile 2010: Riordino e revisione della disciplina in materia di fertilizzanti, a norma dell’articolo 13 della legge 7 Luglio 2009 n. 88. Gazzetta Ufficiale della Repubblica Italiana 2010, 106.”

277

Table 3: arrange data in the table

Data was arranged for a better visualization

306

what about the shoots height?

We included shoot height.

325

Between found and among: delete space

Done

10.1±0.9 mgQE/g DW: what kind of variant is this?

Mg quercetin equivalent

347

Opuntia: italic character

Done

360

However, in the composting campaigns herein described, none of these emissions were revealed although they were periodically monitored: insert these data in the supplementary file

We modified the sentence and added data in the supplementary material.

358

what about april? how do you explain the low pH value, below 7?

We consider normal a small variation around neutrality, but we checked the data, and we noticed a type error for the May 5^th. The correct value is 7.1 in Figure 3.

373

present also the data for the other chemical elements in correlation with other literature data, not only Fe, Zn, Mn, K.

We added [36]; Other authors reported that SCG co-composted with cat manure, when applied to soil at 5% (w/w on a wet basis), improved the height of spinach and did not increase the content of Zn, Cu, Pb, and Cd

414

Instead of tallest=smallest

Done

442

in the C3 variant the polyphenol content is lower than in the other two. how do you explain this?

Also discuss data on flavonoids.

Following statistical analysis, the mean values of TPC and flavonoids  are not significantly different among plants grown on soil added with compost.  Furthermore other authors reported contrasting results on SCG applications [42].

Done

445

Full stop after literature

Done

484

Author contributions: re-write according to the authors guidelines

Done

448

References: re-write all the references according to the authors guidelines

Done

Lines

Comments

Responses

146

Position of citations 27 and 28

We changed the position of citations 27 and 28 from 137 to 146 line

268

Replace with Figure 2 and 3

Done

247

But you use only one variant C1. Why are there two values in terms of flavonoids content?

We modified the following sentence:

A similar trend was observed for flavonoid content (Figure 7 B) which were highest in the 344 control plants (22.2± 3.3 mgQE g-1 DW) and a significant difference was found with plants 345 from soil mixed to 10% of C1 (13.0± 2.14, and 10.1±0.9 mgQE g-1 DW respectively).

In:

A similar trend was observed for flavonoid content (Figure 7 B), the control plants showed the highest value (22.2±3.3 mgQE g^-1 DW) which was significantly different from the flavonoid amount of plants grown in soil mixed to 10% of C0 or 10% C3 (13.0±2.14, and 10.1±0.9 mgQE g^-1 DW respectively).

406

change citation order

Done

418

italic character

Done

523

Author contribution: re-write following the authors guideline

Done